@article {9119, title = {Asrij/OCIAD1 expression delineates functionally distinct hematopoietic stem cells in the bone marrow. [Next Generation Sequencing facility]}, journal = {Exp Hematol}, year = {2025}, month = {2025 Oct 01}, pages = {105266}, abstract = {

Hematopoietic stem cells (HSCs) within the bone marrow (BM) display significant molecular and functional heterogeneity. Deciphering intrinsic factors that govern HSC diversity is key to enriching specific HSC subtypes for predictable and clinically relevant differentiation outcomes. Here, we show that the mitochondrial protein Asrij/OCIAD1, a conserved regulator of hematopoietic homeostasis, contributes to HSC heterogeneity. Asrij depletion is known to cause loss of quiescence, myeloid bias and aging-like changes in mouse BM HSCs. Interestingly, Asrij expression is inherently heterogeneous and enriched in only 47\% of the HSC population. To investigate whether Asrij expression levels influence HSC fate, we generated a novel Asrij-mNeonGreen knock-in reporter mouse using CRISPR-Cas9 technology. We show that the Asrij reporter faithfully recapitulates its heterogeneous expression in the BM HSCs, allowing isolation of live cells based on Asrij expression levels. Ex-vivo culture of HSCs demonstrated that Asrij HSCs exhibit enhanced self-renewal capacity, whereas Asrij HSCs are primed for differentiation. Transplantation assays further revealed that Asrij HSCs have enhanced reconstitution in the BM hematopoietic stem and progenitor cell (HSPC) and myeloid cell compartments. Transcriptomic analysis uncovered signatures of quiescence in Asrij HSCs, while Asrij HSCs exhibit hallmarks of HSC activation. In summary, we show that Asrij levels impact the quiescence, self-renewal, and differentiation potential of HSCs, thereby contributing to the functional diversity of the HSC pool. Further, the Asrij-mNeonGreen reporter mouse provides a powerful and versatile model for investigating the molecular underpinnings of functional diversity within the HSC compartment. TEASER ABSTRACT: Our study addresses the complexities of hematopoietic stem cell (HSC) heterogeneity and uncovers novel determinants that govern long-term (LT) reconstituting HSC function. Using a novel Asrij-mNeonGreen fluorescent reporter mouse, we show that the mitochondrial protein Asrij/OCIAD1 regulates HSC heterogeneity. We show that low Asrij expression marks quiescent HSCs with robust self-renewal capacity, whereas high Asrij expression identifies activated, differentiation-primed HSCs. These findings position Asrij as a novel determinant of HSC heterogeneity and introduce the Asrij-mNeonGreen reporter as a versatile tool for dissecting stem cell fate decisions in-vivo.

}, issn = {1873-2399}, doi = {10.1016/j.exphem.2025.105266}, author = {Prakash, Aishwarya and Halder, Souvik and Inamdar, Maneesha S} } @article {9056, title = {Cryo-EM structures of Mycobacterium tuberculosis imidazole glycerol phosphate dehydratase in the apo state and in the presence of small molecules [National Cryo-EM Facility, BLiSC]}, journal = {Acta Crystallogr F Struct Biol Commun}, year = {2025}, month = {2025 Jul 01}, abstract = {

Unlike humans, Mycobacterium tuberculosis (Mtb), the causative agent of human tuberculosis, has a de novo histidine-biosynthesis pathway. The enzyme imidazole glycerol phosphate dehydratase (IGPD), which catalyses the conversion of imidazole glycerol phosphate to imidazole acetol phosphate, has been studied extensively from various organisms and has become a major target for the development of antibacterial, antiweed and antifungal small molecules. In our previous studies, we have shown that in crystals IGPD forms a 24-mer oligomeric state in which the monomers are arranged in 432 symmetry. In order to gain insights into the oligomeric state of Mtb IGPD in solution, we determined cryogenic sample electron microscopy (cryo-EM) structures of apo IGPD at 2.2 and 3.1 {\r A} resolution. In addition, we also determined the cryo-EM structure of IGPD in the presence of 3-amino-1,2,4-triazole (ATZ) to 2.8 {\r A} resolution. The results of this work, which corroborate those from the crystallographic studies, indicate that IGPD forms a homo-oligomeric structure in solution comprising of 24 subunits. ATZ binds in the active-site pocket of the enzyme, which is located at the interface of three monomers and tethers 24 ATZ molecules. The results of this study suggest that cryo-EM, in addition to being a rapidly evolving and complementary imaging technology for elucidating 3D structures of biological macromolecules, can be useful in pinpointing the mode of binding small molecules of low mass (here \~{}85 Da) and mapping protein-ligand interactions, which could assist in the design of accurate (high-potency) inhibitors.

}, issn = {2053-230X}, doi = {10.1107/S2053230X25004595}, author = {Raina, Rahul and Kar, Deepsikha and Singla, Mohini and Tiwari, Satish and Kumari, Swati and Aneja, Sonanjali and Kumar, Varun and Banerjee, Soumya and Goyal, Shivika and Pal, Ravi Kant and Vinothkumar, Kutti R and Biswal, Bichitra} } @inbook {9067, title = {The Diagnostic Success Story: C-CAMP COVID-19 Innovations Deployment Accelerator (C-CIDA)}, booktitle = {At War with the Single Strand: How Indian Science Fought the COVID-19 Pandemic}, year = {2025}, pages = {53{\textendash}56}, publisher = {Springer Nature Singapore}, organization = {Springer Nature Singapore}, address = {Singapore}, abstract = {

Novel coronavirus COVID-19 emerged as one of the gravest public healthcare challenges in recent history. Even before the first lockdown started, we at C-CAMP had already felt the need to identify and work with such indigenously developed and indigenously produced COVID-19 innovations for rapid deployment. One of the first such innovations we fast-tracked in early March was a rapid diagnostics kit that has since received regulatory nods and was deployed in the field. As the virus marched on with no sign of reprieve, we recognized the importance of being more organized and focused in our efforts. Through the last decade or so of India{\textquoteright}s biotech boom, India{\textquoteright}s bio start-ups have been putting forward Made-in-India globally competitive solutions. Now was the time to capitalize on these technologies and align them with unmet needs for rapid deployment at an unprecedented scale.

}, isbn = {978-981-96-8122-8}, doi = {10.1007/978-981-96-8122-8_5}, url = {https://doi.org/10.1007/978-981-96-8122-8_5}, author = {Saiyed, Taslimarif and Chandavarkar, Nandita and Chakraborty, Debarshini and Janardan, Neelanjana and Alam, Mohammad Atif and Patel, Paresh and Gupta, Preksha and Bhardwaj, Vishal and Wala, Bhavisha and Joshi, Niranjan and Mukherjee, Titash and Vuppala, Kiran and Kiran, Amritanjali and Das, Deboleena Raha and Jain, Neeraj and Kumar, Manoj and Chandran, Subhashini and Rao, Madhusudhana and Joshi, Madhav and Kakkar, Abhishek and Khirani, Sekhar}, editor = {Swarup, Renu and Padmanaban, Govindarajan and Arora, Narendra Kumar} } @article {8961, title = {Integrated omics analysis of rice gene expression profiles and microRNAs identifies crucial target genes for nitrogen use efficiency [Next Gen Genomics Facility]}, journal = {Plant Physiology Reports}, year = {2025}, month = {Feb}, abstract = {

Nitrogen use efficiency is a complex trait in rice involving nitrogen uptake, utilization, and assimilation. The coding mRNAs and non-coding regulatory RNAs have combinatorial involvement in molecular mechanisms underlying nutrient use efficiency. In this study, we generated the microRNA expression profiles of panicle tissues in N-efficient (Basmati370, Thurur bhog) and non-efficient genotypes (Kola Joha3, IC463254) at the booting stage using microRNA sequencing and microarray expression profiles of root and shoot tissues at the seedling stage in N-efficient (Varadhan) and non-efficient (Prasanna) genotypes under hydroponic condition with N (+) and without N supplementation N (-) using Affymetrix rice microarray chip. To understand holistically the differential expression of mRNAs and microRNAs regulating NUE, we integrated the newly generated microRNA sequencing and microarray datasets with earlier generated mRNA gene expression profile data using bioinformatics tools. We present a consolidated set of genes operating tissue and stage-wise in rice for NUE. The microRNAs viz. miR2118o, miR1442, miR5149 were up-regulated and miR164, miR2867, miR171i were down-regulated in N efficient genotypes under low N conditions. Interestingly, we have identified the consistent differential expression of known and novel microRNAs in N efficient genotypes viz. miR1859 (down-regulated), miR1441, miR3979-5p, miR5149 (up-regulated) under low nitrogen conditions. The GARP-MYB/SANT domain, early nodulin genes were exclusively present in the shoot of Varadhan; genes related to polycomb complex, enhancer of zeste, were downregulated under N0 condition. Under low N conditions, brassinosteroid insensitive receptor kinase-1, IAA31, DUF581, DUF617, DUF803, NIN, nodulin, oligopeptide transporter, OsPOP1, calmodulin kinases were upregulated across roots and shoots. The genes upregulated in efficient genotypes expressed in shoots as well as in panicle tissues and regulated by downregulation of their miRNAs are targets for further studies. Remarkably, the downregulation of specific metabolic pathways was observed as an adaptive strategy to cope with nitrogen limitation, highlighting the complexity of NUE regulation in rice. These findings elucidate the intricate molecular networks governing NUE in rice and underscore the potential of specific microRNAs and genes as targets for precision breeding strategies aimed at enhancing nitrogen utilization efficiency in rice cultivation. The integrative transcriptomic approach presented here provides a foundation for further functional genomic studies, facilitating the development of novel breeding strategies to optimize NUE in rice.

}, issn = {2662-2548}, doi = {10.1007/s40502-024-00846-9}, url = {https://doi.org/10.1007/s40502-024-00846-9}, author = {Barbadikar, Kalyani M. and Talapanti, Krishna Kant and Bej, Sonali and Karre, Suman and Kothandaraman, Harish and Malathi, S. and Puskur, Raghuveer Rao and Mangrauthia, Satendra Kumar and Desiraju, Subrahmanyam and Voleti, S. R. and Sureshrao Sakhare, Akshay and Anantha, M. S. and Sundaram, R. M. and Chirravuri, Neeraja N.} } @article {9105, title = {Macrophage metabolic reprogramming during dietary stress influences adult body size in Drosophila [Central Imaging and Flow Cytometry Facility (INT)]}, journal = {EMBO Rep}, year = {2025}, month = {2025 Sep 09}, abstract = {

Immune cells are increasingly recognized as nutrient sensors; however, their developmental role in regulating growth under homeostasis or dietary stress remains elusive. Here, we show that Drosophila larval macrophages, in response to excessive dietary sugar (HSD), reprogram their metabolic state by activating glycolysis, thereby enhancing TCA-cycle flux, and increasing lipogenesis-while concurrently maintaining a lipolytic state. Although this immune-metabolic configuration correlates with growth retardation under HSD, our genetic analyses reveal that enhanced lipogenesis supports growth, whereas glycolysis and lipolysis are growth-inhibitory. Notably, promoting immune-driven lipogenesis offsets early growth inhibition in imaginal discs caused by glycolytic and lipolytic immune-metabolic states. Our findings reveal a model of immune-metabolic imbalance, where growth-suppressive states (glycolysis, lipolysis) dominate over a growth-supportive lipogenic state, thereby impairing early organ size control and ultimately affecting adult size. Overall, this study provides important insights into dietary stress-induced immune-metabolic reprogramming and its link to organ size regulation and early developmental plasticity.

}, issn = {1469-3178}, doi = {10.1038/s44319-025-00574-7}, author = {Mahanta, Anusree and Najar, Sajad Ahmad and Hariharan, Nivedita and Bhowmick, Ajit and Rizvi, Syed Iqra and Goyal, Manisha and Parupalli, Preethi and Subramanian, Ramaswamy and Giangrande, Angela and Palakodeti, Dasaradhi and Mukherjee, Tina} } @article {9122, title = {Microwave-assisted synthesis of silica quantum dots: a novel approach for targeting PI3K/AKT signaling in breast cancer therapy [Central Imaging and Flow Cytometry Facility, CIFF, Bangalore Life Science Cluster - NCBS - C-CAMP - InStem - TIGS]}, journal = {RSC Adv}, volume = {15}, year = {2025}, month = {2025 Oct 20}, pages = {39677-39688}, abstract = {

In this study, a facile and efficient approach was adopted for the first time to synthesize silica quantum dots (SiQDs) using tetraethyl orthosilicate (TEOS) as a precursor and ascorbic acid as a reducing agent. The synthesis was achieved microwave-assisted hydrolysis of TEOS in the presence of ascorbic acid, resulting in the formation of stable, highly fluorescent SiQDs. Extensive high-throughput characterization techniques were employed to investigate the optical, morphological, and structural properties of the as-prepared SiQDs, confirming their uniform size distribution, strong photoluminescence, and moderate stability. The therapeutic potential of SiQDs was explored against breast cancer cell lines (MCF-7 and MDA-MB-231), where they exhibited significant cytotoxicity and selective anticancer activity. Mechanistic investigations revealed that SiQDs effectively induce apoptosis in cancer cells by downregulating the PI3K/AKT signaling pathway, a critical regulator of cell survival and proliferation. These findings underscore the potential of SiQDs as promising anticancer agents in precision cancer therapy, offering a novel strategy for targeted cancer treatment. The study also paves the way for the design and development of silica-based nanomaterials with tailored properties for biomedical applications.

}, issn = {2046-2069}, doi = {10.1039/d5ra04715c}, author = {Selvaraj, Anitha and Ponnuchamy, Kumar} } @article {8677, title = {Distinct developmental patterns in Anopheles stephensi organ systems [Electron Microscopy (Micro-CT) Facility]}, journal = {Dev Biol}, year = {2024}, month = {2024 Jan 23}, abstract = {

Anatomical profiles of insects inform vector biology, comparative development and evolutionary studies with applications in forensics, agriculture and disease control. This study presents a comprehensive, high-resolution developmental profile of Anopheles stephensi, encompassing larval, pupal, and adult stages, obtained through microCT scanning. The results indicate in situ anatomical changes in most organ systems, including the central nervous system, eyes, musculature, alimentary canal, salivary glands, and ovaries, among other organ systems, except for the developing heart. We find significant differences in the mosquito gut, body-wall, and flight muscle development during metamorphosis from other dipterans like Drosophila. Specifically, indirect flight muscle specification and growth can be traced back at least to the 4th instar A. stephensi larvae, as opposed to post-puparial development in other Dipterans like Drosophila and Calliphora. Further, while Drosophila larval body-wall muscles and gut undergo histolysis, changes to these organs during mosquito metamorphosis are less pronounced. These observations, and raw data therein may serve as a reference for studies on the development and the genetics of mosquitoes. Overall, the detailed developmental profile of A. stephensi presented here illuminates the unique anatomy and developmental processes of Culicidae, with important implications for vector biology, disease control, and comparative evolutionary studies.

}, issn = {1095-564X}, doi = {10.1016/j.ydbio.2024.01.008}, author = {Agrawal, Khushboo and Prabhakar, Sunil and Bakthavachalu, Baskar and Chaturvedi, Dhananjay} } @article {8841, title = {Exploring advances in single particle CryoEM with apoferritin: From blobs to true atomic resolution [Electron Microscopy (Micro-CT) Facility (INT)]}, journal = {Int J Biochem Cell Biol}, volume = {169}, year = {2024}, month = {2024 Apr}, pages = {106536}, abstract = {

Deciphering the three-dimensional structures of macromolecules is of paramount importance for gaining insights into their functions and roles in human health and disease. Single particle cryoEM has emerged as a powerful technique that enables direct visualization of macromolecules and their complexes, and through subsequent averaging, achieve near atomic-level resolution. A major breakthrough was recently achieved with the determination of the apoferritin structure at true atomic resolution. In this review, we discuss the latest technological innovations across the entire single-particle workflow, which have been instrumental in driving the resolution revolution and in transforming cryoEM as a mainstream technique in structural biology. We illustrate these advancements using apoferritin as an example that has served as an excellent benchmark sample for assessing emerging technologies. We further explore whether the existing technology can routinely generate atomic structures of dynamic macromolecules that more accurately represent real-world samples, the limitations in the workflow, and the current approaches employed to overcome them.

}, keywords = {Apoferritins, Cryoelectron microscopy, Humans, Macromolecular Substances}, issn = {1878-5875}, doi = {10.1016/j.biocel.2024.106536}, author = {Premageetha, Gowtham ThambraRajan and Vinothkumar, Kutti R and Bose, Sucharita} } @article {8678, title = {Exploring Advances in Single Particle CryoEM with Apoferritin: from Blobs to True Atomic Resolution [National Cryo-EM Facility, BLiSC (INT)]}, journal = {The International Journal of Biochemistry \& Cell Biology}, year = {2024}, month = {02/2024}, type = {Review}, abstract = {

Deciphering the three-dimensional structures of macromolecules is of paramount importance for gaining insights into their functions and roles in human health and disease. Single particle cryoEM has emerged as a powerful technique that enables direct visualization of macromolecules and their complexes, and through subsequent averaging, achieve near atomic-level resolution. A major breakthrough was recently achieved with the determination of the apoferritin structure at true atomic resolution. In this review, we discuss the latest technological innovations across the entire single-particle workflow, which have been instrumental in driving and expanding the resolution revolution and in transforming cryoEM as a mainstream technique in structural biology. We illustrate these advancements using apoferritin as an example that has served as an excellent benchmark sample for assessing emerging technologies. We further explore whether the existing technology can routinely generate atomic structures of dynamic macromolecules that more accurately represent real-world samples, the limitations in the workflow and the current approaches employed to overcome them.

}, doi = {https://doi.org/10.1016/j.biocel.2024.106536}, url = {https://www.sciencedirect.com/science/article/abs/pii/S135727252400027X}, author = {Gowtham ThambraRajan Premageetha and Kutti R. Vinothkumar and Sucharita Bose} } @article {8675, title = {Generation and characterization of retinal pigment epithelium from patient iPSC line to model oculocutaneous albinism (OCA)1A disease [EyeStem Research Pvt. Ltd. - a C-CAMP Startup and Discovery to Innovation Accelerator, C-CAMP]}, journal = {J. Biosciences }, volume = {49}, year = {2024}, month = {01/2024}, abstract = {

Oculocutaneous albinism (OCA) is characterized by reduced melanin biosynthesis affecting the retina, thus impairing visual function. The disease pathology of OCA is poorly understood at the cellular level due to unavailability of suitable biological model systems. This study aimed to develop a disease-specific\ in vitro\ model for OCA type 1A, the most severe form caused by\ TYR\ (tyrosinase) gene mutations, using retinal pigment epithelium (RPE) differentiated from patient-derived human-induced pluripotent stem cells (hiPSCs). A comparative study between healthy and OCA1A RPE cells revealed that while healthy RPE cells exhibited timely onest of pigmentation during differentiation, OCA1A RPE cells failed to pigment even after an extended culture period. This observation was validated by ultrastructural studies using electron microscopy, hinting at melanosome-specific defects. Immunocytochemistry demonstrated abnormal expression patterns of melanogenesis-specific protein markers in OCA1A RPE cells, indicating reduced or absence of melanin synthesis. Next, a quantitative assay was\ performed to\ confirm the\ absence of melanin production in\ OCA1A RPE cells. Tyrosinase assay showed no activity in OCA1A compared with healthy RPE, suggesting non-functionality of\ TYR, further corroborated by western blot analysis showing complete absence of the protein. Gene expression by RNA sequencing of healthy and OCA1A RPE cells uncovered differential gene expression associated with lens development, visual perception, transmembrane transporter activity, and key signaling pathways. This disease-in-a-dish model of OCA1A provides an excellent platform to understand disease mechanism, identify potential therapeutic targets, and facilitate gene therapy or gene correction.

}, doi = {/doi.org/10.1007/s12038-023-00406-7}, url = {https://link.springer.com/article/10.1007/s12038-023-00406-7}, author = {Janavi Subramani and Niharika Patlolla and Rajani Battu and Taslimarif Saiyed and Rajarshi Pal} } @article {8957, title = {Lactobacillus fermentum SNR1: Probiotic efficiency and characterization of the antimicrobial peptide against dental caries and foodborne pathogens [Mass Spectrometry - Proteomics]}, journal = {The Microbe}, volume = {5}, year = {2024}, pages = {100190}, abstract = {

In this study, fermented rice gruel, a traditional food in South Telangana, India, was used as a source for the isolation of biologically significant probiotics. A total of 56 morphologically diverse strains were isolated and screened for their ability to inhibit dental biofilms as well as foodborne and aquaculture bacterial pathogens. Among these, isolate SNR1 was selected for further characterization due to its potential antimicrobial activity against bacterial pathogens such as Streptococcus mutans, Listeria monocytogenes, Staphylococcus aureus, Vibrio parahaemolyticus and Salmonella typhi. The results of 16S rDNA sequencing showed that isolate SNR1 was closely related to Lactobacillus fermentum strains from different sources. The bacteriocin peptide with potential activity was extracted and purified from the cell-free supernatant using ammonium sulphate precipitation, gel permeation chromatography and reversed-phase high performance liquid chromatography (RP-HPLC), and was named peptide SNR1. Tricine SDS-PAGE followed by MALDI-TOF-MS analysis revealed that the purified peptide had a molecular mass of 4.33 kDa, with the N-terminal sequence identified as MPTTSHFHLSPQGK. The purified peptide SNR1 showed good stability with various physiological enzymes, temperatures and pH conditions.

}, keywords = {Anti-biofilm activity, Anti-microbial activity, Dental caries, SNR1}, issn = {2950-1946}, doi = {https://doi.org/10.1016/j.microb.2024.100190}, url = {https://www.sciencedirect.com/science/article/pii/S2950194624001572}, author = {Ayyanna Repally and Esakkiraj Palanichamy and Natwar Jha and Arul Venkatesan and Ankaiah Dasari} } @article {8863, title = {LC/MS Based Characterization Workflow of GLP-1 Therapeutic Peptide Liraglutide and Its Impurities [Agilent Technologies - C-CAMP Collaboration]}, year = {2024}, abstract = {

This application note demonstrates the power of the Agilent 1290 Infinity II liquid chromatograph (LC) coupled with the Agilent 6545XT AdvanceBio liquid chromatography/quadrupole time-of-flight mass spectrometer (LC/Q-TOF MS) for identification and characterization of therapeutic peptides. This application is exemplified for the GLP-1 agonist liraglutide and its impurities. The therapeutic peptide workflow within Agilent MassHunter BioConfirm software allows for accurate mass-based identification, sequence conformation, and identification of impurities, with extra or missing amino acid impurities that can arise during solid phase synthesis. These impurities may pose a potential safety risk and eventually may impact the efficacy of the product.

}, url = {https://www.agilent.com/cs/library/applications/an-liraglutide-impurities-characterization-5994-7727en-agilent.pdf}, author = {Shadab Ahmad and Nirpendra Singh and Ashish Pargaonkar and Dheeraj Vig and Mike Knierman} } @article {8824, title = {Macrophage-conditioned medium enhances tunneling nanotube formation in breast cancer cells via PKC, Src, NF-κB, and p38 MAPK signaling [Next Gen Genomics Facility]}, journal = {Cell Signal}, year = {2024}, month = {2024 Jun 25}, pages = {111274}, abstract = {

Tumor-associated macrophages (TAMs) secrete cytokines, chemokines, and growth factors in the tumor microenvironment (TME) to support cancer progression. Higher TAM infiltration in the breast TME is associated with a poor prognosis. Previous studies have demonstrated the role of macrophages in stimulating long-range intercellular bridges referred to as tunneling nanotubes (TNTs) in cancer cells. Intercellular communication between cancer cells via TNTs promotes cancer growth, invasion, metastasis, and therapy resistance. Given the important role of TNTs and macrophages in cancer, the role of macrophage-induced TNTs in chemotherapy drug doxorubicin resistance is not known. Furthermore, the mechanism of macrophage-mediated TNT formation is elusive. In this study, it is shown that the macrophage-conditioned medium (MΦCM) partially mimicked inflammatory TME, induced an EMT phenotype, and increased migration in MCF-7 breast cancer cells. Additionally, secreted proteins in MΦCM induced TNT formation in MCF-7 cells, which led to increased resistance to doxorubicin. Transcriptomic analysis of MΦCM-treated MCF-7 cells showed enrichment of the NF-κB and focal adhesion pathways, as well as upregulation of genes involved in EMT, extracellular remodeling, and actin cytoskeleton reorganization. Interestingly, inhibitors of PKC, Src, NF-κB, and p38 decreased macrophage-induced TNT formation in MCF-7 cells. These results reveal the novel role of PKC and Src in inducing TNT formation in cancer cells and suggest that inhibition of PKC and Src activity may likely contribute to reduced macrophage-breast cancer cell interaction and the potential therapeutic strategy of cancer.

}, issn = {1873-3913}, doi = {10.1016/j.cellsig.2024.111274}, author = {Melwani, Pooja Kamal and Balla, Murali Mohan Sagar and Bhamani, Aman and Nandha, Shivani R and Checker, Rahul and Pandey, Badri Narain} } @article {8708, title = {Multiplexed fluorescence and scatter detection with single cell resolution using on-chip fiber optics for droplet microfluidic applications [Discovery to Innovation Accelerator, C-CAMP]}, journal = {Microsyst Nanoeng}, volume = {10}, year = {2024}, month = {2024}, pages = {35}, abstract = {

Droplet microfluidics has emerged as a critical component of several high-throughput single-cell analysis techniques in biomedical research and diagnostics. Despite significant progress in the development of individual assays, multiparametric optical sensing of droplets and their encapsulated contents has been challenging. The current approaches, most commonly involving microscopy-based high-speed imaging of droplets, are technically complex and require expensive instrumentation, limiting their widespread adoption. To address these limitations, we developed the OptiDrop platform; this platform is a novel optofluidic setup that leverages the principles of flow cytometry. Our platform enables on-chip detection of the scatter and multiple fluorescence signals from the microfluidic droplets and their contents using optical fibers. The highly customizable on-chip optical fiber-based signal detection system enables simplified, miniaturized, low-cost, multiparametric sensing of optical signals with high sensitivity and single-cell resolution within each droplet. To demonstrate the ability of the OptiDrop platform, we conducted a differential expression analysis of the major histocompatibility complex (MHC) protein in response to IFN stimulation. Our results showed the platform{\textquoteright}s ability to sensitively detect cell surface biomarkers using fluorescently labeled antibodies. Thus, the OptiDrop platform combines the versatility of flow cytometry with the power of droplet microfluidics to provide wide-ranging, scalable optical sensing solutions for research and diagnostics.

}, issn = {2055-7434}, doi = {10.1038/s41378-024-00665-w}, author = {Gupta, Preksha and Mohan, Ambili and Mishra, Apurv and Nair, Atindra and Chowdhury, Neeladri and Balekai, Dhanush and Rai, Kavyashree and Prabhakar, Anil and Saiyed, Taslimarif} } @article {8813, title = {A novel acidic pH-dependent metacaspase governs defense-response against pathogens in tomato [Mass Spectrometry - Proteomics Facility]}, journal = {Plant Physiology and Biochemistry}, volume = {213}, year = {2024}, pages = {108850}, abstract = {

The importance of metacaspases in programmed cell death and tissue differentiation is known, but their significance in disease stress response, particularly in a crop plant, remained enigmatic. We show the tomato metacaspase expression landscape undergoes differential reprogramming during biotrophic and necrotrophic modes of pathogenesis; also, the metacaspase activity dynamics correlate with the disease progression. These stresses have contrasting effects on the expression pattern of SlMC8, a Type II metacaspase, indicating that SlMC8 is crucial for stress response. In accordance, selected biotic stress-related transcription factors repress SlMC8 promoter activity. Interestingly, SlMC8 exhibits maximum proteolysis at an acidic pH range of 5{\textendash}6. Molecular dynamics simulation identified the low pH-driven protonation event of Glu246 as critical to stabilize the interaction of SlMC8 with its substrate. Mutagenesis of Glu246 to charge-neutral glutamine suppressed SlMC8{\textquoteright}s proteolytic activity, corroborating the importance of the amino acid in SlMC8 activation. The glutamic acid residue is found in an equivalent position in metacaspases having acidic pH dependence. SlMC8 overexpression leads to heightened ROS levels, cell death, and tolerance to PstDC3000, and SlMC8 repression reversed the phenomena. However, the overexpression of SlMC8 increases tomato susceptibility to necrotrophic Alternaria solani. We propose that SlMC8 activation due to concurrent changes in cellular pH during infection contributes to the basal resistance of the plant by promoting cell death at the site of infection, and the low pH dependence acts as a guard against unwarranted cell death. Our study confirms the essentiality of a low pH-driven Type II metacaspase in tomato biotic stress-response regulation.

}, keywords = {Cell death, host-pathogen interaction, Metacaspase, pH sensitivity, Tomato}, issn = {0981-9428}, doi = {https://doi.org/10.1016/j.plaphy.2024.108850}, url = {https://www.sciencedirect.com/science/article/pii/S0981942824005187}, author = {Shrabani Basak and Debarati Paul and Rohit Das and Shubhra Ghosh Dastidar and Pallob Kundu} } @article {8739, title = {Proteomic and metabolomic insights into seed germination of Ferula assa-foetida [Mass Spectrometry - Proteomics Facility]}, journal = {J Proteomics}, volume = {300}, year = {2024}, month = {2024 May 30}, pages = {105176}, abstract = {

Cold stratification is known to affect the speed of seed germination; however, its regulation at the molecular level in Ferula assa-foetida remains ambiguous. Here, we used cold stratification (4\ {\textdegree}C in the dark) to induce germination in F. assa-foetida and adopted a proteomic and metabolomic approach to understand the molecular mechanism of germination. Compared to the control, we identified 209 non-redundant proteins and 96 metabolites in germinated F. assa-foetida seed. Results highlight the common and unique regulatory mechanisms like signaling cascade, reactivation of energy metabolism, activation of ROS scavenging system, DNA repair, gene expression cascade, cytoskeleton, and cell wall modulation in F. assa-foetida germination. A protein-protein interaction network identifies 18 hub protein species central to the interactome and could be a key player in F. assa-foetida germination. Further, the predominant metabolic pathways like glucosinolate biosynthesis, arginine and proline metabolism, cysteine and methionine metabolism, aminoacyl-tRNA biosynthesis, and carotenoid biosynthesis in germinating seed may indicate the regulation of carbon and nitrogen metabolism is prime essential to maintain the physiology of germinating seedlings. The findings of this study provide a better understanding of cold stratification-induced seed germination, which might be utilized for genetic modification and traditional breeding of Ferula assa-foetida. SIGNIFICANCE: Seed germination is the fundamental checkpoint for plant growth and development, which has ecological significance. Ferula assa-foetida L., commonly known as "asafoetida," is a medicinal and food crop with huge therapeutic potential. To date, our understanding of F. assa-foetida seed germination is rudimentary. Therefore, studying the molecular mechanism that governs dormancy decay and the onset of germination in F. assa-foetida is essential for understanding the basic principle of seed germination, which could offer to improve genetic modification and traditional breeding.

}, keywords = {Ferula, Gene Expression Regulation, Plant, Germination, Metabolomics, Plant Proteins, Protein Interaction Maps, Proteome, Proteomics, Seeds}, issn = {1876-7737}, doi = {10.1016/j.jprot.2024.105176}, author = {Punia, Ashwani and Kumari, Manglesh and Chouhan, Monika and Saini, Vishal and Joshi, Robin and Kumar, Ashok and Kumar, Rajiv} } @article {8562, title = {Comparative stability study and aggregate analysis of Bevacizumab marketed formulations using advanced analytical techniques [Biologics / Biopharmaceutical Characterization Facility]}, journal = {Heliyon}, volume = {9}, year = {2023}, pages = {e19478}, abstract = {

Bevacizumab (Bvz) is the most preferred recombinant humanized monoclonal antibody in biosimilar development due to its prominence as a standard treatment in the oncology space. Therapeutic monoclonal antibodies are typically more complex and unlikely to produce a replica. As a result, regulatory agencies allow approval of biosimilars that differ structurally and functionally from their reference product, but these differences should not have any clinical significance. To identify these significant discrepancies, it is essential to perform a thorough characterization of critical product attributes both in real-time and after storage until the product{\textquoteright}s expiration. In the present study, two Bvz biosimilar brands (Bio-1 and Bio-2) marketed in India were evaluated and compared with the reference product Avastin{\textregistered} to assess their degree of similarity. A comprehensive physicochemical characterization of biosimilars and reference product was performed using orthogonal techniques including LC-ESI-QTOF, MALDI-TOF, FTIR-ATR, iCIEF, rCE, nrCE, UV280, and RP-HPLC. Furthermore, Bvz formulations under study were subjected to various stress conditions of thermal (elevated temperature 50\ {\textpm}\ 2\ {\textdegree}C), chemical (acidic pH 3.0\ {\textpm}\ 0.2, neutral pH 7.0\ {\textpm}\ 0.2, and basic pH 10.0\ {\textpm}\ 0.2), and mechanical (agitation 200\ rpm) for comparative stability evaluation. Any alteration in the secondary structure of the native protein was detected and quantified using far-UV circular dichroism (CD), indicating an average of 15\% and 11\% loss in native antiparallel β-sheet conformation respectively in Bio-1 and Bio-2 upon exposure to elevated temperature and high pH. Additionally, covalent or non-covalent aggregates formed as a function of elevated temperature and agitation were quantified using SEC-MALS.

}, keywords = {Aggregate analysis, Bevacizumab, Biosimilars, Intact mass analysis, stress study}, issn = {2405-8440}, doi = {https://doi.org/10.1016/j.heliyon.2023.e19478}, url = {https://www.sciencedirect.com/science/article/pii/S2405844023066860}, author = {Arpit Arunkumar Bana and Nithin Sajeev and Sabyasachi Halder and Haidar Abbas Masi and Shikha Patel and Priti Mehta} } @article {8062, title = {CRISPR/Cas9 and FLP-FRT mediated regulatory dissection of the BX-C of Drosophila melanogaster [Transgenic Fly Facility]}, journal = {Chromosome Res}, volume = {31}, year = {2023}, month = {2023 Jan 31}, pages = {7}, abstract = {

The homeotic genes or Hox define the anterior-posterior (AP) body axis formation in bilaterians and are often present on the chromosome in an order collinear to their function across the AP axis. However, there are many cases wherein the Hox are not collinear, but their expression pattern is conserved across the AP axis. The expression pattern of Hox is attributed to the cis-regulatory modules (CRMs) consisting of enhancers, initiators, or repressor elements that regulate the genes in a segment-specific manner. In the Drosophila melanogaster Hox complex, the bithorax complex (BX-C) and even the CRMs are organized in an order that is collinear to their function in the thoracic and abdominal segments. In the present study, the regulatorily inert regions were targeted using CRISPR/Cas9 to generate a series of transgenic lines with the insertion of FRT sequences. These FRT lines are repurposed to shuffle the CRMs associated with Abd-B to generate modular deletion, duplication, or inversion of multiple CRMs. The rearrangements yielded entirely novel phenotypes in the fly suggesting the requirement of such complex manipulations to address the significance of higher order arrangement of the CRMs. The functional map and the transgenic flies generated in this study are important resources to decipher the collective ability of multiple regulatory elements in the eukaryotic genome to function as complex modules.

}, keywords = {Animals, CRISPR-Cas Systems, Drosophila melanogaster, Drosophila Proteins, Gene Expression Regulation, Developmental, Homeodomain Proteins, Regulatory Sequences, Nucleic Acid}, issn = {1573-6849}, doi = {10.1007/s10577-023-09716-w}, author = {Hajirnis, Nikhil and Pandey, Shubhanshu and Mishra, Rakesh K} } @article {8466, title = {Cryo-EM structure of GABA transporter 1 reveals substrate recognition and transport mechanism [National Cryo-Electron Microscopy Facility]}, journal = {Nat Struct Mol Biol}, year = {2023}, month = {2023 Jul 03}, abstract = {

The inhibitory neurotransmitter γ-aminobutyric acid (GABA) is cleared from the synaptic cleft by the sodium- and chloride-coupled GABA transporter GAT1. Inhibition of GAT1 prolongs the GABAergic signaling at the synapse and is a strategy to treat certain forms of epilepsy. In this study, we present the cryo-electron microscopy structure of Rattus norvegicus GABA transporter 1 (rGAT1) at a resolution of 3.1 {\r A}. The structure elucidation was facilitated by epitope transfer of a fragment-antigen binding (Fab) interaction site from the Drosophila dopamine transporter (dDAT) to rGAT1. The structure reveals rGAT1 in a cytosol-facing conformation, with a linear density in the primary binding site that accommodates a molecule of GABA, a displaced ion density proximal to Na site 1 and a bound chloride ion. A unique insertion in TM10 aids the formation of a compact, closed extracellular gate. Besides yielding mechanistic insights into ion and substrate recognition, our study will enable the rational design of specific antiepileptics.

}, issn = {1545-9985}, doi = {10.1038/s41594-023-01011-w}, url = {https://www.nature.com/articles/s41594-023-01011-w}, author = {Nayak, Smruti Ranjan and Joseph, Deepthi and H{\"o}fner, Georg and Dakua, Archishman and Athreya, Arunabh and Wanner, Klaus T and Kanner, Baruch I and Penmatsa, Aravind} } @article {8544, title = {Distinct evolution of type I glutamine synthetase in Plasmodium and its species-specific requirement [Mass Spectrometry Facility - Metabolomics]}, journal = {Nat Commun}, volume = {14}, year = {2023}, month = {2023 Jul 14}, pages = {4216}, abstract = {

Malaria parasite lacks canonical pathways for amino acid biosynthesis and depends primarily on hemoglobin degradation and extracellular resources for amino acids. Interestingly, a putative gene for glutamine synthetase (GS) is retained despite glutamine being an abundant amino acid in human and mosquito hosts. Here we show Plasmodium GS has evolved as a unique type I enzyme with distinct structural and regulatory properties to adapt to the asexual niche. Methionine sulfoximine (MSO) and phosphinothricin (PPT) inhibit parasite GS activity. GS is localized to the parasite cytosol and abundantly expressed in all the life cycle stages. Parasite GS displays species-specific requirement in Plasmodium falciparum (Pf) having asparagine-rich proteome. Targeting PfGS affects asparagine levels and inhibits protein synthesis through eIF2α phosphorylation leading to parasite death. Exposure of artemisinin-resistant Pf parasites to MSO and PPT inhibits the emergence of viable parasites upon artemisinin treatment.

}, keywords = {Amino Acids, Animals, Artemisinins, Asparagine, Glutamate-Ammonia Ligase, Glutamine, Humans, Parasites, Plasmodium falciparum}, issn = {2041-1723}, doi = {10.1038/s41467-023-39670-4}, author = {Ghosh, Sourav and Kundu, Rajib and Chandana, Manjunatha and Das, Rahul and Anand, Aditya and Beura, Subhashree and Bobde, Ruchir Chandrakant and Jain, Vishal and Prabhu, Sowmya Ramakant and Behera, Prativa Kumari and Mohanty, Akshaya Kumar and Chakrapani, Mahabala and Satyamoorthy, Kapaettu and Suryawanshi, Amol Ratnakar and Dixit, Anshuman and Padmanaban, Govindarajan and Nagaraj, Viswanathan Arun} } @article {8318, title = {High-quality single amplicon sequencing method for illumina MiSeq platform using pool of {\textquoteright}N{\textquoteright} (0-10) spacer-linked target specific primers without PhiX spike-in [Next Gen Genomics Facility (INT)]}, journal = {BMC Genomics}, volume = {24}, year = {2023}, month = {2023 Mar 23}, pages = {141}, abstract = {

BACKGROUND: Illumina sequencing platform requires base diversity in the initial 11 cycles for efficient cluster identification and colour matrix estimation. This limitation yields low-quality data for amplicon libraries having homogeneous base composition. Spike-in of PhiX library ensures base diversity but reduces the overall number of sequencing reads for data analysis. To overcome such low diversity issues during amplicon sequencing on illumina platforms, we developed a high throughput single amplicon sequencing method by introducing {\textquoteright}N{\textquoteright} (0-10) spacers in target gene amplification primers that are pooled for simple handling.

RESULT: We evaluated the efficiency of {\textquoteright}N{\textquoteright} (0-10) spacer-linked primers by targeting bacterial 16S V3-V4 region, demonstrating heterogeneous base library construction. The addition of {\textquoteright}N{\textquoteright} (0-10) spacers causes sequencing frameshift at every base that leads to base diversity and produces heterogeneous high quality reads within a single amplicon library. We have written a python based command-line software,"MetReTrim", to trim the {\textquoteright}N{\textquoteright} (0-10) spacers from the raw reads ( https://github.com/Mohak91/MetReTrim ). We further demonstrated the accuracy of this method by comparative mock community analysis with standard illumina V3-V4 primer method. The ZymoBIOMICS{\texttrademark} microbial community DNA standard was used as a control for this study. We performed data analysisusing the DADA2 pipeline where taxonomy was assigned using SILVA database as reference. We observed no difference between the communities represented by our method and standard illumina V3-V4 primer method.

CONCLUSION: This method eliminates the need for PhiX spike-in for single amplicon sequencing on illumina MiSeq platform. This allows for sequencing of more number of samples in a run and a reduction in the overall cost. Given that Illumina sequencing works on SBS chemistry irrespective of the platform (such as HiSeq, MiSeq, NextSeq, NovaSeq, etc.) we propose that this strategy of using {\textquoteright}N{\textquoteright} (0-10) spacer-linked primer design can be adopted for generating high-quality single locus amplicon sequencing in a high throughput manner across the illumina platform subject to further validation.

}, keywords = {Bacteria, Gene Library, High-Throughput Nucleotide Sequencing, Microbiota, RNA, Ribosomal, 16S, Sequence Analysis, DNA}, issn = {1471-2164}, doi = {10.1186/s12864-023-09233-4}, author = {Naik, Tejali and Sharda, Mohak and C P, Lakshminarayanan and Virbhadra, Kumar and Pandit, Awadhesh} } @article {8472, title = {Marinobacterium lacunae sp. nov. isolated from estuarine sediment [Next Gen Genomics Facility]}, journal = {Arch Microbiol}, volume = {205}, year = {2023}, month = {2023 Jul 22}, pages = {294}, abstract = {

A novel motile bacterium was isolated from a sediment sample collected in Kochi backwaters, Kerala, India. This bacterium is Gram negative, rod shaped, 1.0-1.5\ {\textmu}m wide, and 2.0-3.0\ {\textmu}m long. It was designated as strain AK27. Colonies were grown on marine agar displayed circular, off-white, shiny, moist, translucent, flat, margin entire, 1-2\ mm in diameter. The major fatty acids identified in this strain were C ω7c, C, and summed in feature 3. The composition of polar lipids in the strain AK27 included phosphatidylglycerol, phosphatidylethanolamine, diphosphatidylglycerol, one unidentified amino lipid, two unidentified aminophospholipids, two unidentified phospholipids, and six unidentified lipids. The genomic DNA of strain AK27 exhibited a G+C content of 56.4\ mol\%. Based on the analysis of 16S rRNA gene sequence, strain AK27 showed sequence similarity to M. ramblicola D7 and M. zhoushanense WM3 as 98.99\% and 98.58\%, respectively. Compared to other type strains of the Marinobacterium genus, strain AK27 exhibited sequence similarities ranging from 91.7\% to 96.4\%. When compared to Marinobacterium zhoushanense WM3 and Marinobacterium ramblicola D7, strain AK27 exhibited average nucleotide identity values of 80.25\% and 79.97\%, and dDDH values of 22.9\% and 22.6\%, respectively. The genome size of the strain AK27 was 4.55\ Mb, with 4,229 coding sequences. Based on the observed phenotypic and chemotaxonomic features, and the results of phylogenetic and phylogenomic analysis, this study proposes the classification of strain AK27 as a novel species within the genus Marinobacterium. The proposed name for this novel species is Marinobacterium lacunae sp. nov.

}, keywords = {Agar, Alteromonadaceae, Cardiolipins, Phylogeny, RNA, Ribosomal, 16S}, issn = {1432-072X}, doi = {10.1007/s00203-023-03627-4}, author = {Tanuku, Srinivas Naga Radha and Pinnaka, Anil Kumar and Behera, Swarnaprava and Singh, Aditya and Pydi, Sudharani and Vasudeva, Gunjan and Vaidya, Bhumika and Sharma, Gunjan and Ganta, Sampath Kumar and Garbhapu, Naveen Sagar} } @article {8319, title = {Significance of Plasmodium berghei Amino Acid Transporter 1 in Food Vacuole Functionality and Its Association with Cerebral Pathogenesis [Mass Spectrometry Facility - Metabolomics]}, journal = {Microbiol Spectr}, year = {2023}, month = {2023 Mar 28}, pages = {e0494322}, abstract = {

The food vacuole plays a central role in the blood stage of parasite development by digesting host hemoglobin acquired from red blood cells and detoxifying the host heme released during hemoglobin digestion into hemozoin. Blood-stage parasites undergo periodic schizont bursts, releasing food vacuoles containing hemozoin. Clinical studies in malaria-infected patients and animal studies have shown the association of hemozoin with disease pathogenesis and abnormal host immune responses in malaria. Here, we perform a detailed characterization of putative Plasmodium berghei amino acid transporter 1 localized in the food vacuole to understand its significance in the malaria parasite. We show that the targeted deletion of amino acid transporter 1 in Plasmodium berghei leads to a swollen food vacuole phenotype with the accumulation of host hemoglobin-derived peptides. Plasmodium berghei amino acid transporter 1-knockout parasites produce less hemozoin, and the hemozoin crystals display a thin morphology compared with wild-type parasites. The knockout parasites show reduced sensitivity to chloroquine and amodiaquine by showing recrudescence. More importantly, mice infected with the knockout parasites are protected from cerebral malaria and display reduced neuronal inflammation and cerebral complications. Genetic complementation of the knockout parasites restores the food vacuole morphology with hemozoin levels similar to that of wild-type parasites, causing cerebral malaria in the infected mice. The knockout parasites also show a significant delay in male gametocyte exflagellation. Our findings highlight the significance of amino acid transporter 1 in food vacuole functionality and its association with malaria pathogenesis and gametocyte development. Food vacuoles of the malaria parasite are involved in the degradation of red blood cell hemoglobin. The amino acids derived from hemoglobin degradation support parasite growth, and the heme released is detoxified into hemozoin. Antimalarials such as quinolines target hemozoin formation in the food vacuole. Food vacuole transporters transport hemoglobin-derived amino acids and peptides from the food vacuole to the parasite cytosol. Such transporters are also associated with drug resistance. Here, we show that the deletion of amino acid transporter 1 in Plasmodium berghei leads to swollen food vacuoles with the accumulation of hemoglobin-derived peptides. The transporter-deleted parasites generate less hemozoin with thin crystal morphology and show reduced sensitivity to quinolines. Mice infected with transporter-deleted parasites are protected from cerebral malaria. There is also a delay in male gametocyte exflagellation, affecting transmission. Our findings uncover the functional significance of amino acid transporter 1 in the life cycle of the malaria parasite.

}, issn = {2165-0497}, doi = {10.1128/spectrum.04943-22}, author = {Anand, Aditya and Chandana, Manjunatha and Ghosh, Sourav and Das, Rahul and Singh, Nalini and Vaishalli, Pradeep Mini and Gantasala, Nagavara Prasad and Padmanaban, Govindarajan and Nagaraj, Viswanathan Arun} } @article {8587, title = {Systematic identification of CAZymes and transcription factors in the hypercellulolytic fungus Penicillium funiculosum NCIM1228 involved in lignocellulosic biomass degradation [Next Gen Genomics Facility]}, journal = {Biotechnol Biofuels Bioprod}, volume = {16}, year = {2023}, month = {2023 Oct 04}, pages = {150}, abstract = {

BACKGROUND: Penicillium funiculosum NCIM1228 is a filamentous fungus that was identified in our laboratory to have high cellulolytic activity. Analysis of its secretome suggested that it responds to different carbon substrates by secreting\ specific enzymes capable of digesting those substrates. This phenomenon indicated the presence of a regulatory system guiding the expression of these hydrolyzing enzymes. Since transcription factors (TFs) are the key players in regulating the expression of enzymes, this study aimed first to identify the complete repertoire of Carbohydrate Active Enzymes (CAZymes) and TFs coded in its genome. The regulation of CAZymes was then analysed by studying the expression pattern of these CAZymes and TFs in different carbon substrates-Avicel (cellulosic substrate), wheat bran (WB; hemicellulosic substrate), Avicel + wheat bran, pre-treated wheat straw (a potential substrate for lignocellulosic ethanol), and glucose (control).

RESULTS: The P. funiculosum NCIM1228 genome was sequenced, and 10,739 genes were identified in its genome. These genes included a total of 298 CAZymes and 451 TF coding genes. A distinct expression pattern of the CAZymes was observed in different carbon substrates tested. Core cellulose hydrolyzing enzymes were highly expressed in the presence of Avicel, while pre-treated wheat straw and Avicel + wheat bran induced a mixture of CAZymes because of their heterogeneous nature. Wheat bran mainly induced hemicellulases, and the least number of CAZymes were expressed in glucose. TFs also exhibited distinct expression patterns in each of the carbon substrates. Though most of these TFs have not been functionally characterized before, homologs of NosA, Fcr1, and ATF21, which have been known to be involved in fruiting body development, protein secretion and\ stress response, were identified.

CONCLUSIONS: Overall, the P. funiculosum NCIM1228 genome was sequenced, and the CAZymes and TFs present in its genome were annotated. The expression of the CAZymes and TFs in response to various polymeric sugars present in the lignocellulosic biomass was identified. This work thus provides a comprehensive mapping of transcription factors (TFs) involved in regulating the production of biomass hydrolyzing enzymes.

}, issn = {2731-3654}, doi = {10.1186/s13068-023-02399-9}, author = {Pasari, Nandita and Gupta, Mayank and Sinha, Tulika and Ogunmolu, Funso Emmanuel and Yazdani, Syed Shams} } @article {2916, title = {Characterization of ACE inhibitory and antioxidant peptides in yak and cow milk hard chhurpi cheese of the Sikkim Himalayan region [Mass Spectrometry Proteomics Facility]}, journal = {Food Chemistry: X}, volume = {13}, year = {2022}, month = {03/2022}, pages = {100231}, type = {Journal Article}, abstract = {

In this study, simulated in vitro GI digestion of the Himalayan hard chhurpi cheese resulted in the increase of hydrolyzed protein content, antioxidant and ACE-inhibitory activities. LC-MS/MS-based peptidomics revealed a total of 1473 peptides in the samples originating from different milk proteins, including α-S1-casein, α-S2-casein, β-casein, κ-casein, α-lactalbumin, and β-lactoglobulin, out of which 60 peptides have been reported for different functional properties. A total of 101 peptides were predicted to be antihypertensive using the bioactivity prediction web servers, AHTpin and mAHTPred. In silico molecular docking studies predicted 20 antihypertensive peptides, exhibiting non-bond interactions between hard chhurpi peptides and ACE catalytic residues. A peptide, SLVYPFPGPI, identified in GI digested cow hard chhurpi and undigested, and GI digested samples of yak hard chhurpi, showed a stronger binding affinity towards ACE. Identifying antioxidant and ACE inhibitory peptides in hard cheese products adds value to them as functional foods of the Himalayan region.

}, keywords = {Antihypertensive, Bioactive peptides, Hard chhurpi, Molecular docking, Proteomics, Sikkim Himalaya}, doi = {https://doi.org/10.1016/j.fochx.2022.100231}, url = {https://www.sciencedirect.com/science/article/pii/S2590157522000293}, author = {Abedin, Md Minhajul and Chourasia, Rounak and Phukon, Loreni Chiring and Singh, Sudhir P and Rai, Amit Kumar} } @article {4869, title = {Dual control of dopamine in Drosophila myeloid-like progenitor cell proliferation and regulation of lymph gland growth. [Central Imaging \& Flow Cytometry Facility]}, journal = {EMBO Rep}, year = {2022}, month = {2022 Apr 27}, pages = {e52951}, type = {Journal Article}, abstract = {

In Drosophila, definitive haematopoiesis takes place in a specialized organ termed "lymph gland". It harbours multi-potent stem-like blood progenitor cells whose development controls overall growth of this haematopoietic tissue and formation of mature blood cells. With respect to its development, neurotransmitters have emerged as potent regulators of blood-progenitor cell development and function. In this study, we extend our understanding of neurotransmitters and show that progenitors are self-sufficient with regard to synthesizing dopamine, a well-established neurotransmitter. These cells also have modules for dopamine sensing through the receptor and transporter. We found that modulating expression of these components in progenitor cells affected lymph gland growth, which suggested growth-promoting function of dopamine in blood-progenitor cells. Cell-cycle analysis of developing lymph glands revealed an unexpected requirement for intracellular dopamine in moderating the progression of early progenitor cells from S to G2 phase of the cell cycle, while activation of dopamine receptor signalling later in development regulated their progression from G2 and entry into mitosis. The dual capacity in which dopamine operated, first intracellularly to coordinate S/G2 transition and later extracellularly in G2/M transition, was critical for the growth of the lymph gland. Overall, the data presented highlight a novel non-canonical use of dopamine in the myeloid system that reveals an uncharacterized function of intracellular dopamine in cell-cycle phasing with outcomes on haematopoietic growth and immunity as well.

}, keywords = {cell cycle, dopamine signalling, dopamine synthesis, haematopoietic progenitors, proliferation}, issn = {1469-3178}, doi = {10.15252/embr.202152951}, url = {embopress.org/doi/abs/10.15252/embr.202152951}, author = {Kapoor, Ankita and Padmavathi, Achalla and Madhwal, Sukanya and Mukherjee, Tina} } @article {3706, title = {Effectiveness of a novel, non-intrusive, continuous-use air decontamination technology to reduce microbial contamination in clinical settings: a multi-centric study [Biomoneta Research Pvt. Ltd., a C-CAMP Startup]}, journal = {J Hosp Infect}, volume = {123}, year = {2022}, month = {2022 Feb 16}, pages = {15-22}, abstract = {

BACKGROUND: Despite rigorous disinfection and fumigation, healthcare-associated infection (HAI) remains a significant concern in healthcare settings. We have developed a novel airborne-microbicidal technology {\textquoteright}ZeBox{\textquoteright} which clears \>99.999\% of airborne microbial load under controlled laboratory conditions.

AIM: To evaluate the clinical performance of ZeBox in reducing airborne and surface microbial load.

METHODS: The study was conducted in single-bed and multi-bed intensive care units (ICUs) of two hospitals. Airborne and surface microbial loads were sampled pre and post deployment of ZeBox at pre-determined sites. Statistical significance of the reduction was determined using the Mann-Whitney U-test.

FINDINGS: ZeBox brought statistically significant reduction of both airborne and surface bacterial and fungal load. In both hospital ICUs, airborne and surface bacterial load decreased by 90\% and 75\% on average respectively, providing a low bioburden zone of 10-15 feet diameter around the unit. The reduced microbial level was maintained during ZeBox{\textquoteright}s operation over several weeks. Most clinical bacterial isolates recovered from one of the hospitals were antibiotic resistant, highlighting ZeBox{\textquoteright}s ability to eliminate antimicrobial-resistant bacteria among others.

CONCLUSION: ZeBox significantly reduces airborne and surface microbial burden in clinical settings. It thereby serves an unmet need for reducing the incidence of HAI.

}, issn = {1532-2939}, doi = {10.1016/j.jhin.2022.02.002}, author = {Nagaraj, S and Chandrasingh, S and Jose, S and Sofia, B and Sampath, S and Krishna, B and Menon, I and Kundu, D and Parekh, S and Madival, D and Nandi, V and Ghatak, A} } @article {3630, title = {Immune profile and responses of a novel dengue DNA vaccine encoding an EDIII-NS1 consensus design based on Indo-African sequences [C-CAMP Bioincubation Facility]}, journal = {Molecular Therapy - Cell Press}, year = {2022}, month = {2022 Jan 07}, type = {Journal Article}, abstract = {

The ongoing COVID-19 pandemic highlights the need to tackle viral variants, expand the number of antigens, and assess diverse delivery systems for vaccines against emerging viruses. In the present study, a DNA vaccine candidate was generated by combining in tandem envelope protein domain III (EDIII) of dengue virus serotypes 1-4 and a dengue virus (DENV)-2 non-structural protein 1 (NS1) protein-coding region. Each domain was designed as a serotype-specific consensus coding sequence derived from different genotypes based on the whole genome sequencing of clinical isolates in\ India and complemented with data from Africa. This sequence was further optimized for protein expression. In silico structural analysis of the EDIII consensus sequence revealed that epitopes are structurally conserved and immunogenic. The vaccination of mice with this construct induced pan-serotype neutralizing antibodies and antigen-specific T\ cell responses. Assaying intracellular interferon (IFN)-γ staining, immunoglobulin IgG2(a/c)/IgG1 ratios, and immune gene profiling suggests a strong Th1-dominant immune\ response. Finally, the passive transfer of immune sera protected AG129 mice challenged with a virulent, non-mouse-adapted DENV-2 strain. Our findings collectively suggest an alternative strategy for dengue vaccine design by offering a novel vaccine candidate with a possible broad-spectrum protection and a successful clinical translation either as a stand alone or in a mix and match strategy.

}, keywords = {antibody-dependent enhancement, consensus sequence, dengue, dengue surveillance, DNA vaccine, EDIII domain, NS1 protein}, issn = {1525-0024}, doi = {10.1016/j.ymthe.2022.01.013}, url = {https://www.cell.com/molecular-therapy-family/molecular-therapy/fulltext/S1525-0016(22)00013-2$\#$secsectitle0165}, author = {Sankaradoss, Arun and Jagtap, Suraj and Nazir, Junaid and Moula, Shefta E and Modak, Ayan and Fialho, Joshuah and Iyer, Meenakshi and Shastri, Jayanthi S and Dias, Mary and Gadepalli, Ravisekhar and Aggarwal, Alisha and Vedpathak, Manoj and Agrawal, Sachee and Pandit, Awadhesh and Nisheetha, Amul and Kumar, Anuj and Bordoloi, Mahasweta and Shafi, Mohamed and Shelar, Bhagyashree and Balachandra, Swathi S and Damodar, Tina and Masika, Moses Muia and Mwaura, Patrick and Anzala, Omu and Muthumani, Kar and Sowdhamini, Ramanathan and Medigeshi, Guruprasad R and Roy, Rahul and Pattabiraman, Chitra and Krishna, Sudhir and Sreekumar, Easwaran} } @article {3480, title = {Production and characterization of bioactive peptides from rice beans using Bacillus subtilis [Mass Spectrometry - Proteomics Facility]}, journal = {Bioresource Technology}, year = {2022}, month = {01/2022}, pages = {126932}, type = {Journal Article}, abstract = {

A bioprocess was developed for production of bioactive peptides on microbial fermentation of rice beans using proteolytic Bacillus subtilis strains. The peptides produced were identified by LC-MS/MS analysis, revealing the presence of many unique peptide sequences to individual hydrolysates. On functional properties prediction, antihypertensive peptides (3.90\%) were found to be higher in comparison to other bioactive peptides. Among different strains, B. subtilis KN2B fermented hydrolysate exhibited highest angiotensin converting enzyme (ACE)-inhibitory activity (45.73\%). Furthermore, 19 selected peptides, including the common and unique peptides were examined for their affinity towards the binding cavity of ACE using molecular docking. The results showed a common peptide PFPIPFPIPIPLP, and another IPFPPIPFLPPI unique to B. subtilis KN2B fermented hydrolysate exhibited promising binding at the ACE binding site with substantial free binding energy. The process developed can be used for the production of bioactive peptides from rice bean for application in nutraceutical industries.

}, keywords = {ACE-inhibitory peptide, Bacillus subtilis, Fermented legume, Molecular docking, Rice bean}, doi = {https://doi.org/10.1016/j.biortech.2022.126932}, url = {https://www.sciencedirect.com/science/article/abs/pii/S0960852422002619}, author = {Padhi, Srichandan and Chourasia, Rounak and Kumari, Megha and Singh, Sudhir P and Rai, Amit Kumar} } @article {6120, title = {Quantitative insight into the metabolism of isoprene-producing Synechocystis sp. PCC 6803 using steady state C-MFA. [Mass Spectrometry Facility]}, journal = {Photosynth Res}, year = {2022}, month = {2022 Sep 07}, abstract = {

Cyanobacteria are photosynthetic bacteria, widely studied for the conversion of atmospheric carbon dioxide to useful platform chemicals. Isoprene is one such industrially important chemical, primarily used for production of synthetic rubber and biofuels. Synechocystis sp. PCC 6803, a genetically amenable cyanobacterium, produces isoprene on heterologous expression of isoprene synthase gene, albeit in very low quantities. Rationalized metabolic engineering to re-route the carbon flux for enhanced isoprene production requires in-dept knowledge of the metabolic flux distribution in the cell. Hence, in the present study, we undertook steady state 13C-metabolic flux analysis of glucose-tolerant wild-type (GTN) and isoprene-producing recombinant (ISP) Synechocystis sp. to understand and compare the carbon flux distribution in the two strains. The R-values for amino acids, flux analysis predictions and gene expression profiles emphasized predominance of Calvin cycle and glycogen metabolism in GTN. Alternatively, flux analysis predicted higher activity of the anaplerotic pathway through phosphoenolpyruvate carboxylase and malic enzyme in ISP. The striking difference in the Calvin cycle, glycogen metabolism and anaplerotic pathway activity in GTN and ISP suggested a possible role of energy molecules (ATP and NADPH) in regulating the carbon flux distribution in GTN and ISP. This claim was further supported by the transcript level of selected genes of the electron transport chain. This study provides the first quantitative insight into the carbon flux distribution of isoprene-producing cyanobacterium, information critical for developing Synechocystis sp. as a single cell factory for isoprenoid/terpenoid production.

}, keywords = {Cyanobacterium, Flux analysis, MEP pathway, Metabolic model, qRT-PCR}, issn = {1573-5079}, doi = {10.1007/s11120-022-00957-0}, url = {https://link.springer.com/article/10.1007/s11120-022-00957-0}, author = {Nirati, Yasha and Purushotham, Nidhish and Alagesan, Swathi} } @article {4451, title = {SARS-CoV-2 infection of human-induced pluripotent stem cells-derived lung lineage cells evokes inflammatory and chemosensory responses by targeting mitochondrial pathways [Eyestem Research Pvt. Ltd., a C-CAMP Startup]}, journal = {J Cell Physiol}, year = {2022}, month = {2022 Apr 23}, abstract = {

The COVID-19 disease caused by severe acute respiratory syndrome coronavirus 2\ (SARS-CoV-2) primarily affects the lung, particularly the proximal airway and distal alveolar cells. NKX2.1+ primordial lung progenitors of the foregut (anterior) endoderm are the developmental precursors to all adult lung epithelial lineages and are postulated to play an important role in viral tropism. Here, we show that SARS-CoV-2 readily infected and replicated in human-induced pluripotent stem cell-derived proximal airway cells, distal alveolar cells, and lung progenitors. In addition to the upregulation of antiviral defense and immune responses, transcriptomics data uncovered a robust epithelial cell-specific response, including perturbation of metabolic processes and disruption in the alveolar maturation program. We also identified spatiotemporal dysregulation of mitochondrial heme oxygenase 1 (HMOX1), which is associated with defense against antioxidant-induced lung injury. Cytokines, such as TNF-α, INF-γ, IL-6, and IL-13, were upregulated in infected cells sparking mitochondrial ROS production and change in electron transport chain complexes. Increased mitochondrial ROS then activated additional proinflammatory cytokines leading to an aberrant cell cycle resulting in apoptosis. Notably, we are the first to report a chemosensory response resulting from SARS-CoV-2 infection similar to that seen in COVID-19 patients. Some of our key findings were validated using COVID-19-affected postmortem lung tissue sections. These results suggest that our in vitro system could serve as a suitable model to investigate the pathogenetic mechanisms of SARS-CoV-2 infection and to discover and test therapeutic drugs against COVID-19 or its consequences.

}, issn = {1097-4652}, doi = {10.1002/jcp.30755}, author = {Surendran, Harshini and Kumar, Saurabh and Narasimhaiah, Swathi and Ananthamurthy, Anuradha and Varghese, P S and D{\textquoteright}Souza, George A and Medigeshi, Guruprasad and Pal, Rajarshi} } @article {6781, title = {Techno-commercial application of microencapsulated basil oil and thyme oil and their antibacterial activity against aquatic pathogens [Electron Microscopy Facility]}, journal = {International Journal of Fisheries and Aquatic Studies}, volume = {10}, year = {2022}, month = {10/2022}, chapter = {101-106}, abstract = {

A complete study was conducted through manufacturing of Microencapsulation of Basil Oil Powder (MEBOP) and Microencapsulation of Thyme Oil Powder (METOP) to evaluate the anti-bacterial activity against aquatic pathogens. An oil-in-water emulsion were prepared using gum with dextrin and the resultant matrix was spray dried with the average yield of 45\% w/w of MEBOP and 50\% w/w of METOP. Total oil content in the encapsulated powder in both were found to be 24.39\% w/w of MEBOP 25.80\% w/w of METOP respectively. Scanning Electron Microscopic (SEM) analysis was also done to confirm the encapsulation of oil and study possible structure of matrix. Microencapsulated powders were further subjected for antibacterial activity against aquaculture pathogens like Vibrio harvey, Vibrio parahaemolyticus, Bacillus cereus, Aeromonas sp, E. coli and Salmonella sp. Results were compared against the respective oils at different concentrations. Phyto- constituents from Basil oil and Thyme oil like Thymol, Linalool, and Methyl chavicol were quantified using HPLC (High-Performance Liquid Chromatography) and GC (Gas Chromatography) which may be responsible for this activity. Furthermore, stress studies were conducted at 60 {\textdegree}C to understand the stability of Encapsulated oil powders to this and its commercial usage in formulations.

}, keywords = {anti-bacterial, aqua gut health, Microencapsulated oil powder, stress study}, issn = {2394-0506}, doi = {10.22271/fish.2022.v10.i5b.2730}, url = {https://www.fisheriesjournal.com/archives/2022/vol10issue5/PartB/10-4-44-468.pdf}, author = {Sampath M and Ganesh Muguli and Sameer Pai and Babu UV} } @article {7318, title = {Techno-commercial application of microencapsulated basil oil and thyme oil and their antibacterial activity against aquatic pathogens [EM Facility]}, journal = {International Journal of Fisheries and Aquatic Studies 2022}, volume = {10(5)}, year = {2022}, month = {23/06/2022}, pages = {101-106}, type = {Journal Article}, chapter = {101}, abstract = {

A complete study was conducted through manufacturing of Microencapsulation of Basil Oil Powder (MEBOP) and Microencapsulation of Thyme Oil Powder (METOP) to evaluate the anti-bacterial activity against aquatic pathogens. An oil-in-water emulsion were prepared using gum with dextrin and the resultant matrix was spray dried with the average yield of 45\% w/w of MEBOP and 50\% w/w of METOP. Total oil content in the encapsulated powder in both were found to be 24.39\% w/w of MEBOP 25.80\% w/w of METOP respectively. Scanning Electron Microscopic (SEM) analysis was also done to confirm the encapsulation of oil and study possible structure of matrix. Microencapsulated powders were further subjected for antibacterial activity against aquaculture pathogens like Vibrio harvey, Vibrio parahaemolyticus, Bacillus cereus, Aeromonas sp, E. coli and Salmonella sp. Results were compared against the respective oils at different concentrations. Phyto- constituents from Basil oil and Thyme oil like Thymol, Linalool, and Methyl chavicol were quantified using HPLC (High-Performance Liquid Chromatography) and GC (Gas Chromatography) which may be responsible for this activity. Furthermore, stress studies were conducted at 60 {\textdegree}C to understand the stability of Encapsulated oil powders to this and its commercial usage in formulations.

}, keywords = {anti-bacterial, aqua gut health, Microencapsulated oil powder, stress study}, issn = {E-ISSN: 2347-5129}, doi = {https://doi.org/10.22271/fish.2022.v10.i5b.2730}, url = {https://www.fisheriesjournal.com/archives/2022/vol10issue5/PartB/10-4-44-468.pdf}, author = {Sampath, M and Muguli, Ganesh and Pai, Sameer and Babu, UV} } @article {2733, title = {Transcriptome analysis at mid-stage seed development in litchi with contrasting seed size [Next Gen Genomics Facility]}, journal = {3 Biotech }, volume = {12}, year = {2022}, month = {01/2022}, abstract = {

Litchi is a sub-tropical fruit crop with genotypes that bear fruits with variable seed size. Small seed size is a desirable trait\ in litchi, as it improves consumers{\textquoteright} preference and facilitates fruit processing. Seed specific transcriptome analysis was performed in two litchi genotypes with contrasting seed size to identify the genes associated with seed development. The transcriptomic sequence data from seeds at mid-development stages (16{\textendash}28\ days after anthesis) were de-novo assembled into 1,39,608 Trinity transcripts. Out of these, 6325 transcripts expressed differentially between the two contrasting genotypes. Several putative genes for salicylic acid, jasmonic acid and brassinosteriod pathways were down-regulated in seeds of the small-seeded litchi. The putative regulators of seed maturation and seed storage were down-regulated in the small-seeded genotype. Embryogenesis, cell\ expansion, seed size and stress related Trinity transcripts exhibited differential expression. Further studies on gene characterization will reveal the early regulators of seed size in litchi.

}, keywords = {Embryogenesis, Litchi, Seed size, Small seed, Transcriptome}, doi = {doi.org/10.1007/s13205-021-03098-8}, url = {https://link.springer.com/article/10.1007/s13205-021-03098-8$\#$citeas}, author = {Ashish K. Pathak and Sudhir P. Singh and Ritika Sharma and Vishal Nath and Rakesh Tuli} } @article {1866, title = {Alterations of Primary Metabolites in Root Exudates of Intercropped Cajanus cajan{\textendash}Zea mays Modulate the Adaptation and Proteome of Ensifer (Sinorhizobium) fredii NGR234 [Mass Spectrometry - Proteomics Facility]}, journal = {Microbial Ecology}, year = {2021}, month = {08/2021}, pages = {1{\textendash}18}, type = {Journal Article}, abstract = {

Legume-cereal intercropping systems, in the context of diversity, ecological function, and better yield have been widely studied. Such systems enhance nutrient phytoavailability by balancing root-rhizosphere interactions. Root exudates (RE) play an important role in the rhizospheric interactions of plant-plant and/or plant-microbiome interaction. However, the influence of the primary metabolites of RE on plant-rhizobia interactions in a legume-cereal intercrop system is not known. To understand the plant communication with rhizobia, Cajanus cajan-Zea mays intercropped plants and the broad host range legume nodulating Ensifer fredii NGR234 as the model plants and rhizobium used respectively. A metabolomics-based approach revealed a clear separation between intercropped and monocropped RE of the two plants. Intercropped C. cajan showed an increase in the myo-inositol, and proline, while intercropped Z. mays showed enhanced galactose, D-glucopyranoside, and arginine in the RE. Physiological assays of NGR234 with the RE of intercropped C. cajan exhibited a significant enhancement in biofilm formation, while intercropped Z. mays RE accelerated the bacterial growth in the late log phase. Further, using label-free proteomics, we identified a total of 2570 proteins of NGR234 covering 50\% annotated protein sequences upon exposure to Z. mays RE. Furthermore, intercropped Z. mays RE upregulated bacterioferritin comigratory protein (BCP), putative nitroreductase, IlvD, LeuC, D (branched-chain amino acid proteins), and chaperonin proteins GroEL2. Identification offered new insights into the metabolome of the legume-cereal intercrop and proteome of NGR234-Z. mays interactions that underline the new molecular candidates likely to be involved in the fitness of rhizobium in the intercropping system.

}, keywords = {Cajanus cajan Zea mays, Ensifer fredii NGR234, Metabolomics, Proteomics, Root exudates (RE)}, doi = {https://doi.org/10.1007/s00248-021-01818-4}, url = {https://link.springer.com/article/10.1007/s00248-021-01818-4}, author = {Vora, Siddhi M and Ankati, Sravani and Patole, Chhaya and Podile, Appa Rao and Archana, G} } @article {1858, title = {Azaindole Based Potentiator of Antibiotics against Gram-Negative Bacteria [C-CAMP Startup Bugworks]}, journal = {ACS Infectious Diseases}, year = {2021}, month = {10/2021}, type = {Journal Article}, abstract = {
We discovered azaindole-based compounds with weak innate activity that exhibit substantial potentiation of antibacterial activities of different antibiotics, viz., rifampicin, erythromycin, solithromycin, and novobiocin in Gram-negative bacteria. In the presence of the azaindole derivatives, these antibiotics exhibited submicromolar minimum inhibitory concentrations (MICs) against Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Acinetobacter baumannii. The fold improvements in MIC of these antibiotics that were otherwise weak or inactive on their own against these bacteria were also observed against drug-resistant clinical isolates. Our studies indicate that this selective potentiation is probably through destabilization of the outer membrane{\textquoteright}s integrity, known to be regulated by the lipopolysaccharides (LPS). Thus, the azaindole based compounds described here open opportunities for those antibiotics that are otherwise ineffective due to LPS mediated entry barriers in Gram-negative bacteria.
}, keywords = {Azaindole, Bacterial Permeability, Lipopolysachhardies (LPS), Polymyxin, Synergy}, doi = {10.1021/acsinfecdis.1c00171}, url = {https://pubs.acs.org/doi/abs/10.1021/acsinfecdis.1c00171$\#$}, author = {Sharma, Sreevalli and Rao, Ranga and Reeve, Stephanie M and Phelps, Gregory A and Bharatham, Nagakumar and Katagihallimath, Nainesh and Ramachandran, Vasanthi and Raveendran, Savitha and Sarma, Maitrayee and Nath, Anubha} } @article {1679, title = {Chemical intervention for enhancing growth and reducing grain arsenic accumulation in rice [Mass Spectrometry - Metabolomics Facility]}, journal = {Environ Pollut}, volume = {276}, year = {2021}, month = {2021 Feb 13}, pages = {116719}, abstract = {

Arsenic (As) is a ubiquitous environmental carcinogen that enters the human food chain mainly through rice grains. In the present study, we evaluated the potential of thiourea (TU; non-physiological reactive oxygen species scavenger) in mitigating the negative effects of arsenic (As) stress in indica rice variety IR64, with the overall aim to reduce grain As accumulation. At seedling stage, As\ +\ TU treatment induced the formation of more numerous and longer crown roots compared with As alone. The As accumulation in main root, crown root, lower leaf and upper leaf was significantly reduced to 0.1-, 0.14-, 0.16-, 0.14-fold, respectively in As\ +\ TU treated seedlings compared with those of As alone. This reduced As accumulation was also coincided with light-dependent suppression in the expression levels of aquaporins and photosynthesis-related genes in As\ +\ TU treated roots. In addition, the foliar-supplemented TU under As-stress maintained reducing redox conditions which decreased the rate of As accumulation in flag leaves and, eventually grain As by 0.53-fold compared with those of As treatment. The agronomic feasibility of TU was validated under naturally As contaminated sites of Nadia (West Bengal, India). The tiller numbers and crop productivity (kg seed/ha) of TU-sprayed plants were increased by 1.5- and 1.18-fold, respectively; while, grain As accumulation was reduced by 0.36-fold compared with those of water-sprayed control. Thus, this study established TU application as a sustainable solution for cultivating rice in As-contaminated field conditions.

}, issn = {1873-6424}, doi = {10.1016/j.envpol.2021.116719}, author = {Srivastava, Ashish Kumar and Pandey, Manish and Ghate, Tejashree and Kumar, Vikash and Upadhyay, Munish Kumar and Majumdar, Arnab and Sanjukta, Abhay Kumar and Agrawal, Ashish Kumar and Bose, Sutapa and Srivastava, Sudhakar and Suprasanna, Penna} } @article {1699, title = {Derivation of Induced Pluripotent Stem Cell (iPSC) Lines from Patient-Specific Peripheral Blood Mononuclear Cells (PBMC) Using Episomal Vectors [Eyestem Research Pvt. Ltd., a C-CAMP Startup]}, journal = {Methods Mol Biol}, year = {2021}, month = {2021 Mar 27}, abstract = {

Inherited retinal diseases (IRDs) are a diverse group of rare eye disorders, resulting in vision loss or blindness. The underlying reason is mutation in one or more than 250 different genes associated with the development and normal physiology of retina largely comprising of rod/cone photoreceptors and retinal pigment epithelium. Interestingly, the sub retinal region of an eye has been shown to be immune privileged, broadening the scope of cell-replacement therapies for patients suffering from retinal degeneration. Several groups around the globe, including ours, have demonstrated safety and efficacy in preclinical studies by employing various approaches of retinal cell therapy. This had largely been possible with the advent of induced pluripotent stem cells (iPSC)-reprogrammed from adult somatic cells, that serves as a starting material for generating retinal cells de novo. Here, we describe a detailed procedure for reprogramming peripheral blood mononuclear cells (PBMC) into iPSC using episomal vectors without any physical disruption in the host genome. The lines thus created were tested for sterility, cytogenetic stability, identity, absence of episomal plasmids and further authenticated for pluripotency and tri-lineage differentiation capacity by embryoid body formation and immunocytochemistry. We believe that this feeder-cell free, animal-product free and gene-insertion free protocol would help people to develop and bank patient-specific cell lines for autologous cell therapies for incurable rare diseases.

}, issn = {1940-6029}, doi = {10.1007/7651_2021_385}, author = {Konala, Vijay Bhaskar Reddy and Nandakumar, Swapna and Surendran, Harshini and Pal, Rajarshi} } @article {1717, title = { Differential proteins associated with plasma membrane in X- and/or Y-chromosome bearing spermatozoa in indicus cattle [Mass Spectrmetry - Propteomics Facility]}, year = {2021}, month = {04/2021}, abstract = {

The differential proteins associated with plasma membrane of spermatozoa are less known, identification of which shall help overcome limitations of currently used methods of sperm sexing, considered as a high priority for livestock sector of many countries. This study has reported plasma membrane proteomics of unsorted spermatozoa and differential expression of plasma membrane-associated proteins between X- and Y-chromosome bearing spermatozoa of indicus cattle (Bos indicus). Isolation of plasma membrane fraction using percoll gradient, relatively a rapid method, from bovine spermatozoa has been reported to enrich isolation of plasma membrane proteins. Significant enrichment for plasma membrane-associated proteins was observed in plasma membrane fraction (p \< .05) as compared to the total cell lysate using LC-MS/MS. Furthermore, these experiments were conducted in flow cytometry sorted, sexed-semen samples. Thirteen proteins were identified as differentially abundant between X- and Y-sorted spermatozoa. Among these, two proteins were downregulated in Y-sorted spermatozoa compared to the X-sorted spermatozoa (p \< .05), while four and seven proteins could be noted in X- and Y-sorted spermatozoa, respectively. Proteins that are presumed to support sperm capacitation and sperm migration velocity were found to be abundant in Y-sorted spermatozoa while those associated with structural molecule activity were identified as abundant in X-sorted spermatozoa in the present study. Our study provides better insight into the plasma membrane proteomics of spermatozoa of indicus cattle and furnishes data that might aid in design and development of alternate and open technology for sex-sorting of semen.

https://pubmed.ncbi.nlm.nih.gov/33829570/

}, keywords = {X/Y-sorted spermatozoa; plasma membrane; pre-determination of sex; proteome; sperm sexing; unsorted spermatozoa.}, doi = {10.1111/rda.13936}, author = {Rongala Laxmivandana and Chhaya Patole and Tilak Raj Sharma and Kewal Krishan Sharma and Soumen Naskar} } @article {1691, title = {Geometry encoded functional programming of tumor homing peptides for targeted drug delivery [Image Analysis Support]}, journal = {J Control Release}, year = {2021}, month = {2021 Mar 12}, abstract = {

Poly-peptide molecules have shown promising applications in drug delivery and tumor targeting. A series of tumor homing peptides were designed by exhaustively sampling low energy geometrical basins of amino acids at specific sites of a peptide molecule to induce a conformational lock. This peptide library was pruned to a limited set of eight molecules, employing electrostatic interactions, docking, and molecular dynamics simulations. These designed and optimized peptides were synthesized and tested on various cell lines, including breast cancer (MDA-MB-231), cervical cancer (HeLa), osteosarcoma (U2-OS), and non-cancerous mammary epithelial cells (MCF-10A) using confocal microscopy and flow cytometry. Peptides show differential uptake in cancerous MDA-MB-231, HeLa, U2-OS, and non-cancerous MCF-10A cells. Confocal imaging verified their ability to penetrate even in 3D tumorospheres of MDA-MB-231 cells. Further, experiments of mitochondrial membrane potential depolarization and Caspase-3 activation confirmed that their cytotoxic effects are by apoptosis. Homing ability of the designed peptides in in vivo system and fluorescence imaging with clinical samples of human origin have further confirmed that the in vitro studies are qualitatively identical and quantitatively comparable in their ability to selectively recognize tumor cells. Overall, we present a roadmap for the functional programming of peptide-based homing and penetrating molecules that can perform selective tumor targeting.

}, issn = {1873-4995}, doi = {10.1016/j.jconrel.2021.03.010}, author = {Goyal, Ruchika and Jerath, Gaurav and Akhil, R and Chandrasekharan, Aneesh and Puppala, Eswara Rao and Ponneganti, Srikanth and Sarma, Anupam and Naidu, V G M and Santhoshkumar, T R and Ramakrishnan, Vibin} } @article {1860, title = {Glycomic and glycotranscriptomic profiling of mucin-type O-glycans in planarian Schmidtea mediterranea [Mass Spectrometry - Glycomics]}, journal = {Glycobiology}, year = {2021}, month = {09/2021}, type = {Journal Article}, abstract = {

O-Glycans on cell surfaces play important roles in cell-cell, cell-matrix, and receptor-ligand interaction. Therefore, glycan-based interactions are important for tissue regeneration and homeostasis. Free-living flatworm Schmidtea mediterranea, because of its robust regenerative potential, is of great interest in the field of stem cell biology and tissue regeneration. Nevertheless, information on the composition and structure of O-glycans in planaria is unknown. Using mass spectrometry and in silico approaches, we characterized the glycome and the related transcriptome of mucin-type O-glycans of planarian S. mediterranea. Mucin-type O-glycans were composed of multiple isomeric, methylated, and unusually extended mono- and di-substituted O-GalNAc structures. Extensions made of hexoses and 3-O methyl hexoses were the glycoforms observed. From glycotranscriptomic analysis, sixty genes belonging to five distinct enzyme classes were identified to be involved in mucin-type O-glycan biosynthesis. These genes shared homology with those in other invertebrate systems. While a majority of the genes involved in mucin-type O-glycan biosynthesis was highly expressed during organogenesis and in differentiated cells, a few select genes in each enzyme class were specifically enriched during early embryogenesis. Our results indicate a unique temporal and spatial role for mucin-type O-glycans during embryogenesis and organogenesis and in adulthood. In summary, this is the first report on O-glycans in planaria. This study expands the structural and biosynthetic possibilities in cellular glycosylation in the invertebrate glycome and provides a framework towards understanding the biological role of mucin-type O-glycans in tissue regeneration using planarians.

}, keywords = {Flatworm, GC-MS, Invertebrates, MALDI-TOF mass spectrometry, ScRNAseq}, doi = {https://doi.org/10.1093/glycob/cwab097}, url = {https://academic.oup.com/glycob/advance-article-abstract/doi/10.1093/glycob/cwab097/6361079}, author = {Subramanian, Sabarinath Peruvemba and Lakshmanan, Vairavan and Palakodeti, Dasaradhi and Subramanian, Ramaswamy} } @article {1854, title = {Interventional Strategies for Parkinson Disease: Can Neural Precursor Cells Forge a Path Ahead? [Eyestem Research, a C-CAMP startup]}, journal = {ACS Chemical Neuroscience}, year = {2021}, month = {10/2021}, type = {Journal Article}, abstract = {

Neural precursor cells (NPCs), derived from pluripotent stem cells (PSCs), with their unique ability to generate multiple neuronal and glial cell types are extremely useful for understanding biological mechanisms in normal and diseased states. However, generation of specific neuronal subtypes (mature) from NPCs in large numbers adequate for cell therapy is challenging due to lack of a thorough understanding of the cues that govern their differentiation. Interestingly, neural stem cells (NSCs) themselves are in consideration for therapy given their potency to form different neural cell types, release of trophic factors, and immunomodulatory effects that confer neuroprotection. With the recent COVID-19 outbreak and its accompanying neurological indications, the immunomodulatory role of NSCs may gain additional significance in the prevention of disease progression in vulnerable populations. In this regard, small-molecule mediated NPC generation from PSCs via NSC formation has become an important strategy that ensures consistency and robustness of the process. The development of the mammalian brain occurs along the rostro-caudal axis, and the establishment of anterior identity is an early event. Wnt signaling, along with fibroblast growth factor and retinoic acid, acts as a caudalization signal. Further, the increasing amount of epigenetic data available from human fetal brain development has enhanced both our understanding of and ability to experimentally manipulate these developmental regulatory programs in vitro. However, the impact on homing and engraftment after transplantation and subsequently on therapeutic efficacy of NPCs based on their derivation strategy is not yet clear. Another formidable challenge in cell replacement therapy for neurodegenerative disorders is the mode of delivery. In this Perspective, we discuss these core ideas with insights from our preliminary studies exploring the role of PSC-derived NPCs in rat models of MPTP-induced Parkinson{\textquoteright}s disease following intranasal injections.

}, keywords = {cell transplantation, induced pluripotent stem cells, neural precursor cell, neural stem cell., Parkinson disease}, doi = {https://doi.org/10.1021/acschemneuro.1c00525}, url = {https://pubs.acs.org/doi/abs/10.1021/acschemneuro.1c00525}, author = {Nandakumar, Swapna and Shahani, Pradnya and Datta, Indrani and Pal, Rajarshi} } @article {1681, title = {Live cells extracts of freshly cut Chicken and of Baby Sprouts of Mung Beans with UV Absorption and Proton NMR spectra [NMR Facility]}, journal = {Current Nutrition \& Food Science}, year = {2021}, abstract = {

Background: This article discloses information related to a recent patent filed by the author on Extracts of freshly cut farm birds and animals.

Objective: The objective was to produce {\textquotedblleft}Liquid-Protein{\textquotedblright} extracts obtained from live cells of protein rich meat of farmbirds or animals as well as from baby plants of pulses.

Method: Freshly cut meat pieces or sprouts of pulses are put in water and pulse-heated for 30 minutes. The nutritious water extract of these is taken that contains amino-acids/proteins and some signaling chemicals emitted from the stressed live cells.

Results: The heat-stressed animal cells (of Chicken) released Creatine and many other nutrients in the extract along with Guanosine triphosphate/Guanosine diphosphate/ Guanosine monophosphate/ Inosine Mono Phosphate (GTP/GDP/ GMP/IMP) showing a UV absorption peak at 249 nm. This paper analyses the UV -Visual Absorption spectra and proton NMR data for the extracts. It is disclosed that the vegetarian baby plant cells of pulse seeds released (ATP) Adenosine Tri Phosphate (264 nm peak) along with Resveratrol (306 nm peak) but did not produce Creatine and such an extract had side effects.

Conclusion: Cells of birds/animals are similar to those for humans, and the signaling chemicals in the non-vegetarian extract are non-toxic and 100\% compatible to humans as compared to plant cell extracts with incompatible chemicals. Since meat-cells manage to {\textquotedblleft}live{\textquotedblright} for longer than 10 hours with out blood/oxygen supply, by an unclear anaerobic cell respiration involving Creatine and GTP/GDP/GMP/IMP, the extract from these meat cells holds the key for metabolic cell repair for anti-aging of humans.

Keywords:\ Proton NMR spectra, UV Absorption spectra, Extract of chicken cells, Extract of Mumg and Kidney beans, Anti Aging, Creatine, IMP, GTP, ATP

}, doi = {10.2174/1573401317666210122091014}, author = {Pandian, G. Soundra} } @article {1862, title = {Mito-targeted antioxidant prevents cardiovascular remodelling in spontaneously hypertensive rat by modulation of energy metabolism [Mass Spectrometry - Proteomics Facility]}, journal = {Clinical and Experimental Pharmacology and Physiology}, year = {2021}, month = {08/2021}, type = {Journal Article}, abstract = {

Hypertension induced left ventricular hypertrophy (LVH) augments the risk of cardiovascular anomalies. Mitochondrial alterations result in oxidative stress, accompanied by decrease in fatty acid oxidation, leading to the activation of the hypertrophic program. Targeted antioxidants are expected to reduce mitochondrial reactive oxygen species more effectively than general antioxidants. This study was designed to assess whether the mito-targeted antioxidant, Mito-Tempol (Mito-TEMP) is more effective than the general oxidant, Tempol (TEMP) in reduction of hypertension and hypertrophy and prevention of shift in cardiac energy metabolism. Spontaneously hypertensive rats were administered either TEMP (20 mg/kg/day) or Mito-TEMP (2 mg/kg/day) intraperitoneally for 30 days. Post treatment, animals were subjected to 2D-echocardiography. Myocardial lysates were subjected to RPLC {\textendash} LTQ-Orbitrap-MS analysis. Mid-ventricular sections were probed for markers of energy metabolism and fibrosis. The beneficial effect on cardiovascular structure and function was significantly higher for Mito-TEMP. Increase in mitochondrial antioxidants and stimulation of fatty acid metabolism; with significant improvement in cardiovascular function was apparent in spontaneously hypertensive rats (SHR) treated with Mito-TEMP. The study indicates that Mito-TEMP is superior to its non- targeted isoform in preventing hypertension induced LVH, and the beneficial effects on heart are possibly mediated by reversal of metabolic remodelling.

}, keywords = {cardiac hypertrophy, cardiac metabolism, LC-MS, mitochondria targeted antioxidant, proteomic analysis, spontaneously hypertensive rat}, doi = {https://doi.org/10.1111/1440-1681.13585}, url = {https://onlinelibrary.wiley.com/doi/10.1111/1440-1681.13585}, author = {Potnuri, Ajay Godwin and Purushothaman, Sreeja and Saheera, Sherin and Nair, Renuka R} } @article {1917, title = {Novel non intrusive continuous use ZeBox technology to trap and kill airborne microbes [Biomoneta Research Pvt. Ltd., a C-CAMP Startup / Electron Microscopy Facility]}, journal = {Scientific Reports}, volume = {11}, year = {2021}, month = {11/2021}, pages = {Article number: 22779 }, abstract = {

Preventing nosocomial infection is a major unmet need of our times. Existing air decontamination technologies suffer from demerits such as toxicity of exposure, species specificity, noxious gas emission, environment-dependent performance and high power consumption. Here, we present a novel technology called {\textquotedblleft}ZeBox{\textquotedblright} that transcends the conventional limitations and achieves high microbicidal efficiency. In ZeBox, a non-ionizing electric field extracts naturally charged microbes from flowing air and deposits them on engineered microbicidal surfaces. The surface{\textquoteright}s three dimensional topography traps the microbes long enough for them to be inactivated. The electric field and chemical surfaces synergistically achieve rapid inactivation of a broad spectrum of microbes. ZeBox achieved near complete kill of airborne microbes in challenge tests (5{\textendash}9 log reduction) and\ \>90\%\ efficiency in a fully functional stem cell research facility in the presence of humans. Thus, ZeBox fulfills the dire need for a real-time, continuous, safe, trap-and-kill air decontamination technology.

}, doi = {doi.org/10.1038/s41598-021-02184-4}, author = {Kruttika S. Phadke and Deepak G. Madival and Janani Venkataraman and Debosmita Kundu and K. S. Ramanujan and Nisha Holla and Jaywant Arakeri and Gaurav Tomar and Santanu Datta and Arindam Ghatak} } @article {1639, title = {Transplantation of retinal pigment epithelium and photoreceptors generated concomitantly via small molecule-mediated differentiation rescues visual function in rodent models of retinal degeneration [Eyestem Research Pvt. Ltd., a C-CAMP Startup]}, journal = {Stem Cell Res Ther}, volume = {12}, year = {2021}, month = {2021 Jan 19}, pages = {70}, abstract = {

BACKGROUND: Age-related macular degeneration (AMD) is a result of degeneration/damage of the retinal pigment epithelium (RPE) while retinitis pigmentosa (RP), an inherited early-onset disease, results from premature loss of photoreceptors. A promising therapeutic approach for both is the replacement of lost/damaged cells with human induced pluripotent stem cell (hiPSC)-derived retinal cells.

METHODS: The aim of this study was to investigate the in vivo functionality of RPE and photoreceptor progenitor (PRP) cells derived from a clinical-grade hiPSC line through a unified protocol. De novo-generated RPE and PRP were characterized extensively to validate their identity, purity, and potency.

RESULTS: RPE expressed tight junction proteins, showed pigmentation and ciliation, and secreted polarization-related factors vascular endothelial growth factor (VEGF) and pigment epithelium-derived factor (PEDF). PRP expressed neural retina proteins and cone and rod markers, and responded to KCl-induced polarization. Transcriptomic analysis demonstrated an increase in the expression of mature retinal tissue-specific genes coupled with concomitant downregulation of genes from undesired lineages. RPE transplantation rescued visual function in RCS rats shown via optokinetic tracking and photoreceptor rescue. PRP transplantation improved light perception in NOD.SCID-rd1 mice, and positive electroretinography signals indicated functional photoreceptor activity in the host{\textquoteright}s outer nuclear layer. Graft survival and integration were confirmed using immunohistochemistry, and no animals showed teratoma formation or any kind of ectopic growth in the eye.

CONCLUSIONS: To our knowledge, this is the first demonstration of a unified, scalable, and GMP-adaptable protocol indicating strong animal efficacy and safety data with hiPSC-derived RPE and PRP cells. These findings provide robust proof-of-principle results for IND-enabling studies to test these potential regenerative cell therapies in patients.

}, issn = {1757-6512}, doi = {10.1186/s13287-021-02134-x}, author = {Surendran, Harshini and Nandakumar, Swapna and Reddy K, Vijay Bhaskar and Stoddard, Jonathan and Mohan K, Varsha and Upadhyay, Pramod K and McGill, Trevor J and Pal, Rajarshi} } @article {1323, title = {Dataset for the combined transcriptome assembly of M. oleifera and functional annotation [Next Gen Genomics Facility (INT)]}, journal = {Data in Brief}, year = {2020}, pages = {105416}, abstract = {

In this paper, we present the data acquired during transcriptome analysis of the plant Moringa oleifera [1] from five different tissues (root, stem, leaf, flower and seed) by RNA sequencing. A total of 271 million reads were assembled with an N50 of 2094bp. The combined transcriptome was assessed for transcript abundance across five tissues. The protein coding genes identified from the transcripts were annotated and used for orthology analysis. Further, enzymes involved in the biosynthesis of select medicinally important secondary metabolites, vitamins and ion transporters were identified and their expression levels across tissues were examined. The data generated by RNA sequencing has been deposited to NCBI public repository under the accession number PRJNA394193 (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA394193).

}, keywords = {Annotation, Enrichment analysis, Gene Expression, Metabolic pathway, Orthology, Transcriptome}, issn = {2352-3409}, doi = {https://doi.org/10.1016/j.dib.2020.105416}, url = {http://www.sciencedirect.com/science/article/pii/S2352340920303103}, author = {K. Mohamed Shafi and Adwait G. Joshi and Iyer Meenakshi and Shaik Naseer Pasha and K. Harini and Jarjapu Mahita and Radha Sivarajan Sajeevan and Snehal D. Karpe and Pritha Ghosh and Sathyanarayanan Nitish and A. Gandhimathi and Oommen K. Mathew and Subramanian Hari Prasanna and Manoharan Malini and Eshita Mutt and Mahantesha Naika and Nithin Ravooru and Rajas M. Rao and Prashant N. Shingate and Anshul Sukhwal and Margaret S. Sunitha and Atul K. Upadhyay and Rithvik S. Vinekar and Ramanathan Sowdhamini} } @article {1447, title = {Dietary supplementation of extracts of red sea weed (Kappaphycus alvarezii) improves growth, intestinal morphology, expression of intestinal genes and immune responses in broiler chickens [Sea6Energy Pvt. Ltd, a C-CAMP Startup]}, journal = {Journal of the Science of Food and Agriculture}, year = {2020}, month = {08, 2020}, type = {Research Article}, abstract = {

BACKGROUND

Effects of supplementation of dried alkaline (referred to as MVP1) and aqueous (referred to as PBD1) extracts of\ K. alvarezii\ , were evaluated in broiler (Vencobb 400) chickens (1{\textendash}35 d post-hatch). In experiment I, each of the seven diets (basal diet with three levels (0.5, 1.5 or 5.0 g kg-1\ diet) of MVP1 or PBD1 and a negative control) was fed to twelve pen replicates containing five birds in each. In experiment II, each of three diets (a negative control, and PBD1 at two levels (1.0 or 1.5 g kg-1\ diet)) was fed to sixteen pen replicates of five chicks in each.

RESULTS

Concentrations of total phenolics, phycobillins and free radical scavenging activity were higher (P\<0.01) whereas carrageenan was lower in PBD1 than in MVP1. In the experiment I, PBD1 at 1.5 g kg-1\ diet improved (P\<0.05) body weight (7.11\% higher). In the experiment II, both the treatments improved (P\<0.01) BW (9.18\% and 8.47\%, respectively) as compared to control. The group fed with PBD1@ 1.0 g kg-1\ had higher (P\<0.05) HI titre, expression of intestinal claudin 2, TLR2A, NOD1, avian beta defensin 4, interleukin 2 and 6 genes than control. Treatments did not influence feed efficiency or levels of most of the antioxidant enzymes. Villus width and crypt depth were significantly higher in the group fed with 1.5 g kg-1\ of PBD1.

CONCLUSION

Supplementing dried aqueous extract of\ Kappaphycus alvarezii\ at 1 g kg-1\ diet may be an effective strategy to increase growth and immunity in broiler chicken.

}, doi = {https://doi.org/10.1002/jsfa.10708}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/jsfa.10708}, author = {Paul, Shyam Sundar and Venkata, Hemanth Giri Rao Vantharam and Raju, MVLN and Rama Rao, SV and Nori, Sri Sailaja and Suryanarayan, Shrikumar and Kumar, Vikas and Perveen, Zeba and Srinivas Prasad, Cadaba} } @article {1530, title = {Elucidating the processes and pathways enriched in buffalo sperm proteome in regulating semen quality [Mass Spectrometry Facility - Proteomics]}, journal = {Cell Tissue Res}, year = {2020}, month = {2020 Nov 05}, abstract = {

Sperm carries a reservoir of proteins regulating the molecular functions to attain functional competence. Semen samples collected from buffalo bulls were assessed for sperm functional attributes (n\ =\ 11) and proteome profiling (n\ =\ 6). Sperm proteins were extracted and profiled by employing LC-MS/MS. Overall, the buffalo sperm contained 1365 proteins, of which 458 were common between the groups. The unique proteins were 477 and 430 in good and poor quality semen, respectively. In the whole proteome of buffalo sperm, sexual reproduction with phosphatidylethanolamine-binding protein1 (PEBP1), fetuin-B (FETUB) and acrosin (ACR) was the most enriched (p\ =\ 8.44E-19) biological process, also with thermogenesis (p\ =\ 0.003), oocyte meiosis (p\ =\ 0.007) and vascular smooth muscle contraction (p\ =\ 0.009) apart from metabolic pathways. In good quality semen, mesenchyme migration (p\ =\ 1.24E-07) and morphogenesis (p\ =\ 0.001) were abundant biological processes. In good quality semen, the fluid shear stress (p\ =\ 0.01) and, in poor quality semen, valine, leucine and isoleucine degradation (p\ =\ 3.8E-05) pathways were enriched. In good quality semen, 7 proteins were significantly (p\ \<\ 0.05) upregulated and 33 proteins were significantly (p\ \<\ 0.05) downregulated. On validating the abundantly expressed sperm proteins, serine protease inhibitor Kazal-type 2-like (SPINK2; 2.17-fold) and neddylin (NEDD8; 1.13-fold) were upregulated and YBX2 was downregulated (0.41-fold) in good quality semen as compared with poor quality semen (1-fold). The present findings revealed the importance of sperm proteins in oocyte maturation, fertilization process and early embryonic development. The variations in the proteomic composition can be used as potential markers for the selection of breeding bulls.

}, issn = {1432-0878}, doi = {10.1007/s00441-020-03303-9}, author = {Binsila, Bala Krishnan and Archana, Santhanahalli Siddalingappa and Ramya, Laxman and Swathi, Divakar and Selvaraju, Sellappan and Gowda, N K Shivakumar and Pal, Din Taran and Rafay, Abu and Bhatta, Raghavendra} } @article {1464, title = {Human induced pluripotent stem cell-derived lung epithelial system for SARS-CoV-2 infection modeling and its potential in drug repurposing [Eyestem Research Pvt. Ltd., a C-CAMP Startup]}, journal = {Stem Cells Dev}, year = {2020}, month = {2020 Aug 31}, abstract = {

The lung is the most vulnerable target for the SARS-CoV-2 infection and respiratory failure causing Acute Respiratory Distress Syndrome (ARDS) is its foremost outcome. However, the current primary in vitro models in use for SARS-CoV-2 display apparent limitations for modeling such complex human respiratory disease. While patient cells can directly model the effects of a drug, their availability and capacity for expansion are limited compared to the transformed/immortalized cells or the tumor-derived cell lines. What{\textquoteright}s more, the latter may harbor genetic and metabolic abnormalities in the course of their derivation, making them unsuitable for drug screening. Therefore, it is important to create physiologically relevant human-cell models that can replicate the pathophysiology of SARS-CoV-2, thus, facilitating drug testing. Here, we show preliminary data on how human induced pluripotent stem cells (hiPSC)-based lung epithelial cell system could emerge as a relevant and sensitive platform for modelling SARS-CoV-2 infection and drug screening.

}, issn = {1557-8534}, doi = {10.1089/scd.2020.0152}, author = {Surendran, Harshini and Nandakumar, Swapna and Pal, Rajarshi} } @article {1465, title = {A knowledge-driven protocol for prediction of proteins of interest with an emphasis on biosynthetic pathways [Next Gen Genomics Facility (INT)]}, journal = {MethodsX}, year = {2020}, pages = {101053}, abstract = {

This protocol describes a stepwise process to identify proteins of interest from a query proteome derived from NGS data. We implemented this protocol on Moringa oleifera transcriptome to identify proteins involved in secondary metabolite and vitamin biosynthesis and ion transport. This knowledge-driven protocol identifies proteins using an integrated approach involving sensitive sequence search and evolutionary relationships. We make use of functionally important residues (FIR) specific for the query protein family identified through its homologous sequences and literature. We screen protein hits based on the clustering with true homologues through phylogenetic tree reconstruction complemented with the FIR mapping. The protocol was validated for the protein hits through qRT-PCR and transcriptome quantification. Our protocol demonstrated a higher specificity as compared to other methods, particularly in distinguishing cross-family hits. This protocol was effective in transcriptome data analysis of M. oleifera as described in Pasha et. al.{\textbullet}Knowledge-driven protocol to identify secondary metabolite synthesizing protein in a highly specific manner.{\textbullet}Use of functionally important residues for screening of true hits.{\textbullet}Beneficial for metabolite pathway reconstruction in any (species, metagenomics) NGS data.

}, keywords = {functionally important residue, homology, multiple sequence alignment, Pathway, phylogenetic analysis}, issn = {2215-0161}, doi = {https://doi.org/10.1016/j.mex.2020.101053}, url = {http://www.sciencedirect.com/science/article/pii/S2215016120302739}, author = {Adwait G. Joshi and K. Harini and Iyer Meenakshi and K. Mohamed Shafi and Shaik Naseer Pasha and Jarjapu Mahita and Radha Sivarajan Sajeevan and Snehal D. Karpe and Pritha Ghosh and Sathyanarayanan Nitish and A. Gandhimathi and Oommen K. Mathew and Subramanian Hari Prasanna and Manoharan Malini and Eshita Mutt and Mahantesha Naika and Nithin Ravooru and Rajas M. Rao and Prashant N. Shingate and Anshul Sukhwal and Margaret S. Sunitha and Atul K. Upadhyay and Rithvik S. Vinekar and Ramanathan Sowdhamini} } @article {1480, title = {A microfluidic flow analyzer with integrated lensed optical fibers [Discovery to Innovation Accelerator]}, journal = {Biomicrofluidics}, volume = {14}, year = {2020}, pages = {054104}, doi = {10.1063/5.0013250}, url = {https://doi.org/10.1063/5.0013250}, author = {Mohan,A. and Gupta,P. and Nair,A. P. and Prabhakar,A. and Saiyed,T.} } @article {1554, title = {Pitchers of Nepenthes khasiana express several digestive-enzyme encoding genes, harbor mostly fungi and probably evolved through changes in the expression of leaf polarity genes [Next Gen Genomics Facility]}, journal = {BMC Plant Biol}, volume = {20}, year = {2020}, month = {2020 Nov 17}, pages = {524}, abstract = {

BACKGROUND: A structural phenomenon seen in certain lineages of angiosperms that has captivated many scholars including Charles Darwin is the evolution of plant carnivory. Evidently, these structural features collectively termed carnivorous syndrome, evolved to aid nutritional acquisition from attracted, captured and digested prey. We now understand why plant carnivory evolved but how carnivorous plants acquired these attributes remains a mystery. In an attempt to understand the evolution of Nepenthes pitcher and to shed more light on its role in prey digestion, we analyzed the transcriptome data of the highly specialized Nepenthes khasiana leaf comprising the leaf base lamina, tendril and the different parts/zones of the pitcher tube viz. digestive zone, waxy zone and lid.

RESULTS: In total, we generated around 262 million high-quality Illumina reads. Reads were pooled, normalized and de novo assembled to generate a reference transcriptome of about 412,224 transcripts. We then estimated transcript abundance along the N. khasiana leaf by mapping individual reads from each part/zone to the reference transcriptome. Correlation-based hierarchical clustering analysis of 27,208 commonly expressed genes indicated functional relationship and similar cellular processes underlying the development of the leaf base and the pitcher, thereby implying that the Nepenthes pitcher is indeed a modified leaf. From a list of 2386 differentially expressed genes (DEGs), we identified transcripts encoding key enzymes involved in prey digestion and protection against pathogen attack, some of which are expressed at high levels in the digestive zone. Interestingly, many of these enzyme-encoding genes are also expressed in the unopened N. khasiana pitcher. Transcripts showing homology to both bacteria and fungi were also detected; and in the digestive zone, fungi are more predominant as compared to bacteria. Taking cues from histology and scanning electron microscopy (SEM) photomicrographs, we found altered expressions of key regulatory genes involved in leaf development. Of particular interest, the expression of class III HOMEODOMAIN-LEUCINE ZIPPER (HD-ZIPIII) and ARGONAUTE (AGO) genes were upregulated in the tendril.

CONCLUSIONS: Our findings suggest that N. khasiana pitchers employ a wide range of enzymes for prey digestion and plant defense, harbor microbes and probably evolved through altered expression of leaf polarity genes.

}, issn = {1471-2229}, doi = {10.1186/s12870-020-02663-2}, author = {Dkhar, Jeremy and Bhaskar, Yogendra Kumar and Lynn, Andrew and Pareek, Ashwani} } @article {1627, title = {RNA-binding proteins La and HuR cooperatively modulate translation repression of PDCD4 mRNA [Mass Spectrometry Facility - Proteomics]}, journal = {J Biol Chem .}, volume = {doi: 10.1074/jbc.RA120.014894. Online ahead of print.}, year = {2020}, month = {12/2020}, abstract = {

Post-transcriptional regulation of gene expression plays a critical role in controlling the inflammatory response. An uncontrolled inflammatory response results in chronic inflammation, often leading to tumorigenesis. Programmed cell death 4 (PDCD4) is a pro-inflammatory tumor-suppressor gene which helps to prevent the transition from chronic inflammation to cancer. PDCD4 mRNA translation is regulated by an interplay between the oncogenic microRNA miR-21 and the RNA-binding protein (RBP) HuR in response to LPS stimulation, but the role of other regulatory factors remain unknown. Here we report that the RBP Lupus antigen (La) interacts with the 3{\textquoteright}UTR of PDCD4 mRNA and prevents miR-21-mediated translation repression. While LPS causes nuclear-cytoplasmic translocation of HuR, it enhances cellular La expression. Remarkably, La and HuR were found to bind cooperatively to the PDCD4 mRNA and mitigate miR-21-mediated translation repression. The cooperative action of La and HuR reduced cell proliferation and enhanced apoptosis, reversing the pro-oncogenic function of miR-21. Together, these observations demonstrate a cooperative interplay between two RBPs, triggered differentially by the same stimulus, which exerts a synergistic effect on PDCD4 expression and thereby helps maintain a balance between inflammation and tumorigenesis.

}, author = {Kumar, Ravi and Poria, Dipak Kumar and Ray, Partho Sarothi} } @article {1422, title = {Structural Characterization of an Exopolysaccharide Isolated from Enterococcus faecalis, and Study on its Antioxidant Activity, and Cytotoxicity Against HeLa Cells [Mass Spectrometry - Glycomics]}, journal = {Curr Microbiol}, year = {2020}, month = {2020 Jul 28}, abstract = {

An exopolysaccharide (EPS-I) having the molecular weight ~ 2.6 {\texttimes} 10\ Da, was isolated from a Zinc resistant strain of Enterococcus faecalis from costal area. The exopolysaccharide consists of D-mannose, D-glucose, and L-fucose in molar ratio of 9:4:1. The monosaccharide units in the EPS-1 were determined through chemical (total acid hydrolysis and methylation analysis) and spectroscopic (FTIR and H NMR experiment) analysis. The mannose-rich EPS-1 showed total antioxidant activity (1\ mg\ mL of EPS-I as functional as approximately to 500 {\textpm} 5.2\ {\textmu}M of ascorbic acid) and Fe metal ion chelation activity (EC = 405.6\ {\textmu}g\ mL) and hydroxyl radical scavenging activity (EC = 219.5\ {\textmu}g\ mL). The in vitro cytotoxicity experiment of EPS-I against cervical carcinoma cell line, HeLa cells showed strong cytotoxic effect (LC = 267.3\ {\textmu}g\ mL) and at that concentration, it found almost nontoxic against normal healthy cells (HEK-293).

}, issn = {1432-0991}, doi = {10.1007/s00284-020-02130-z}, author = {Choudhuri, Indranil and Khanra, Kalyani and Pariya, Prasenjit and Maity, Gajendra Nath and Mondal, Soumitra and Pati, Bikas Ranjan and Bhattacharyya, Nandan} } @article {1524, title = {Tracking the time-dependent and tissue-specific processes of arsenic accumulation and stress responses in rice (Oryza sativa L.) [Mass Spectrometry - Metabolomics Facility]}, journal = {Journal of Hazardous Materials}, year = {2020}, pages = {124307}, abstract = {

The present study analysed time (0.5h to 24h) and tissue [roots, old leaves (OL) and young leaves (YL)] dependent nature of arsenic (As) accumulation and ensuing responses in two contrasting varieties of rice (Oryza sativa L.); Pooja (tolerant) and CO-50 (moderately sensitive). Arsenic accumulation was 5.4-, 4.7- and 7.3-fold higher at 24h in roots, OL and YL, respectively of var. CO-50 than that in var. Pooja. Arsenic accumulation in YL depicted a delayed accumulation; at 2h onwards in var. Pooja (0.23{\textmu}gg-1 dw) while at 1h onwards in var. CO50 (0.26{\textmu}gg-1 dw). The responses of oxidative stress parameters, antioxidant enzymes, metabolites and ions were also found to be tissue- and time-dependent and depicted differential pattern in the two varieties. Among hormone, salicylic acid and abscisic acid showed variable response in var. Pooja and var. CO-50. Metabolite analysis depicted an involvement of various metabolites in As stress responses of two varieties. In conclusion, an early sensing of the As stress, proper coordination of hormones, biochemical responses, ionic and metabolic profiles allowed var. Pooja to resist As stress and reduce As accumulation more effectively as compared to that of var. CO-50.

}, keywords = {Arsenic, Hormones, Metabolites, ROS, Signalling, Stress}, issn = {0304-3894}, doi = {https://doi.org/10.1016/j.jhazmat.2020.124307}, url = {http://www.sciencedirect.com/science/article/pii/S0304389420322974}, author = {Poonam Yadav and Sudhakar Srivastva and Tanmayi Patil and Rishiraj Raghuvanshi and Ashish K. Srivastava and Penna Suprasanna} } @article {1145, title = {Aspartic protease from Aspergillus niger: Molecular characterization and interaction with pepstatin A [Mass Spectrometry Facility - Proteomics].}, journal = {Int J Biol Macromol}, year = {2019}, month = {2019 Jul 30}, abstract = {

In the pursuit of industrial aspartic proteases, aspergillopepsin A-like endopeptidase from the fungi Aspergillus niger, was identified and cultured by solid state fermentation. Conventional chromatographic techniques were employed to purify the extracellular aspartic protease to apparent homogeneity. The enzyme was found to have single polypeptide chain with a molecular mass of 50 {\textpm} 0.5 kDa. The optimum pH and temperature for the purified aspartic protease was found to be 3.5 and 60 {\textdegree}C respectively. The enzyme was stable for 60 min at 50 {\textdegree}C. The purified enzyme had specific activity of 40,000 {\textpm} 1800 U/mg. The enzyme had 85\% homology with the reported aspergillopepsin A-like aspartic endopeptidase from Aspergillus niger CBS 513.88, based on tryptic digestion and peptide analysis. Pepstatin A reversibly inhibited the enzyme with a K value of 0.045 μM. Based on homology modeling and predicted secondary structure, it was inferred that the aspartic protease is rich in β-structures, which was also confirmed by CD measurements. Interaction of pepstatin A with the enzyme did not affect the conformation of the enzyme as evidenced by CD and fluorescence measurements. Degree of hydrolysis of commercial substrates indicated the order of cleaving ability of the enzyme to be hemoglobin \> defatted soya flour \> gluten \> gelatin \> skim milk powder. The enzyme also improved the functional characteristics of defatted soya flour. This aspartic protease was found to be an excellent candidate for genetic manipulation for biotechnological application in food and feed industries, due to its high catalytic turn over number and thermostability.

}, issn = {1879-0003}, doi = {10.1016/j.ijbiomac.2019.07.133}, author = {Purushothaman, Kavya and Bhat, Sagar Krishna and Singh, Sridevi Annapurna and Marathe, Gopal Kedihithlu and Appu Rao, Appu Rao G} } @article {1061, title = {Differentiating Human Induced Pluripotent Stem Cells (iPSCs) Into Lung Epithelial Cells. [Eyestem, a C-CAMP Startup]}, journal = {Curr Protoc Stem Cell Biol}, year = {2019}, month = {2019 Apr 18}, pages = {e86}, abstract = {

Human induced pluripotent\ stem cells\ (hiPSCs) not only offer great opportunities for the study of human development but also have tremendous potential for future clinical\ cell-based therapies.\ The protocol outlined here is used to differentiate hiPSCs into lung epithelial cell types through a process that faithfully recapitulates the stepwise events observed in vivo. From pluripotency, cells are differentiated to a definitive endoderm fate, followed by progression into anteriorized foregut endoderm that has the ability to give rise to both proximal and distal epithelial cells. Furthermore, this methodology allows for the study of lung dysfunction and disease modeling using patient-derived cells, as well as high-throughput pharmacological screening and eventually personalized therapies. Recently we were able to reproduce this protocol using the working cell bank of an hiPSC line made under current Good Manufacturing Practice (cGMP) conditions, a necessary step for the future clinical application of these\ cells. {\textcopyright} 2019 by John Wiley \& Sons, Inc.

}, issn = {1938-8969}, doi = {10.1002/cpsc.86}, author = {Surendran, Harshini and Rajamoorthy, Mohanapriya and Pal, Rajarshi} } @article {1138, title = {Discovery of a deeply divergent new lineage of vine snake (Colubridae: Ahaetuliinae: Proahaetulla gen. nov.) from the southern Western Ghats of Peninsular India with a revised key for Ahaetuliinae [Next Gen Genomics Facility].}, journal = {PLoS One}, volume = {14}, year = {2019}, month = {2019}, pages = {e0218851}, abstract = {

The Western Ghats are well known as a biodiversity hotspot, but the full extent of its snake diversity is yet to be uncovered. Here, we describe a new genus and species of vine snake Proahaetulla antiqua gen. et sp. nov., from the Agasthyamalai hills in the southern Western Ghats. It was found to be a member of the Ahaetuliinae clade, which currently comprises the arboreal snake genera Ahaetulla, Dryophiops, Dendrelaphis and Chrysopelea, distributed in South and Southeast Asia. Proahaetulla shows a sister relationship with all currently known taxa belonging to the genus Ahaetulla, and shares ancestry with Dryophiops. In addition to its phylogenetic position and significant genetic divergence, this new taxon is also different in morphology from members of Ahaetuliinae in a combination of characters, having 12-13 partially serrated keels on the dorsal scale rows, 20 maxillary teeth and 3 postocular scales. Divergence dating reveals that the new genus is ancient, dating back to the Mid-Oligocene, and is one of the oldest persisting monotypic lineages of snakes in the Western Ghats. This discovery adds to the growing list of ancient lineages endemic to the Agasthyamalai hills and underscores the biogeographic significance of this isolated massif in the southern Western Ghats.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0218851}, author = {Mallik, Ashok Kumar and Achyuthan, N Srikanthan and Ganesh, Sumaithangi R and Pal, Saunak P and Vijayakumar, S P and Shanker, Kartik} } @article {986, title = {Enhancement of the gut barrier integrity by a microbial metabolite through the Nrf2 pathway [Discovery to Innovation Accelerator]}, journal = {Nat Commun}, volume = {10}, year = {2019}, month = {2019 Jan 09}, pages = {89}, abstract = {

The importance of gut microbiota in human health and pathophysiology is undisputable. Despite the abundance of metagenomics data, the functional dynamics of gut microbiota in human health and disease remain elusive. Urolithin A (UroA), a major microbial metabolite derived from polyphenolics of berries and pomegranate fruits displays anti-inflammatory, anti-oxidative, and anti-ageing activities. Here, we show that UroA and its potent synthetic analogue (UAS03) significantly enhance gut barrier function and inhibit unwarranted inflammation. We demonstrate that UroA and UAS03 exert their barrier functions through activation of aryl hydrocarbon receptor (AhR)- nuclear factor erythroid 2-related factor 2 (Nrf2)-dependent pathways to upregulate epithelial tight junction proteins. Importantly, treatment with these compounds attenuated colitis in pre-clinical models by remedying barrier dysfunction in addition to anti-inflammatory activities. Cumulatively, the results highlight how microbial metabolites provide two-pronged beneficial activities at gut epithelium by enhancing barrier functions and reducing inflammation to protect from colonic diseases.

}, issn = {2041-1723}, doi = {10.1038/s41467-018-07859-7}, author = {Singh, Rajbir and Chandrashekharappa, Sandeep and Bodduluri, Sobha R and Baby, Becca V and Hegde, Bindu and Kotla, Niranjan G and Hiwale, Ankita A and Saiyed, Taslimarif and Patel, Paresh and Vijay-Kumar, Matam and Langille, Morgan G I and Douglas, Gavin M and Cheng, Xi and Rouchka, Eric C and Waigel, Sabine J and Dryden, Gerald W and Alatassi, Houda and Zhang, Huang-Ge and Haribabu, Bodduluri and Vemula, Praveen K and Jala, Venkatakrishna R} } @article {1201, title = {Overexpression of native Musa-miR397 enhances plant biomass without compromising abiotic stress tolerance in banana [Mass Spectrometry - Metabolomics Facility].}, journal = {Sci Rep}, volume = {9}, year = {2019}, month = {2019 Nov 11}, pages = {16434}, abstract = {

Plant micro RNAs (miRNAs) control growth, development and stress tolerance but are comparatively unexplored in banana, whose cultivation is threatened by abiotic stress and nutrient deficiencies. In this study, a native Musa-miR397 precursor harboring 11 copper-responsive GTAC motifs in its promoter element was identified from banana genome. Musa-miR397 was significantly upregulated (8-10) fold in banana roots and leaves under copper deficiency, correlating with expression of root copper deficiency marker genes such as Musa-COPT and Musa-FRO2. Correspondingly, target laccases were significantly downregulated (\>-2 fold), indicating miRNA-mediated silencing for Cu salvaging. No significant expression changes in the miR397-laccase module were observed under iron stress. Musa-miR397 was also significantly upregulated (\>2 fold) under ABA, MV and heat treatments but downregulated under NaCl stress, indicating universal stress-responsiveness. Further, Musa-miR397 overexpression in banana significantly increased plant growth by 2-3 fold compared with wild-type but did not compromise tolerance towards Cu deficiency and NaCl stress. RNA-seq of transgenic and wild type plants revealed modulation in expression of 71 genes related to diverse aspects of growth and development, collectively promoting enhanced biomass. Summing up, our results not only portray Musa-miR397 as a candidate for enhancing plant biomass but also highlight it at the crossroads of growth-defense trade-offs.

}, issn = {2045-2322}, doi = {10.1038/s41598-019-52858-3}, author = {Patel, Prashanti and Yadav, Karuna and Srivastava, Ashish Kumar and Suprasanna, Penna and Ganapathi, Thumballi Ramabhatta} } @article {1232, title = {Phosphoproteomic analysis reveals Akt isoform-specific regulation of cytoskeleton proteins in human temporal lobe epilepsy with hippocampal sclerosis [Mass Spectrometry - Proteomics]}, journal = {Neurochem Int}, year = {2019}, month = {2019 Dec 26}, pages = {104654}, abstract = {

Akt is one of the most important downstream effectors of phosphatidylinositol 3-kinase/mTOR pathway. Hyper activation and expression of this pathway are shown in a variety of neurological disorders including human temporal lobe epilepsy with hippocampal sclerosis (TLE-HS). Nevertheless, the expression and activation profiles of the Akt isoforms, Akt1, Akt2, and Akt3 and their functional roles in human TLE-HS have not been studied. We examined the protein expression and activation (phosphorylation) patterns of Akt and its isoforms in human hippocampal tissue from TLE and non-TLE patients. A phosphoproteomic approach followed by interactome analysis of each Akt isoform was used to understand protein-protein interactions and their role in TLE-HS pathology. Our results demonstrated activation of the Akt/mTOR pathway as well as activation of Akt downstream substrates like GSK3β, mTOR, and S6 in TLE-HS samples. Akt1 isoform levels were significantly increased in the TLE-HS samples as compared to the non-TLE samples. Most importantly, different isoforms were activated in different TLE-HS samples, Akt2 was activated in three samples, Akt2 and Akt1 were simultaneously activated in one sample and Akt3 was activated in two samples. Our phosphoproteomic screen across six TLE-HS samples identified 183 proteins phosphorylated by Akt isoforms, 29 of these proteins belong to cytoskeletal modification. Also, we were able to identify proteins of several other classes involved in glycolysis, neuronal development, protein folding and excitatory amino acid transport functions as Akt substrates. Taken together, our data offer clues to understand the role of Akt and its isoforms in underlying the pathology of TLE-HS and further, modulation of Akt/mTOR pathway using Akt isoforms specific inhibitors may offer a new therapeutic window for treatment of human TLE-HS.

}, issn = {1872-9754}, doi = {10.1016/j.neuint.2019.104654}, author = {Valmiki, Rajesh Ramanna and Venkatesalu, Subhashini and Chacko, Ari George and Prabhu, Krishna and Thomas, Maya Mary and Mathew, Vivek and Yoganathan, Sangeetha and Muthusamy, Karthik and Chacko, Geeta and Vanjare, Harshad Arvind and Krothapalli, Srinivasa Babu} } @article {1178, title = {Serotonin is essential for eye regeneration in planaria Schmidtea~mediterranea [Mass Spectrometry (Metabolomics) and Central Imaging \& Flow Cytometry Facilities (INT)]}, journal = {FEBS Lett}, year = {2019}, month = {2019 Sep 17}, abstract = {

Planaria is an ideal system to study factors involved in regeneration and tissue homeostasis. Little is known about the role of metabolites and small molecules in stem cell maintenance and lineage specification in planarians. Using liquid chromatography and mass spectrometry (LC-MS)-based quantitative metabolomics, we determined the relative levels of metabolites in stem cells, progenitors, and differentiated cells of the planarian Schmidtea\ mediterranea. Tryptophan and its metabolic product serotonin are significantly enriched in stem cells and progenitor population. Serotonin biosynthesis in these cells is brought about by a noncanonical enzyme, phenylalanine hydroxylase. Knockdown of Smed-pah leads to complete disappearance of eyes in regenerating planaria, while exogenous supply of serotonin and its precursor rescues the eyeless phenotype. Our results demonstrate a key role for serotonin in eye regeneration.

}, issn = {1873-3468}, doi = {10.1002/1873-3468.13607}, author = {Sarkar, Arunabha and Mukundan, Namita and Sowndarya, Sai and Dubey, Vinay Kumar and Babu, Rosana and Lakshmanan, Vairavan and Rangiah, Kannan and Panicker, Mitradas M and Palakodeti, Dasaradhi and Subramanian, Sabarinath Peruvemba and Subramanian, Ramaswamy} } @article {1016, title = {Serum biomarkers identification by iTRAQ and verification by MRM: S100A8/S100A9 levels predict tumor-stroma involvement and prognosis in Glioblastoma [Mass Spectrometry - Metabolomics Facility].}, journal = {Sci Rep}, volume = {9}, year = {2019}, month = {2019 Feb 26}, pages = {2749}, abstract = {

Despite advances in biology and treatment modalities, the prognosis of glioblastoma (GBM) remains poor. Serum reflects disease macroenvironment and thus provides a less invasive means to diagnose and monitor a diseased condition. By employing 4-plex iTRAQ methodology, we identified 40 proteins with differential abundance in GBM sera. The high abundance of serum S100A8/S100A9 was verified by multiple reaction monitoring (MRM). ELISA and MRM-based quantitation showed a significant positive correlation. Further, an integrated investigation using stromal, tumor purity and cell type scores demonstrated an enrichment of myeloid cell lineage in the GBM tumor microenvironment. Transcript levels of S100A8/S100A9 were found to be independent poor prognostic indicators in GBM. Medium levels of pre-operative and three-month post-operative follow-up serum S100A8 levels predicted poor prognosis in GBM patients who lived beyond median survival. In vitro experiments showed that recombinant S100A8/S100A9 proteins promoted integrin signalling dependent glioma cell migration and invasion up to a threshold level of concentrations. Thus, we have discovered GBM serum marker by iTRAQ and verified by MRM. We also demonstrate interplay between tumor micro and macroenvironment and identified S100A8 as a potential marker with diagnostic and prognostic value in GBM.

}, issn = {2045-2322}, doi = {10.1038/s41598-019-39067-8}, author = {Arora, Anjali and Patil, Vikas and Kundu, Paramita and Kondaiah, Paturu and Hegde, A S and Arivazhagan, A and Santosh, Vani and Pal, Debnath and Somasundaram, Kumaravel} } @article {1202, title = {Small molecule modulator of aggrephagy regulates neuroinflammation to curb pathogenesis of neurodegeneration [Discovery to Innovation Accelerator (INT)].}, journal = {EBioMedicine}, year = {2019}, month = {2019 Nov 11}, abstract = {

BACKGROUND: Plethora of efforts fails to yield a single drug to reverse the pathogenesis of Parkinson{\textquoteright}s disease (PD) and related α-synucleopathies.

METHODS: Using chemical biology, we identified a small molecule inhibitor of c-abl kinase, PD180970 that could potentially clear the toxic protein aggregates. Genetic, molecular, cell biological and immunological assays were performed to understand the mechanism of action. In vivo preclinical disease model of PD was used to assess its neuroprotection efficacy.

FINDINGS: In this report, we show the ability of a small molecule inhibitor of tyrosine kinases, PD180970, to induce autophagy (cell lines and mice midbrain) in an mTOR-independent manner and ameliorate the α-synuclein mediated toxicity. PD180970 also exerts anti-neuroinflammatory potential by inhibiting the release of proinflammatory cytokines such as IL-6 (interleukin-6) and MCP-1 (monocyte chemoattractant protein-1) through reduction of TLR-4 (toll like receptor-4) mediated NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) activation. In vivo studies show that PD180970 is neuroprotective by degrading the toxic protein oligomers through induction of autophagy and subsiding the microglial activation.

INTERPRETATION: These protective mechanisms ensure the negation of Parkinson{\textquoteright}s disease related motor impairments. FUND: This work was supported by Wellcome Trust/DBT India Alliance Intermediate Fellowship (500159-Z-09-Z), DST-SERB grant (EMR/2015/001946), DBT (BT/INF/22/SP27679/2018) and JNCASR intramural funds to RM, and SERB, DST (SR/SO/HS/0121/2012) to PAA, and DST-SERB (SB/YS/LS-215/2013) to JPC and BIRAC funding to ETA C-CAMP.

}, issn = {2352-3964}, doi = {10.1016/j.ebiom.2019.10.036}, author = {Sn, Suresh and Pandurangi, Janhavi and Murumalla, Ravi and Dj, Vidyadhara and Garimella, Lakshmi and Acharya, Achyuth and Rai, Shashank and Paul, Abhik and Yarreiphang, Haorei and Pillai, Malini S and Giridharan, Mridhula and Clement, James P and Alladi, Phalguni Anand and Saiyed, Taslimarif and Manjithaya, Ravi} } @article {1007, title = {SOD1 activity threshold and TOR signalling modulate VAP(P58S) aggregation via reactive oxygen species-induced proteasomal degradation in a model of amyotrophic lateral sclerosis. [High Throughput Screening Facility]}, journal = {Dis Model Mech}, volume = {12}, year = {2019}, month = {2019 Feb 07}, abstract = {

Familial amyotrophic lateral sclerosis (ALS) is an incurable, late-onset motor neuron disease, linked strongly to various causative genetic loci. codes for a missense mutation, P56S, in VAMP-associated protein B (VAPB) that causes the protein to misfold and form cellular aggregates. Uncovering genes and mechanisms that affect aggregation dynamics would greatly help increase our understanding of the disease and lead to potential therapeutics. We developed a quantitative high-throughput S2R+ cell-based kinetic assay coupled with fluorescent microscopy to score for genes involved in the modulation of aggregates of the fly orthologue, VAP(P58S), fused with GFP. A targeted RNA interference screen against 900 genes identified 150 hits that modify aggregation, including the ALS loci and (also known as ), as well as genes belonging to the mTOR pathway. Further, a system to measure the extent of VAP(P58S) aggregation in the larval brain was developed in order to validate the hits from the cell-based screen. In the larval brain, we find that reduction of SOD1 levels or decreased mTOR signalling reduces aggregation, presumably by increasing the levels of cellular reactive oxygen species (ROS). The mechanism of aggregate clearance is, primarily, proteasomal degradation, which appears to be triggered by an increase in ROS. We have thus uncovered an interesting interplay between SOD1, ROS and mTOR signalling that regulates the dynamics of VAP aggregation. Mechanistic processes underlying such cellular regulatory networks will lead to better understanding of the initiation and progression of ALS.This article has an associated First Person interview with the first author of the paper.

}, issn = {1754-8411}, doi = {10.1242/dmm.033803}, author = {Chaplot, Kriti and Pimpale, Lokesh and Ramalingam, Balaji and Deivasigamani, Senthilkumar and Kamat, Siddhesh S and Ratnaparkhi, Girish S} } @article {1135, title = {Spoiled for Choice: Diverse Endocytic Pathways Function at the Cell Surface [BLiSC - NCBS, inStem, C-CAMP]}, journal = {Annu Rev Cell Dev Biol}, year = {2019}, month = {2019 Jul 05}, abstract = {

Endocytosis has long been identified as a key cellular process involved in bringing in nutrients, in clearing cellular debris in tissue, in the regulation of signaling, and in maintaining cell membrane compositional homeostasis. While clathrin-mediated endocytosis has been most extensively studied, a number of clathrin-independent endocytic pathways are continuing to be delineated. Here we provide a current survey of the different types of endocytic pathways available at the cell surface and discuss a new classification and plausible molecular mechanisms for some of the less characterized pathways. Along with an evolutionary perspective of the origins of some of these pathways, we provide an appreciation of the distinct roles that these pathways play in various aspects of cellular physiology, including the control of signaling and membrane tension. Expected final online publication date for the Volume 35 is October 7, 2019. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

}, issn = {1530-8995}, doi = {10.1146/annurev-cellbio-100617-062710}, author = {Thottacherry, Joseph Jose and Sathe, Mugdha and Prabhakara, Chaitra and Mayor, Satyajit} } @article {1175, title = {Stromal cells downregulate miR-23a-5p to activate protective autophagy in acute myeloid leukemia [Next Gen Genomics Facility (INT)].}, journal = {Cell Death Dis}, volume = {10}, year = {2019}, month = {2019 Sep 30}, pages = {736}, abstract = {

Complex molecular cross talk between stromal cells and the leukemic cells in bone marrow is known to contribute significantly towards drug-resistance. Here, we have identified the molecular events that lead to stromal cells mediated therapy-resistance in acute myeloid leukemia (AML). Our work demonstrates that stromal cells downregulate miR-23a-5p levels in leukemic cells to protect them from the chemotherapy induced apoptosis. Downregulation of miR-23a-5p in leukemic cells leads to upregulation of protective autophagy by targeting TLR2 expression. Further, autophagy inhibitors when used as adjuvants along with conventional drugs can improve drug sensitivity in vitro as well in vivo in a mouse model of leukemia. Our work also demonstrates that this mechanism of bone marrow stromal cell mediated regulation of miR-23a-5p levels and subsequent molecular events are relevant predominantly in myeloid leukemia. Our results illustrate the critical and dynamic role of the bone marrow microenvironment in modulating miRNA expression in leukemic cells which could contribute significantly to drug resistance and subsequent relapse, possibly through persistence of minimal residual disease in this environment.

}, issn = {2041-4889}, doi = {10.1038/s41419-019-1964-8}, author = {Ganesan, Saravanan and Palani, Hamenth Kumar and Lakshmanan, Vairavan and Balasundaram, Nithya and Alex, Ansu Abu and David, Sachin and Venkatraman, Arvind and Korula, Anu and George, Biju and Balasubramanian, Poonkuzhali and Palakodeti, Dasaradhi and Vyas, Neha and Mathews, Vikram} } @article {1084, title = {The transcriptome enables the identification of candidate genes behind medicinal value of Drumstick tree (Moringa oleifera) [Next Gen Genomics Facility (INT)].}, journal = {Genomics}, year = {2019}, month = {2019 Apr 29}, abstract = {

Moringa oleifera is a plant well-known for its nutrition value, drought resistance and medicinal properties. cDNA libraries from five different tissues (leaf, root, stem, seed and flower) of M. oleifera cultivar Bhagya were generated and sequenced. We developed a bioinformatics pipeline to assemble transcriptome, along with the previously published M. oleifera genome, to predict 17,148 gene models. Few candidate genes related to biosynthesis of secondary metabolites, vitamins and ion transporters were identified. Expressions were further confirmed by real-time quantitative PCR experiments for few promising leads. Quantitative estimation of metabolites, as well as elemental analysis, was also carried out to support our observations. Enzymes in the biosynthesis of vitamins and metabolites like quercetin and kaempferol are highly expressed in leaves, flowers and seeds. The expression of iron transporters and calcium storage proteins were observed in root and leaves. In general, leaves retain the highest amount of small molecules of interest.

}, issn = {1089-8646}, doi = {10.1016/j.ygeno.2019.04.014}, author = {Pasha, Shaik Naseer and Shafi, K Mohamed and Joshi, Adwait G and Meenakshi, Iyer and Harini, K and Mahita, Jarjapu and Sajeevan, Radha Sivarajan and Karpe, Snehal D and Ghosh, Pritha and Nitish, Sathyanarayanan and Gandhimathi, A and Mathew, Oommen K and Prasanna, Subramanian Hari and Malini, Manoharan and Mutt, Eshita and Naika, Mahantesha and Ravooru, Nithin and Rao, Rajas M and Shingate, Prashant N and Sukhwal, Anshul and Sunitha, Margaret S and Upadhyay, Atul K and Vinekar, Rithvik S and Sowdhamini, Ramanathan} } @article {1020, title = {Transcriptomics analysis of propiconazole-treated Cochliobolus sativus reveals new putative azole targets in the plant pathogen [Next Gen Genomics Facility].}, journal = {Funct Integr Genomics}, year = {2019}, month = {2019 Mar 06}, abstract = {

Cochliobolus sativus (anamorph: Bipolaris sorokiniana) is a filamentous fungus from the class Dothideomycetes. It is a pathogen of cereals including wheat and barley, and causes foliar spot blotch, root rot, black point on grains, head blight, leaf blight, and seedling blight diseases. Annual yields of these economically important cereals are severely reduced due to this pathogen attack. Evolution of fungicide resistant pathogen strains, availability of a limited number of potent antifungal compounds, and their efficacy are the acute issues in field management of phytopathogenic fungi. Propiconazole is a widely used azole fungicide to control the disease in fields. The known targets of azoles are the demethylase enzymes involved in ergosterol biosynthesis. Nonetheless, azoles have multiple modes of action, some of which have not been explored yet. Identifying the off-target effects of fungicides by dissecting gene expression profiles in response to them can provide insights into their modes of action and possible mechanisms of fungicide resistance. Moreover it can also reveal additional targets for development of new fungicides. Hence, we analyzed the global gene expression profile of C. sativus on exposure to sub-lethal doses of propiconazole in a time series. The gene expression patterns were confirmed using quantitative reverse transcriptase PCR (qRT-PCR). This study revealed overexpression of target genes from the sterol biosynthesis pathway supporting the reported mode of resistance against azoles. In addition, some new potential targets have also been identified, which could be explored to develop new fungicides and plant protection strategies.

}, issn = {1438-7948}, doi = {10.1007/s10142-019-00660-9}, author = {Somani, Deepika and Adhav, Ragini and Prashant, Ramya and Kadoo, Narendra Y} } @article {993, title = {Complete assembly of a dengue virus type 3 genome from a recent genotype III clade by metagenomic sequencing of serum.[Next Gen Genomics Facility (INT)]}, journal = {Wellcome Open Res}, volume = {3}, year = {2018}, month = {2018}, pages = {44}, abstract = {

Mosquito-borne flaviviruses, such as dengue and Japanese encephalitis virus (JEV), cause life-threatening diseases, particularly in the tropics. Here we performed unbiased metagenomic sequencing of RNA extracted from the serum of four patients and the plasma of one patient, all hospitalized at a tertiary care centre in South India with severe or prolonged febrile illness, together with the serum from one healthy control, in 2014. We identified and assembled a complete dengue virus type 3 sequence from a case of severe dengue fever. We also identified a small number of JEV sequences in the serum of two adults with febrile illness, including one with severe dengue. Phylogenetic analysis revealed that the dengue sequence belonged to genotype III. It has an estimated divergence time of 13.86 years from the most highly related Indian strains. In total, 11 amino acid substitutions were predicted for this strain in the antigenic envelope protein, when compared to the parent strain used for development of the first commercial dengue vaccine.\  We demonstrate that both genome assembly and detection of a low number of viral sequences are possible through the unbiased sequencing of clinical material. These methods may help ascertain causal agents for febrile illnesses with no known cause.

}, issn = {2398-502X}, doi = {10.12688/wellcomeopenres.14438.2}, author = {Dias, Mary and Pattabiraman, Chitra and Siddappa, Shilpa and Gowda, Malali and Shet, Anita and Smith, Derek and Muehlemann, Barbara and Tamma, Krishnapriya and Solomon, Tom and Jones, Terry and Krishna, Sudhir} } @article {686, title = {Exploiting a water network to achieve enthalpy-driven, bromodomain-selective BET inhibitors}, journal = {Bioorganic \& Medicinal Chemistry}, volume = {26}, year = {2018}, pages = {25 - 36}, issn = {0968-0896}, doi = {https://doi.org/10.1016/j.bmc.2017.10.042}, url = {http://www.sciencedirect.com/science/article/pii/S0968089617315948}, author = {William R. Shadrick and Peter J. Slavish and Sergio C. Chai and Brett Waddell and Michele Connelly and Jonathan A. Low and Cynthia Tallant and Brandon M. Young and Nagakumar Bharatham and Stefan Knapp and Vincent A. Boyd and Marie Morfouace and Martine F. Roussel and Taosheng Chen and Richard E. Lee and R. Kiplin Guy and Anang A. Shelat and Philip M. Potter} } @article {708, title = {First report of the characterization of a snake venom apyrase (Ruviapyrase) from Indian Russell{\textquoteright}s viper (Daboia russelii) venom. [Mass Spectromety Facility - Proteomics]}, journal = {Int J Biol Macromol}, volume = {111}, year = {2018}, month = {2018 Jan 08}, pages = {639-648}, abstract = {

A novel apyrase from Russell{\textquoteright}s viper venom (RVV) was purified and characterized, and it was named Ruviapyrase (Russell{\textquoteright}s viper apyrase). It is a high molecular weight (79.4 kDa) monomeric glycoprotein that contains 2.4\% neutral sugars and 58.4\% N-linked oligosaccharides and strongly binds to Concanavalin A. The LC-MS/MS analysis did not identify any protein in NCBI protein database, nevertheless some de novo sequences of Ruviapyrase showed putative conserved domain of apyrase superfamily. Ruviapyrase hydrolysed adenosine triphosphate (ATP) to a significantly greater extent (p \< .05) as compared to adenosine diphosphate (ADP); however, it was devoid of 5{\textquoteright}-nucleotidase and phosphodiesterase activities. The Km and Vmax values for Ruviapyrase towards ATP were 2.54 μM and 615 μM of Pi released min-1, respectively with a turnover number (Kcat) of 24,600 min-1. Spectrofluorometric analysis demonstrated interaction of Ruviapyrase with ATP and ADP at Kd values of 0.92 nM and 1.25 nM, respectively. Ruviapyrase did not show cytotoxicity against breast cancer (MCF-7) cells and haemolytic activity, it exhibited marginal anticoagulant and strong antiplatelet activity, and dose-dependently reversed the ADP-induced platelet aggregation. The catalytic activity and platelet deaggregation property of Ruviapyrase was significantly inhibited by EDTA, DTT and IAA, and neutralized by commercial monovalent and polyvalent antivenom.

}, issn = {1879-0003}, doi = {10.1016/j.ijbiomac.2018.01.038}, author = {Kalita, Bhargab and Patra, Aparup and Jahan, Shagufta and Mukherjee, Ashis K} } @article {1062, title = {Generation of Transplantable Retinal Pigmented Epithelial (RPE) Cells for Treatment of Age-Related Macular Degeneration (AMD). [Eyestem, a C-CAMP Startup]}, journal = {Methods Mol Biol}, year = {2018}, month = {2018 Jun 13}, abstract = {

Age-related macular degeneration (AMD) is the foremost cause of blindness in people over the age of 60 worldwide. Clinically, this disease starts with distortion in central vision eventually leading to legal blindness. Vision loss has a significant impact on quality of life and incurs a substantial cost to the economy. Furthermore, AMD is a complex and progressive neurodegenerative disorder that triggers visual impairment due to the loss of retinal pigmented epithelium (RPE) and the light-sensitive photoreceptors that they support, protect and provide nutrition. Currently, there is no curative treatment for the most common form of this disease, i.e., dry AMD. A novel approach to treat AMD involves the transplantation of RPE cells derived from human induced pluripotent stem cells (iPSCs) in the outer retina. These iPSC-derived RPE cells not only show characteristics similar to native RPE but also could replace as well as regenerate damaged pathologic RPE and produce supportive growth factors and cytokines. Several clinical trials are being conducted taking advantage of a variety of cell- and tissue engineering-based approaches. Here, we present a simple, cost effective, and scalable cell-culture model for generation of purified RPE thus providing the foundation for developing an allogeneic cell therapy for AMD.

}, issn = {1940-6029}, doi = {10.1007/7651_2018_140}, author = {Surendran, Harshini and Rathod, Reena J and Pal, Rajarshi} } @article {765, title = {Identification of multiple isomeric core chitobiose-modified high-mannose and paucimannose -glycans in the planarian. [Mass Spectrometry Facility - Glycomics (INT)]}, journal = {J Biol Chem}, volume = {293}, year = {2018}, month = {2018 May 04}, pages = {6707-6720}, abstract = {

Cell surface-associated glycans mediate many cellular processes, including adhesion, migration, signaling, and extracellular matrix organization. The galactosylation of core fucose (GalFuc epitope) in paucimannose and complex-type -glycans is characteristic of protostome organisms, including flatworms (planarians). Although uninvestigated, the structures of these glycans may play a role in planarian regeneration. Whole-organism MALDI-MS analysis of -linked oligosaccharides from the planarian revealed the presence of multiple isomeric high-mannose and paucimannose structures with unusual mono-, di-, and polygalactosylated ( = 3-5) core fucose structures; the latter structures have not been reported in other systems. Di- and trigalactosylated core fucoses were the most dominant glycomers. -Glycans showed extensive, yet selective, methylation patterns, ranging from non-methylated to polymethylated glycoforms. Although the majority of glycoforms were polymethylated, a small fraction also consisted of non-methylated glycans. Remarkably, monogalactosylated core fucose remained unmethylated, whereas its polygalactosylated forms were methylated, indicating structurally selective methylation. Using database searches, we identified two potential homologs of the Galβ1-4Fuc-synthesizing enzyme from nematodes (GALT-1) that were expressed in the prepharyngeal, pharyngeal, and mesenchymal regions in The presence of two GALT-1 homologs suggests different requirements for mono- and polygalactosylation of core fucose for the formation of multiple isomers. Furthermore, we observed variations in core fucose glycosylation patterns in different planarian strains, suggesting evolutionary adaptation in fucose glycosylation. The various core chitobiose modifications and methylations create \>60 different glycoforms in These results contribute greatly to our understanding of -glycan biosynthesis and suggest the presence of a GlcNAc-independent biosynthetic pathway in

}, issn = {1083-351X}, doi = {10.1074/jbc.RA117.000782}, author = {Subramanian, Sabarinath Peruvemba and Babu, Ponnusamy and Palakodeti, Dasaradhi and Subramanian, Ramaswamy} } @article {1015, title = {Mechanochemical feedback control of dynamin independent endocytosis modulates membrane tension in adherent cells. [Microfluidics and Microfabrication Facility (INT)]}, journal = {Nat Commun}, volume = {9}, year = {2018}, month = {2018 10 11}, pages = {4217}, abstract = {

Plasma membrane tension regulates many key cellular processes. It is modulated by, and can modulate, membrane trafficking. However, the cellular pathway(s) involved in this interplay is poorly understood. Here we find that, among a number of endocytic processes operating simultaneously at the cell surface, a dynamin independent pathway, the CLIC/GEEC (CG) pathway, is rapidly and specifically upregulated upon a sudden reduction of tension. Moreover, inhibition (activation) of the CG pathway results in lower (higher) membrane tension. However, alteration in membrane tension does not directly modulate CG endocytosis. This requires vinculin, a mechano-transducer recruited to focal adhesion in adherent cells. Vinculin acts by controlling the levels of a key regulator of the CG pathway, GBF1, at the plasma membrane. Thus, the CG pathway directly regulates membrane tension and is in turn controlled via a mechano-chemical feedback inhibition, potentially leading to homeostatic regulation of membrane tension in adherent cells.

}, keywords = {Animals, Biomechanical Phenomena, Cell Adhesion, Cell Membrane, Dynamins, Endocytosis, Feedback, Physiological, Mechanotransduction, Cellular, Mice, Signal Transduction, Temperature, Vinculin}, issn = {2041-1723}, doi = {10.1038/s41467-018-06738-5}, author = {Thottacherry, Joseph Jose and Kosmalska, Anita Joanna and Kumar, Amit and Vishen, Amit Singh and Elosegui-Artola, Alberto and Pradhan, Susav and Sharma, Sumit and Singh, Parvinder P and Guadamillas, Marta C and Chaudhary, Natasha and Vishwakarma, Ram and Trepat, Xavier and Del Pozo, Miguel A and Parton, Robert G and Rao, Madan and Pullarkat, Pramod and Roca-Cusachs, Pere and Mayor, Satyajit} } @article {764, title = {Nitrothiophene carboxamides, a novel narrow spectrum antibacterial series: Mechanism of action and Efficacy [Bugworks Res. Pvt. Ltd., a C-CAMP Startup]}, journal = {Sci Rep}, volume = {8}, year = {2018}, month = {2018 May 08}, pages = {7263}, abstract = {

The mechanism of efflux is a tour-de-force in the bacterial armoury that has thwarted the development of novel antibiotics. We report the discovery of a novel chemical series with potent antibacterial properties that was engineered to overcome efflux liability. Compounds liable to efflux specifically via the Resistance Nodulation and cell Division (RND) pump, AcrAB-TolC were chosen for a hit to lead progression. Using structure-based design, the compounds were optimised to lose their binding to the efflux pump, thereby making them potent on wild-type bacteria. We discovered these compounds to be pro-drugs that require activation in E. coli by specific bacterial nitroreductases NfsA and NfsB. Hit to lead chemistry led to the generation of compounds that were potent on wild-type and multi-drug resistant clinical isolates of E. coli, Shigella spp., and Salmonella spp. These compounds are bactericidal and efficacious in a mouse thigh infection model.

}, issn = {2045-2322}, doi = {10.1038/s41598-018-25407-7}, author = {Hameed P, Shahul and Bharatham, Nagakumar and Katagihallimath, Nainesh and Sharma, Sreevalli and Nandishaiah, Radha and Shanbhag, Anirudh P and Thomas, Teby and Narjari, Riya and Sarma, Maitrayee and Bhowmik, Purnendu and Amar, Prakruthi and Ravishankar, Rajani and Jayaraman, Ramesh and Muthan, Kubendran and Subbiah, Ramesh and Ramachandran, Vasanthi and Balasubramanian, V and Datta, Santanu} } @article {865, title = {Prevention of pesticide-induced neuronal dysfunction and mortality with nucleophilic poly-Oxime topical gel. [BIG Grant Supported Entrepreneur]}, journal = {Sci Adv}, volume = {4}, year = {2018}, month = {2018 Oct}, pages = {eaau1780}, abstract = {

Organophosphate-based pesticides inhibit acetylcholinesterase (AChE), which plays a pivotal role in neuromuscular function. While spraying in the field, farmworkers get exposed to pesticides through the dermal route. Internalized pesticide inhibits AChE, which leads to neurotoxicity, cardiotoxicity, cognitive dysfunction, loss of endurance, and death in severe cases. Here, we present a nucleophilic pyridine-2-aldoxime-functionalized chitosan-based topical gel (-Oxime gel) that rapidly deactivates organophosphates, methyl parathion (MPT), on the skin of rats, which leads to reduced AChE inhibition in the blood and tissues. Testing the robustness of -Oxime gel, we report reduction in AChE inhibition following repeated dermal administration of MPT in the presence of -Oxime gel. Furthermore, -Oxime gel prevented MPT-induced neuromuscular dysfunction, loss of endurance, and locomotor coordination. We observe a 100\% survival in rats following topical MPT administration in the presence of -Oxime gel. This prophylactic gel may therefore help farmworkers by limiting pesticide-induced toxicity and mortality.

}, issn = {2375-2548}, doi = {10.1126/sciadv.aau1780}, author = {Thorat, Ketan and Pandey, Subhashini and Chandrashekharappa, Sandeep and Vavilthota, Nikitha and Hiwale, Ankita A and Shah, Purna and Sreekumar, Sneha and Upadhyay, Shubhangi and Phuntsok, Tenzin and Mahato, Manohar and Mudnakudu-Nagaraju, Kiran K and Sunnapu, Omprakash and Vemula, Praveen K} } @article {815, title = {Quantification of Neurotransmitters from Intact and Regenerating Planarians Using UHPLC-MS/SRM Method. [Mass Spectrometry - Metabolomics Facility (INT)]}, journal = {Methods Mol Biol}, volume = {1774}, year = {2018}, month = {2018}, pages = {555-570}, abstract = {

Freshwater planarian species S. mediterranea is an emerging stem cell model because of its capability of regenerating large portions of missing body parts. It is one of the best model systems available to address the basic biological mechanisms in the regeneration processes. Absolute quantification of metabolites from planarians is imperative to understand their role in the regeneration processes. Here we describe a stable isotope dilution ultrahigh performance liquid chromatography/mass spectrometry/selected reaction monitoring (UHPLC-MS/SRM) assay for a sensitive and quantitative assessment of neurotransmitters (NTs) in planaria. We used this method for the simultaneous quantification of 16 NTs from both intact and regenerating planarians.

}, issn = {1940-6029}, doi = {10.1007/978-1-4939-7802-1_25}, author = {Rangiah, Kannan and Palakodeti, Dasaradhi} } @article {814, title = {Recovery of Five Complete Influenza A(H1N1)pdm09 Genome Sequences from the 2015 Influenza Outbreak in India by Metagenomic Sequencing. [Next Gen Genomics Facility (INT)]}, journal = {Genome Announc}, volume = {6}, year = {2018}, month = {2018 Jun 28}, abstract = {

Five complete (H1N1)pdm09 viral sequences were recovered from hospitalized individuals during the 2015 influenza outbreak by metagenomic sequencing. Four of the genomes are from oropharyngeal swabs, and one is from an isolate. All five sequences belong to an emerging 6B clade. Studying them further is critical for outbreak preparedness.

}, issn = {2169-8287}, doi = {10.1128/genomeA.00511-18}, author = {Dash, Paban Kumar and Pattabiraman, Chitra and Tandel, Kundan and Sharma, Shashi and Kumar, Jyoti S and Siddappa, Shilpa and Gowda, Malali and Krishna, Sudhir and Parida, Manmohan} } @article {8903, title = {Comparative sperm protein profiling in bulls differing in fertility and identification of phosphatidylethanolamine-binding protein 4, a potential fertility marker [Mass Spectrometry - Proteomics]}, journal = {Andrology}, volume = {5}, year = {2017}, month = {2017 Sep}, pages = {1032-1051}, abstract = {

This study aimed to identify sperm proteomic signatures regulating sperm functions and fertility by: (i) comparing the sperm electrophoretic protein profiles and identifying the differentially abundant proteins among breeding bulls differing in fertility status and (ii) elucidating the possible role of one of the identified novel proteins, PEBP4 on sperm function and fertility. The grouping of bulls as fertile (n\ =\ 6) and low fertile (n\ =\ 6) was performed based on bull fertility index and infertile (n\ =\ 6) based on semen rejection rate (\>33\%). The sperm motility, fructolysis index, acrosomal reaction, intracellular calcium levels, and seminal plasma fructose and calcium levels were studied among fertility groups. The differentially expressed sperm proteins observed in single- and two-dimensional gel electrophoresis (2DE) were identified using Nano-LC-MS/MS. In the fertile bulls, the expression levels of calmodulin (CALM1), spermadhesinZ13 (SPADH2), and phosphatidylethanolamine-binding protein 4 (PEBP4) were significantly (p\ \<\ 0.05) higher than in other fertility groups. In bovine, expression of PEBP4 a novel seminal protein was not observed in spermatozoa of infertile bulls. When the bulls were grouped based on the presence (n\ =\ 8) or absence (n\ =\ 10) of PEBP4 protein in spermatozoa, a positive significant (p\ \<\ 0.05) association of this protein with the percentage of motile, type-A spermatozoa, and sperm fructose uptake was observed. Further, PEBP4 was localized in elongated spermatids, Leydig cells, excurrent duct system, and principal piece of spermatozoa. These findings suggest a crucial role for the PEBP4 protein in spermiogenesis, epididymal sperm maturation, and sperm motility. This first study in bovine indicates the positive association of PEBP4 in regulating sperm maturation, functions, and fertility and could be a potential marker for predicting semen quality and fertility.

}, keywords = {Acrosome Reaction, Animals, Biomarkers, Calcium Signaling, Cattle, Cattle Diseases, Epididymis, Fertility, Fructose, Infertility, Male, Male, Phosphatidylethanolamine Binding Protein, Proteome, Semen, Sperm Maturation, Sperm Motility, Spermatozoa}, issn = {2047-2927}, doi = {10.1111/andr.12404}, author = {Somashekar, L and Selvaraju, S and Parthipan, S and Patil, S K and Binsila, B K and Venkataswamy, M M and Karthik Bhat, S and Ravindra, J P} } @article {627, title = {Developmentally regulated higher-order chromatin interactions orchestrate B cell fate commitment.}, journal = {Nucleic Acids Res}, year = {2017}, month = {2017 Aug 17}, abstract = {

Genome organization in 3D nuclear-space is important for regulation of gene expression. However, the alterations of chromatin architecture that impinge on the B cell-fate choice of multi-potent progenitors are still unclear. By integrating in situ Hi-C analyses with epigenetic landscapes and genome-wide expression profiles, we tracked the changes in genome architecture as the cells transit from a progenitor to a committed state. We identified the genomic loci that undergo developmental switch between A and B compartments during B-cell fate determination. Furthermore, although, topologically associating domains (TADs) are stable, a significant number of TADs display structural alterations that are associated with changes in cis-regulatory interaction landscape. Finally, we demonstrate the potential roles for Ebf1 and its downstream factor, Pax5, in chromatin reorganization and transcription regulation. Collectively, our studies provide a general paradigm of the dynamic relationship between chromatin reorganization and lineage-specific gene expression pattern that dictates cell-fate determination.

}, issn = {1362-4962}, doi = {10.1093/nar/gkx722}, author = {Boya, Ravi and Yadavalli, Anurupa Devi and Nikhat, Sameena and Kurukuti, Sreenivasulu and Palakodeti, Dasaradhi and Pongubala, Jagan M R} } @article {8888, title = {Genome-wide changes in microRNA expression during short and prolonged heat stress and recovery in contrasting rice cultivars [Next Gen Genomics Facility]}, journal = {J Exp Bot}, volume = {68}, year = {2017}, month = {2017 Apr 01}, pages = {2399-2412}, abstract = {

MicroRNAs (miRNAs) are known to regulate expression of genes under stress. We report here the deep sequencing of small RNAs expressed during control, short and prolonged heat stress and recovery. Genome-wide identification of miRNAs in tolerant (Nagina 22) and susceptible (Vandana) rice cultivars was performed in 16 samples representing root and shoot of 13-day-old seedlings. The expression profile of miRNAs was analysed in 36 pairwise combinations to identify the genotype-, treatment- and tissue-dependent expression of miRNAs. Small-RNA sequencing of 16 libraries yielded ~271 million high-quality raw sequences; 162 miRNA families were identified. The highly expressed miRNAs in rice tissues were miR166, miR168, miR1425, miR529, mR162, miR1876, and miR1862. Expression of osa-miR1436, osa-miR5076, osa-miR5161, and osa-miR6253 was observed only in stressed tissue of both genotypes indicating their general role in heat stress response. Expression of osa-miR1439, osa-miR1848, osa-miR2096, osa-miR2106, osa-miR2875, osa-miR3981, osa-miR5079, osa-miR5151, osa-miR5484, osa-miR5792, and osa-miR5812 was observed only in Nagina 22 during high temperature, suggesting a specific role of these miRNAs in heat stress tolerance. This study provides details of the repertoire of miRNAs expressed in root and shoot of heat susceptible and tolerant rice genotypes under heat stress and recovery.

}, keywords = {Gene Expression Regulation, Plant, Genome, Plant, High-Throughput Nucleotide Sequencing, Hot Temperature, MicroRNAs, Oryza, RNA, Plant, Seedlings, Sequence Analysis, RNA, Time Factors}, issn = {1460-2431}, doi = {10.1093/jxb/erx111}, author = {Mangrauthia, Satendra K and Bhogireddy, Sailaja and Agarwal, Surekha and Prasanth, Vishnu V and Voleti, S R and Neelamraju, Sarla and Subrahmanyam, Desiraju} } @article {471, title = {Gut microbial degradation of organophosphate insecticides-induces glucose intolerance via gluconeogenesis. [Next Generation Genomics facility]}, journal = {Genome Biol}, volume = {18}, year = {2017}, month = {2017 Jan 24}, pages = {8}, abstract = {

BACKGROUND: Organophosphates are the most frequently and largely applied insecticide in the world due to their biodegradable nature. Gut microbes were shown to degrade organophosphates and cause intestinal dysfunction. The diabetogenic nature of organophosphates was recently reported but the underlying molecular mechanism is unclear. We aimed to understand the role of gut microbiota in organophosphate-induced hyperglycemia and to unravel the molecular mechanism behind this process.

RESULTS: Here we demonstrate a high prevalence of diabetes among people directly exposed to organophosphates in rural India (n = 3080). Correlation and linear regression analysis reveal a strong association between plasma organophosphate residues and HbA1c but no association with acetylcholine esterase was noticed. Chronic treatment of mice with organophosphate for 180\ days confirms the induction of glucose intolerance with no significant change in acetylcholine esterase. Further fecal transplantation and culture transplantation experiments confirm the involvement of gut microbiota in organophosphate-induced glucose intolerance. Intestinal metatranscriptomic and host metabolomic analyses reveal that gut microbial organophosphate degradation produces short chain fatty acids like acetic acid, which induces gluconeogenesis and thereby accounts for glucose intolerance. Plasma organophosphate residues are positively correlated with fecal esterase activity and acetate level of human diabetes.

CONCLUSION: Collectively, our results implicate gluconeogenesis as the key mechanism behind organophosphate-induced hyperglycemia, mediated by the organophosphate-degrading potential of gut microbiota. This study reveals the gut microbiome-mediated diabetogenic nature of organophosphates and hence that the usage of these insecticides should be reconsidered.

}, issn = {1474-760X}, doi = {10.1186/s13059-016-1134-6}, author = {Velmurugan, Ganesan and Ramprasath, Tharmarajan and Swaminathan, Krishnan and Mithieux, Gilles and Rajendhran, Jeyaprakash and Dhivakar, Mani and Parthasarathy, Ayothi and Babu, D D Venkatesh and Thumburaj, Leishman John and Freddy, Allen J and Dinakaran, Vasudevan and Puhari, Shanavas Syed Mohamed and Rekha, Balakrishnan and Christy, Yacob Jenifer and Anusha, Sivakumar and Divya, Ganesan and Suganya, Kannan and Meganathan, Boominathan and Kalyanaraman, Narayanan and Vasudevan, Varadaraj and Kamaraj, Raju and Karthik, Maruthan and Jeyakumar, Balakrishnan and Abhishek, Albert and Paul, Eldho and Pushpanathan, Muthuirulan and Rajmohan, Rajamani Koushick and Velayutham, Kumaravel and Lyon, Alexander R and Ramasamy, Subbiah} } @article {707, title = {Proteomics and antivenomics of Echis carinatus carinatus venom: Correlation with pharmacological properties and pathophysiology of envenomation.}, journal = {Sci Rep}, volume = {7}, year = {2017}, month = {2017 Dec 07}, pages = {17119}, abstract = {

The proteome composition of Echis carinatus carinatus venom (ECV) from India was studied for the first time by tandem mass spectrometry analysis. A total of 90, 47, and 22 distinct enzymatic and non-enzymatic proteins belonging to 15, 10, and 6 snake venom protein families were identified in ECV by searching the ESI-LC-MS/MS data against non-redundant protein databases of Viperidae (taxid 8689), Echis (taxid 8699) and Echis carinatus (taxid 40353), respectively. However, analysis of MS/MS data against the Transcriptome Shotgun Assembly sequences (87 entries) of conger E. coloratus identified only 14 proteins in ECV. Snake venom metalloproteases and snaclecs, the most abundant enzymatic and non-enzymatic proteins, respectively in ECV account for defibrinogenation and the strong in vitro pro-coagulant activity. Further, glutaminyl cyclase, aspartic protease, aminopeptidase, phospholipase B, vascular endothelial growth factor, and nerve growth factor were reported for the first time in ECV. The proteome composition of ECV was well correlated with its biochemical and pharmacological properties and clinical manifestations observed in Echis envenomed patients. Neutralization of enzymes and pharmacological properties of ECV, and immuno-cross-reactivity studies unequivocally point to the poor recognition of \<20 kDa ECV proteins, such as PLA2, subunits of snaclec, and disintegrin by commercial polyvalent antivenom.

}, issn = {2045-2322}, doi = {10.1038/s41598-017-17227-y}, author = {Patra, Aparup and Kalita, Bhargab and Chanda, Abhishek and Mukherjee, Ashis K} } @article {8889, title = {Transcriptional transitions in Alphonso mango (Mangifera indica L.) during fruit development and ripening explain its distinct aroma and shelf life characteristics [Next Gen Genomics Facility]}, journal = {Sci Rep}, volume = {7}, year = {2017}, month = {2017 Aug 18}, pages = {8711}, abstract = {

Alphonso is known as the "King of mangos" due to its unique flavor, attractive color, low fiber pulp and long shelf life. We analyzed the transcriptome of Alphonso mango through Illumina sequencing from seven stages of fruit development and ripening as well as flower. Total transcriptome data from these stages ranged between 65 and 143 Mb. Importantly, 20,755 unique transcripts were annotated and 4,611 were assigned enzyme commission numbers, which encoded 142 biological pathways. These included ethylene and flavor related secondary metabolite biosynthesis pathways, as well as those involved in metabolism of starch, sucrose, amino acids and fatty acids. Differential regulation (p-value <= 0.05) of thousands of transcripts was evident in various stages of fruit development and ripening. Novel transcripts for biosynthesis of mono-terpenes, sesqui-terpenes, di-terpenes, lactones and furanones involved in flavor formation were identified. Large number of transcripts encoding cell wall modifying enzymes was found to be steady in their expression, while few were differentially regulated through these stages. Novel 79 transcripts of inhibitors of cell wall modifying enzymes were simultaneously detected throughout Alphonso fruit development and ripening, suggesting controlled activity of these enzymes involved in fruit softening.

}, keywords = {Cell Wall, Enzyme Inhibitors, Flowers, Fruit, Gene Expression Profiling, Gene Expression Regulation, Developmental, Gene Expression Regulation, Plant, Gene Ontology, Genes, Plant, Glycoside Hydrolases, Mangifera, Odorants, Phylogeny, Reproducibility of Results, RNA, Messenger, Transcription, Genetic, Transcriptome}, issn = {2045-2322}, doi = {10.1038/s41598-017-08499-5}, author = {Deshpande, Ashish B and Anamika, Krishanpal and Jha, Vineet and Chidley, Hemangi G and Oak, Pranjali S and Kadoo, Narendra Y and Pujari, Keshav H and Giri, Ashok P and Gupta, Vidya S} } @article {8890, title = {Transcriptomic resources for the medicinal legume Mucuna pruriens: de novo transcriptome assembly, annotation, identification and validation of EST-SSR markers [Next Gen Genomics Facility]}, journal = {BMC Genomics}, volume = {18}, year = {2017}, month = {2017 May 25}, pages = {409}, abstract = {

BACKGROUND: The medicinal legume Mucuna pruriens (L.) DC. has attracted attention worldwide as a source of the anti-Parkinson{\textquoteright}s drug L-Dopa. It is also a popular green manure cover crop that offers many agronomic benefits including high protein content, nitrogen fixation and soil nutrients. The plant currently lacks genomic resources and there is limited knowledge on gene expression, metabolic pathways, and genetics of secondary metabolite production. Here, we present transcriptomic resources for M. pruriens, including a de novo transcriptome assembly and annotation, as well as differential transcript expression analyses between root, leaf, and pod tissues. We also develop microsatellite markers and analyze genetic diversity and population structure within a set of Indian germplasm accessions.

RESULTS: One-hundred ninety-one million two hundred thirty-three thousand two hundred forty-two\ bp cleaned reads were assembled into 67,561 transcripts with mean length of 626\ bp and N50 of 987\ bp. Assembled sequences were annotated using BLASTX against public databases with over 80\% of transcripts annotated. We identified 7,493 simple sequence repeat (SSR) motifs, including 787 polymorphic repeats between the parents of a mapping population. 134 SSRs from expressed sequenced tags (ESTs) were screened against 23 M. pruriens accessions from India, with 52 EST-SSRs retained after quality control. Population structure analysis using a Bayesian framework implemented in fastSTRUCTURE showed nearly similar groupings as with distance-based (neighbor-joining) and principal component analyses, with most of the accessions clustering per geographical origins. Pair-wise comparison of transcript expression in leaves, roots and pods identified 4,387 differentially expressed transcripts with the highest number occurring between roots and leaves. Differentially expressed transcripts were enriched with transcription factors and transcripts annotated as belonging to secondary metabolite pathways.

CONCLUSIONS: The M. pruriens transcriptomic resources generated in this study provide foundational resources for gene discovery and development of molecular markers. Polymorphic SSRs identified can be used for genetic diversity, marker-trait analyses, and development of functional markers for crop improvement. The results of differential expression studies can be used to investigate genes involved in L-Dopa synthesis and other key metabolic pathways in M. pruriens.

}, keywords = {Data Mining, Expressed Sequence Tags, Gene Expression Profiling, Microsatellite Repeats, Molecular Sequence Annotation, Mucuna, Polymorphism, Genetic, RNA, Messenger, Transcription Factors}, issn = {1471-2164}, doi = {10.1186/s12864-017-3780-9}, author = {Sathyanarayana, N and Pittala, Ranjith Kumar and Tripathi, Pankaj Kumar and Chopra, Ratan and Singh, Heikham Russiachand and Belamkar, Vikas and Bhardwaj, Pardeep Kumar and Doyle, Jeff J and Egan, Ashley N} } @article {706, title = {Dynein Clusters into Lipid Microdomains on Phagosomes to Drive Rapid Transport toward Lysosomes.}, journal = {Cell}, volume = {164}, year = {2016}, month = {2016 Feb 11}, pages = {722-34}, abstract = {

Diverse cellular processes are driven by motor proteins that are recruited to and generate force on lipid membranes. Surprisingly little is known about how membranes control the force from motors and how this may impact specific cellular functions. Here, we show that dynein motors physically cluster into microdomains on the membrane of a phagosome as it matures inside cells. Such geometrical reorganization allows many dyneins within a cluster to generate cooperative force on a single microtubule. This results in rapid directed transport of the phagosome toward microtubule minus ends, likely promoting phagolysosome fusion and pathogen degradation. We show that lipophosphoglycan, the major molecule implicated in immune evasion of Leishmania donovani, inhibits phagosome motion by disrupting the clustering and therefore the cooperative force generation of dynein. These findings appear relevant to several pathogens that prevent phagosome-lysosome fusion by targeting lipid microdomains on phagosomes.

}, keywords = {Animals, Biological Transport, Cell Line, Dictyostelium, Dyneins, Glycosphingolipids, Leishmania donovani, Lysosomes, Membrane Microdomains, Mice, Phagosomes}, issn = {1097-4172}, doi = {10.1016/j.cell.2015.12.054}, author = {Rai, Ashim and Pathak, Divya and Thakur, Shreyasi and Singh, Shampa and Dubey, Alok Kumar and Mallik, Roop} } @article {709, title = {Isolation and Characterization of a Halophilic Cyanobacterium Euhalothece SLVH01 from Sambhar Salt Lake, India. Int.J.Curr.Microbiol.App.Sci (2016) 5(2): 215-224 [Electron Microscopy Facility - TEM]}, year = {2016}, author = {Harshil H. Bhatt and Renu Pasricha and Vivek N. Upasani} } @article {454, title = {Mechanism of apoptosis induction in human breast cancer MCF-7 cell by Ruviprase, a small peptide from Daboia russelii russelii venom.[Mass Spectrometry]}, journal = {Chem Biol Interact}, volume = {258}, year = {2016}, month = {2016 Oct 25}, pages = {297-304}, abstract = {

Ruviprase, a 4.4~kDa peptide isolated from Daboia russelii russelii venom demonstrated antiproliferative activity against EMT6/AR1, U-87MG, HeLa and MCF-7 cancer cells with an IC50 value of 23.0, 8.8, 5.8 and 4.0~μg ml(-1), respectively. However, it was nontoxic to non-cancerous human embryonic kidney cell and human peripheral blood lymphocytes. Flow-cytometric analysis confirmed the apoptosis induction in MCF-7 cells by Ruviprase where it induced DNA condensation but did not cause mitotic blockage or chromosomal aberration in treated-cells. Immunofluorescence microscopic analysis indicated Ruviprase induced apoptosis in MCF-7 cells through p53 and p21-mediated pathways. Ruviprase generated reactive oxygen species (ROS), altered the mitochondrial transmembrane potential, and significantly decreased the cellular glutathione (GSH) content of MCF-7 cells. Immunoblotting and quantitative real-time PCR (qRT-PCR) analyses suggested that Ruviprase down-regulated the expression of anti-apoptotic protein Bcl-2, increased cleavage of poly (ADP-ribose) polymerase (PARP) protein, and up-regulated the expression of pro-apoptotic protein Bax, as well as executer protein caspase-7 to induced apoptosis in MCF-7 cells via intrinsic pathway. This is the first report on the characterization of the anticancer potential of a small, non-toxic and anticoagulant peptide purified from Russell{\textquoteright}s viper venom.

}, issn = {1872-7786}, doi = {10.1016/j.cbi.2016.09.004}, author = {Thakur, Rupamoni and Kini, Sudarshan and Kurkalang, Sillarine and Banerjee, Atanu and Chatterjee, Purba and Chanda, Abhishek and Chatterjee, Anupam and Panda, Dulal and Mukherjee, Ashis K} } @article {455, title = {Unraveling the Proteome Composition and Immuno-profiling of Western India Russell{\textquoteright}s Viper Venom for In-Depth Understanding of Its Pharmacological Properties, Clinical Manifestations, and Effective Antivenom Treatment.[Mass Spectrometry]}, journal = {J Proteome Res}, year = {2016}, month = {2016 Dec 12}, abstract = {

The proteome composition of western India (WI) Russell{\textquoteright}s viper venom (RVV) was correlated with pharmacological properties and pathological manifestations of RV envenomation. Proteins in the 5-19 and 100-110 kDa mass ranges were the most predominate (\~{}35.1\%) and least abundant (\~{}3.4\%) components, respectively, of WI RVV. Non-reduced SDS-PAGE indicated the occurrence of multiple subunits, non-covalent oligomers, self-aggregation, and/or interactions among the RVV proteins. A total of 55 proteins belonging to 13 distinct snake venom families were unambiguously identified by ESI-LC-MS/MS analysis. Phospholipase A2 (32.5\%) and Kunitz-type serine protease inhibitors (12.5\%) represented the most abundant enzymatic and non-enzymatic proteins, respectively. However, ATPase, ADPase, and hyaluronidase, detected by enzyme assays, were not identified by proteomic analysis owing to limitations in protein database deposition. Several biochemical and pharmacological properties of WI RVV were also investigated. Neurological symptoms exhibited by some RV-bite patients in WI may be correlated to the presence of neurotoxic phospholipase A2 enzymes and Kunitz-type serine protease inhibitor complex in this venom. Monovalent antivenom was found to be better than polyvalent antivenom in immuno-recognition and neutralization of the tested pharmacological properties and enzyme activities of WI RVV; nevertheless, both antivenoms demonstrated poor cross-reactivity and neutralization of pharmacological activities shown by low-molecular-mass proteins (<18 kDa) of this venom.

}, issn = {1535-3907}, doi = {10.1021/acs.jproteome.6b00693}, author = {Kalita, Bhargab and Patra, Aparup and Mukherjee, Ashis K} } @article {476, title = {The E3 ligase ube3a is required for learning in Drosophila melanogaster. [Drosophila facility]}, journal = {Biochem Biophys Res Commun}, volume = {462}, year = {2015}, month = {2015 Jun 19}, pages = {71-7}, abstract = {

Angelman syndrome and autism are neurodevelopmental disorders linked to mutations and duplications of an E3 ligase called ube3a respectively. Since cognitive deficits and learning disabilities are hallmark symptoms of both these disorders, we investigated a role for dube3a in the learning ability of flies using the aversive phototaxis suppression assay. We show that down and up-regulation of dube3a are both detrimental to learning in larvae and adults. Using conditional gene expression we found that dube3a is required for normal brain development and during adulthood. Furthermore, we suggest that dube3a could be interacting with other learning and memory genes such as derailed. Along with firmly establishing dube3a as a gene that is required for learning, our work also opens avenues for further understanding the role played by this gene in brain development and behavior.

}, keywords = {Animals, Animals, Genetically Modified, Brain, Drosophila melanogaster, Drosophila Proteins, Gene Expression Regulation, Developmental, Immunoblotting, Larva, Learning, Memory, Motor Activity, Mushroom Bodies, Mutation, Reverse Transcriptase Polymerase Chain Reaction, Time Factors, Ubiquitin-Protein Ligases}, issn = {1090-2104}, doi = {10.1016/j.bbrc.2015.04.110}, author = {Chakraborty, Moumita and Paul, Blesson K and Nayak, Tanmoyita and Das, Aniruddha and Jana, Nihar R and Bhutani, Supriya} } @article {498, title = {Genome sequencing of herb Tulsi (Ocimum tenuiflorum) unravels key genes behind its strong medicinal properties.[Mass spectrometry - Metabolomics]}, journal = {BMC Plant Biol}, volume = {15}, year = {2015}, month = {2015 Aug 28}, pages = {212}, abstract = {

BACKGROUND: Krishna Tulsi, a member of Lamiaceae family, is a herb well known for its spiritual, religious and medicinal importance in India. The common name of this plant is {\textquoteright}Tulsi{\textquoteright} (or {\textquoteright}Tulasi{\textquoteright} or {\textquoteright}Thulasi{\textquoteright}) and is considered sacred by Hindus. We present the draft genome of Ocimum tenuiflurum L (subtype Krishna Tulsi) in this report. The paired-end and mate-pair sequence libraries were generated for the whole genome sequenced with the Illumina Hiseq 1000, resulting in an assembled genome of 374\ Mb, with a genome coverage of 61\ \% (612\ Mb estimated genome size). We have also studied transcriptomes (RNA-Seq) of two subtypes of O. tenuiflorum, Krishna and Rama Tulsi and report the relative expression of genes in both the varieties.

RESULTS: The pathways leading to the production of medicinally-important specialized metabolites have been studied in detail, in relation to similar pathways in Arabidopsis thaliana and other plants. Expression levels of anthocyanin biosynthesis-related genes in leaf samples of Krishna Tulsi were observed to be relatively high, explaining the purple colouration of Krishna Tulsi leaves. The expression of six important genes identified from genome data were validated by performing q-RT-PCR in different tissues of five different species, which shows the high extent of urosolic acid-producing genes in young leaves of the Rama subtype. In addition, the presence of eugenol and ursolic acid, implied as potential drugs in the cure of many diseases including cancer was confirmed using mass spectrometry.

CONCLUSIONS: The availability of the whole genome of O.tenuiflorum and our sequence analysis suggests that small amino acid changes at the functional sites of genes involved in metabolite synthesis pathways confer special medicinal properties to this herb.

}, keywords = {Gene Expression Regulation, Plant, Genome, Plant, India, Ocimum, Plant Leaves, Plants, Medicinal}, issn = {1471-2229}, doi = {10.1186/s12870-015-0562-x}, author = {Upadhyay, Atul K and Chacko, Anita R and Gandhimathi, A and Ghosh, Pritha and Harini, K and Joseph, Agnel P and Joshi, Adwait G and Karpe, Snehal D and Kaushik, Swati and Kuravadi, Nagesh and Lingu, Chandana S and Mahita, J and Malarini, Ramya and Malhotra, Sony and Malini, Manoharan and Mathew, Oommen K and Mutt, Eshita and Naika, Mahantesha and Nitish, Sathyanarayanan and Pasha, Shaik Naseer and Raghavender, Upadhyayula S and Rajamani, Anantharamanan and Shilpa, S and Shingate, Prashant N and Singh, Heikham Russiachand and Sukhwal, Anshul and Sunitha, Margaret S and Sumathi, Manojkumar and Ramaswamy, S and Gowda, Malali and Sowdhamini, Ramanathan} } @article {496, title = {Insulin plant (Costus pictus) extract improves insulin sensitivity and ameliorates atherogenic dyslipidaemia in fructose induced insulin resistant rats: Molecular mechanism [Mass spectrometry - Metabolomics]}, journal = {Journal of Functional Foods}, volume = {17}, year = {2015}, pages = {749 - 760}, abstract = {

Abstract The preventative effect of insulin plant (Costus pictus) leaf extract on hyperglycaemia, insulin sensitivity and dyslipidaemia in fructose-fed insulin resistant rats was investigated in the present study, further the possible underlying mechanism was also explored. It was observed that under insulin resistant condition this plant extract decreases hyperinsulinaemia by improving insulin sensitivity at the peripheral level through its antioxidant and anti-inflammatory effects. At molecular level decreased activation of molecules involved in stress sensitive signalling cascade and down regulation of pro-inflammatory cytokines were noticed. Furthermore the effects of this plant extract on adipokine profile and plasma homocysteine were evaluated. The possible active components responsible for the beneficial effect exerted by this plant extract have also been identified and quantified. To the best of our knowledge this is the first study on insulin plant extract on the aforementioned animal model.

}, keywords = {Insulin plant}, issn = {1756-4646}, doi = {http://dx.doi.org/10.1016/j.jff.2015.06.024}, url = {//www.sciencedirect.com/science/article/pii/S1756464615003187}, author = {S. Ashwini and Zachariah Bobby and Manoj Joseph and Sajini Elizabeth Jacob and Ramamoorthy Padmapriya} } @article {520, title = {Isolation of Exosomes from Blood Plasma: Qualitative and Quantitative Comparison of Ultracentrifugation and Size Exclusion Chromatography Methods. (Mass spectrometry - Proteomics)}, journal = {PLoS One}, volume = {10}, year = {2015}, month = {2015}, pages = {e0145686}, abstract = {

BACKGROUND: Exosomes are emerging targets for biomedical research. However, suitable methods for the isolation of blood plasma-derived exosomes without impurities have not yet been described.

AIM: Therefore, we investigated the efficiency and purity of exosomes isolated with potentially suitable methods; differential ultracentrifugation (UC) and size exclusion chromatography (SEC).

METHODS AND RESULTS: Exosomes were isolated from rat and human blood plasma by various UC and SEC conditions. Efficiency was investigated at serial UC of the supernatant, while in case of SEC by comparing the content of exosomal markers of various fractions. Purity was assessed based on the presence of albumin. We found that the diameter of the majority of isolated particles fell into the size range of exosomes, however, albumin was also present in the preparations, when 1h UC at 4{\textdegree}C was applied. Furthermore, with this method only a minor fraction of total exosomes could be isolated from blood as deduced from the constant amount of exosomal markers CD63 and TSG101 detected after serial UC of rat blood plasma samples. By using UC for longer time or with shorter sedimentation distance at 4{\textdegree}C, or UC performed at 37{\textdegree}C, exosomal yield increased, but albumin impurity was still observed in the isolates, as assessed by transmission electron microscopy, dynamic light scattering and immunoblotting against CD63, TSG101 and albumin. Efficiency and purity were not different in case of using further diluted samples. By using SEC with different columns, we have found that although a minor fraction of exosomes can be isolated without significant albumin content on Sepharose CL-4B or Sephacryl S-400 columns, but not on Sepharose 2B columns, the majority of exosomes co-eluted with albumin.

CONCLUSION: Here we show that it is feasible to isolate exosomes from blood plasma by SEC without significant albumin contamination albeit with low vesicle yield.

}, keywords = {Animals, Chromatography, Gel, Exosomes, Male, Plasma, Rats, Wistar, Ultracentrifugation}, issn = {1932-6203}, doi = {10.1371/journal.pone.0145686}, author = {Baranyai, Tam{\'a}s and Herczeg, Kata and On{\'o}di, Zs{\'o}fia and Voszka, Istv{\'a}n and M{\'o}dos, K{\'a}roly and Marton, Nikolett and Nagy, Gy{\"o}rgy and M{\"a}ger, Imre and Wood, Matthew J and El Andaloussi, Samir and P{\'a}link{\'a}s, Zolt{\'a}n and Kumar, Vikas and Nagy, P{\'e}ter and Kittel, {\'A}gnes and Buz{\'a}s, Edit Ir{\'e}n and Ferdinandy, P{\'e}ter and Giricz, Zolt{\'a}n} } @article {500, title = {Nickel azamacrocyclic complex activated persulphate based oxidative degradation of methyl orange: recovery and reuse of complex using adsorbents [Mass spectrometry - Metabolomics]}, journal = {RSC Adv.}, volume = {5}, year = {2015}, pages = {31716-31724}, abstract = {

Adsorbents are useful for the removal of metal complex based catalysts from the reaction medium. Moreover{,} effective catalysts may be recycled with the use of adsorbents. These facts inspired us to investigate the use of adsorbents for the recovery and reuse of a metal complex that could activate persulphate to effectively degrade an organic pollutant in water. Herein{,} we report the nickel complex (C1) activated persulphate based degradation of methyl orange (MO) in water and the removal of C1 using activated carbon (AC) and Amberlite (Am) as adsorbents. C1 adsorbed onto AC (C1-AC) was reused in the solid form to activate persulphate and degrade MO without leaching C1 into water. Additionally{,} solid C1-Am recovered from the degraded MO solution was ion exchanged using sodium chloride to obtain C1{,} which was reused for MO degradation. The study demonstrates the application of adsorbents such as AC and Am for the adsorptive recovery and reuse of a metal complex based persulphate activator.

}, doi = {10.1039/C5RA03350K}, url = {http://dx.doi.org/10.1039/C5RA03350K}, author = {Subramanian, Gokulakrishnan and Nalawade, Pranav and Hinder, Steven J. and Pillai, Suresh C. and Prakash, Halan} } @article {501, title = {A quantitative metabolomics peek into planarian regeneration. [Mass spectrometry - Metabolomics]}, journal = {Analyst}, volume = {140}, year = {2015}, month = {2015 May 21}, pages = {3445-64}, abstract = {

The fresh water planarian species Schmidtea mediterranea is an emerging stem cell model because of its capability to regenerate a whole animal from a small piece of tissue. It is one of the best model systems to address the basic mechanisms essential for regeneration. Here, we are interested in studying the roles of various amines, thiols and nucleotides in planarian regeneration, stem cell function and growth. We developed mass spectrometry based quantitative methods and validated the differential enrichment of 35 amines, 7 thiol metabolites and 4 nucleotides from both intact and regenerating planarians. Among the amines, alanine in sexual and asparagine in asexual are the highest (\>1000 ng/mg) in the intact planarians. The levels of thiols such as cysteine and GSH are 651 and 1107 ng mg(-1) in planarians. Among the nucleotides, the level of cGMP is the lowest (0.03 ng mg(-1)) and the level of AMP is the highest (187 ng mg(-1)) in both of the planarian strains. We also noticed increasing levels of amines in both anterior and posterior regenerating planarians. The blastema from day 3 regenerating planarians also showed higher amounts of many amines. Interestingly, the thiol (cysteine and GSH) levels are well maintained during planarian regeneration. This suggests an inherent and effective mechanism to control induced oxidative stress because of the robust regeneration and stem cell proliferation. Like in intact planarians, the level of cGMP is also very low in regenerating planarians. Surprisingly, the levels of amines and thiols in head regenerating blastemas are \~{}3 times higher compared to those for tail regenerating blastemas. Thus our results strongly indicate the potential roles of amines, thiols and nucleotides in planarian regeneration.

}, keywords = {Animals, Calibration, Chromatography, High Pressure Liquid, Limit of Detection, Metabolomics, Planarians, Reference Standards, Regeneration, Reproduction, Asexual, Species Specificity, Tandem Mass Spectrometry}, issn = {1364-5528}, doi = {10.1039/c4an02037e}, author = {Natarajan, Nivedita and Ramakrishnan, Padma and Lakshmanan, Vairavan and Palakodeti, Dasaradhi and Rangiah, Kannan} } @article {472, title = {RDGBα, a PtdIns-PtdOH transfer protein, regulates G-protein-coupled PtdIns(4,5)P2 signalling during Drosophila phototransduction.[Drosophila Facility]}, journal = {J Cell Sci}, volume = {128}, year = {2015}, month = {2015 Sep 01}, pages = {3330-44}, abstract = {

Many membrane receptors activate phospholipase C (PLC) during signalling, triggering changes in the levels of several plasma membrane lipids including phosphatidylinositol (PtdIns), phosphatidic acid (PtdOH) and phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2]. It is widely believed that exchange of lipids between the plasma membrane and endoplasmic reticulum (ER) is required to restore lipid homeostasis during PLC signalling, yet the mechanism remains unresolved. RDGBα (hereafter RDGB) is a multi-domain protein with a PtdIns transfer protein (PITP) domain (RDGB-PITPd). We find that, in vitro, the RDGB-PITPd binds and transfers both PtdOH and PtdIns. In Drosophila photoreceptors, which experience high rates of PLC activity, RDGB function is essential for phototransduction. We show that binding of PtdIns to RDGB-PITPd is essential for normal phototransduction; however, this property is insufficient to explain the in vivo function because another Drosophila PITP (encoded by vib) that also binds PtdIns cannot rescue the phenotypes of RDGB deletion. In RDGB mutants, PtdIns(4,5)P2 resynthesis at the plasma membrane following PLC activation is delayed and PtdOH levels elevate. Thus RDGB couples the turnover of both PtdIns and PtdOH, key lipid intermediates during G-protein-coupled PtdIns(4,5)P2 turnover.

}, keywords = {Animals, Drosophila melanogaster, Drosophila Proteins, Eye Proteins, Light Signal Transduction, Membrane Proteins, Phosphatidic Acids, Phosphatidylinositol 4,5-Diphosphate, Type C Phospholipases}, issn = {1477-9137}, doi = {10.1242/jcs.173476}, author = {Yadav, Shweta and Garner, Kathryn and Georgiev, Plamen and Li, Michelle and Gomez-Espinosa, Evelyn and Panda, Aniruddha and Mathre, Swarna and Okkenhaug, Hanneke and Cockcroft, Shamshad and Raghu, Padinjat} } @article {684, title = {Role of Homothorax in region specific regulation of Deformed in embryonic neuroblasts.}, journal = {Mech Dev}, volume = {138 Pt 2}, year = {2015}, month = {2015 Nov}, pages = {190-7}, abstract = {

The expression and regulation of Hox genes in developing central nervous system (CNS) lack important details like specific cell types where Hox genes are expressed and the transcriptional regulatory players involved in these cells. In this study we have investigated the expression and regulation of Drosophila Hox gene Deformed (Dfd) in specific cell types of embryonic CNS. Using Dfd neural autoregulatory enhancer we find that Dfd autoregulates itself in cells of mandibular neuromere. We have also investigated the role of a Hox cofactor Homothorax (Hth) for its role in regulating Dfd expression in CNS. We find that Hth exhibits a region specific role in controlling the expression of Dfd, but has no direct role in mandibular Dfd neural autoregulatory circuit. Our results also suggest that homeodomain of Hth is not required for regulating Dfd expression in embryonic CNS.

}, keywords = {Animals, Central Nervous System, Drosophila, Drosophila Proteins, Enhancer Elements, Genetic, Gene Expression Regulation, Developmental, Genes, Homeobox, Homeodomain Proteins, Neural Stem Cells, Organogenesis}, issn = {1872-6356}, doi = {10.1016/j.mod.2015.09.003}, author = {Kumar, Raviranjan and Chotaliya, Maheshvari and Vuppala, Sruthakeerthi and Auradkar, Ankush and Palasamudrum, Kalyani and Joshi, Rohit} } @article {518, title = {Vertebrate Hedgehog is secreted on two types of extracellular vesicles with different signaling properties. (Mass spectrometry - Proteomics)}, journal = {Sci Rep}, volume = {4}, year = {2014}, month = {2014 Dec 08}, pages = {7357}, abstract = {

Hedgehog (Hh) is a secreted morphogen that elicits differentiation and patterning in developing tissues. Multiple proposed mechanisms to regulate Hh dispersion includes lipoprotein particles and exosomes. Here we report that vertebrate Sonic Hedgehog (Shh) is secreted on two types of extracellular-vesicles/exosomes, from human cell lines and primary chick notochord cells. Although largely overlapping in size as estimated from electron micrographs, the two exosomal fractions exhibited distinct protein and RNA composition. We have probed the functional properties of these vesicles using cell-based assays of Hh-elicited gene expression. Our results suggest that while both Shh-containing exo-vesicular fractions can activate an ectopic Gli-luciferase construct, only exosomes co-expressing Integrins can activate endogenous Shh target genes HNF3β and Olig2 during the differentiation of mouse ES cells to ventral neuronal progenitors. Taken together, our results demonstrate that primary vertebrate cells secrete Shh in distinct vesicular forms, and support a model where packaging of Shh along with other signaling proteins such as Integrins on exosomes modulates target gene activation. The existence of distinct classes of Shh-containing exosomes also suggests a previously unappreciated complexity for fine-tuning of Shh-mediated gradients and pattern formation.

}, keywords = {Animals, Chick Embryo, Exosomes, Extracellular Space, Hedgehog Proteins, HEK293 Cells, Humans, MicroRNAs, Models, Biological, Protein Transport, Signal Transduction, Vertebrates}, issn = {2045-2322}, doi = {10.1038/srep07357}, author = {Vyas, Neha and Walvekar, Ankita and Tate, Dhananjay and Lakshmanan, Vairavan and Bansal, Dhiru and Lo Cicero, Alessandra and Raposo, Graca and Palakodeti, Dasaradhi and Dhawan, Jyotsna} } @article {503, title = {Comprehensive analysis of neurotransmitters from regenerating planarian extract using an ultrahigh-performance liquid chromatography/mass spectrometry/selected reaction monitoring method.}, journal = {Rapid Commun Mass Spectrom}, volume = {27}, year = {2013}, month = {2013 Nov 15}, pages = {2439-52}, abstract = {

RATIONALE: Absolute quantification of neurotransmitters (NTs) from biological systems is imperative to track how changes in concentration of active neurochemicals may affect biological behavior. A sensitive method for the absolute quantification of multiple NTs in a single method is highly needed.

METHODS: A stable-isotope dilution ultrahigh-performance liquid chromatography/mass spectrometry/selected reaction monitoring (UHPLC/MS/SRM) assay has been developed for a sensitive and quantitative assessment of NTs in planaria. We used this method for the simultaneous quantification of 16 NTs. All analytes showed a linear relationship between concentrations (0.78-50 ng/mL), regression coefficients higher than 0.97, accuracy (91-109\%) and low coefficients of variation (CVs). The inter-day CVs for the lowest quality controls (1.56 ng/mL) were in the range between 2-11\%.

RESULTS: The levels of most of the NTs were similar in both sexual and asexual planarians except for glutamic acid, which was about two-fold higher in asexual compared to sexual planarians. We identified high levels of serotonin and failed to detect tryptamine suggesting that the pathway essential for the conversion of tryptophan into tryptamine is absent in planarians. Interestingly, we also found high levels of dopamine and L-DOPA in regenerating planarians suggesting their possible role in regeneration.

CONCLUSIONS: For the first time, we developed novel methodology based on UHPLC/MS/SRM and quantified 16 NTs with high sensitivity and specificity from sexual and asexual strains of planarian Schmidtea mediterranea. This method will also have great application in quantifying various NTs with great precision in different model systems.

}, keywords = {Animals, Chromatography, High Pressure Liquid, Glutamic Acid, Limit of Detection, Neurotransmitter Agents, Planarians, Reproduction, Reproduction, Asexual, Serotonin, Spectrometry, Mass, Electrospray Ionization, Tryptamines}, issn = {1097-0231}, doi = {10.1002/rcm.6706}, author = {Rangiah, Kannan and Palakodeti, Dasaradhi} } @article {493, title = {Deep sequencing reveals unique small RNA repertoire that is regulated during head regeneration in Hydra magnipapillata. [Next Generation Genomics facility]}, journal = {Nucleic Acids Res}, volume = {41}, year = {2013}, month = {2013 Jan 07}, pages = {599-616}, abstract = {

Small non-coding RNAs such as miRNAs, piRNAs and endo-siRNAs fine-tune gene expression through post-transcriptional regulation, modulating important processes in development, differentiation, homeostasis and regeneration. Using deep sequencing, we have profiled small non-coding RNAs in Hydra magnipapillata and investigated changes in small RNA expression pattern during head regeneration. Our results reveal a unique repertoire of small RNAs in hydra. We have identified 126 miRNA loci; 123 of these miRNAs are unique to hydra. Less than 50\% are conserved across two different strains of Hydra vulgaris tested in this study, indicating a highly diverse nature of hydra miRNAs in contrast to bilaterian miRNAs. We also identified siRNAs derived from precursors with perfect stem-loop structure and that arise from inverted repeats. piRNAs were the most abundant small RNAs in hydra, mapping to transposable elements, the annotated transcriptome and unique non-coding regions on the genome. piRNAs that map to transposable elements and the annotated transcriptome display a ping-pong signature. Further, we have identified several miRNAs and piRNAs whose expression is regulated during hydra head regeneration. Our study defines different classes of small RNAs in this cnidarian model system, which may play a role in orchestrating gene expression essential for hydra regeneration.

}, keywords = {Animals, Gene Expression Regulation, Head, High-Throughput Nucleotide Sequencing, Hydra, MicroRNAs, Regeneration, RNA, Small Interfering, RNA, Small Untranslated, Sequence Analysis, RNA, Transcriptome}, issn = {1362-4962}, doi = {10.1093/nar/gks1020}, author = {Krishna, Srikar and Nair, Aparna and Cheedipudi, Sirisha and Poduval, Deepak and Dhawan, Jyotsna and Palakodeti, Dasaradhi and Ghanekar, Yashoda} } @article {488, title = {Genome sequencing unveils a novel sea enterotoxin-carrying PVL phage in Staphylococcus aureus ST772 from India. [Next Generation Genomics facility]}, journal = {PLoS One}, volume = {8}, year = {2013}, month = {2013}, pages = {e60013}, abstract = {

Staphylococcus aureus is a major human pathogen, first recognized as a leading cause of hospital-acquired infections. Community-associated S. aureus (CA-SA) pose a greater threat due to increase in severity of infection and disease among children and healthy adults. CA-SA strains in India are genetically diverse, among which is the sequence type (ST) 772, which has now spread to Australia, Europe and Japan. Towards understanding the genetic characteristics of ST772, we obtained draft genome sequences of five relevant clinical isolates and studied the properties of their PVL-carrying prophages, whose presence is a defining hallmark of CA-SA. We show that this is a novel prophage, which carries the structural genes of the hlb-carrying prophage and includes the sea enterotoxin. This architecture probably emerged early within the ST772 lineage, at least in India. The sea gene, unique to ST772 PVL, despite having promoter sequence characteristics typical of low expression, appears to be highly expressed during early phase of growth in laboratory conditions. We speculate that this might be a consequence of its novel sequence context. The crippled nature of the hlb-converting prophage in ST772 suggests that widespread mobility of the sea enterotoxin might be a selective force behind its {\textquoteright}transfer{\textquoteright} to the PVL prophage. Wild type ST772 strains induced strong proliferative responses as well as high cytotoxic activity against neutrophils, likely mediated by superantigen SEA and the PVL toxin respectively. Both proliferation and cytotoxicity were markedly reduced in a cured ST772 strain indicating the impact of the phage on virulence. The presence of SEA alongside the genes for the immune system-modulating PVL toxin may contribute to the success and virulence of ST772.

}, keywords = {Bacterial Toxins, Base Sequence, Enterotoxins, Exotoxins, Genome, Bacterial, Hemolysin Proteins, Humans, India, Leukocidins, Molecular Sequence Data, Prophages, RNA, Messenger, Sequence Analysis, DNA, Sphingomyelin Phosphodiesterase, Staphylococcus aureus}, issn = {1932-6203}, doi = {10.1371/journal.pone.0060013}, author = {Prabhakara, Sushma and Khedkar, Supriya and Shambat, Srikanth Mairpady and Srinivasan, Rajalakshmi and Basu, Atanu and Norrby-Teglund, Anna and Seshasayee, Aswin Sai Narain and Arakere, Gayathri} } @article {1010, title = {Light driven ultrafast electron transfer in oxidative redding of Green Fluorescent Proteins. [Protein Technology Facility]}, journal = {Sci Rep}, volume = {3}, year = {2013}, month = {2013}, pages = {1580}, abstract = {

Fluorescent proteins undergoing green to red (G/R) photoconversion have proved to be potential tools for investigating dynamic processes in living cells and for photo-localization nanoscopy. However, the photochemical reaction during light induced G/R photoconversion of fluorescent proteins remains unclear. Here we report the direct observation of ultrafast time-resolved electron transfer (ET) during the photoexcitation of the fluorescent proteins EGFP and mEos2 in presence of electron acceptor, p-benzoquinone (BQ). Our results show that in the excited state, the neutral EGFP chromophore accepts electrons from an anionic electron donor, Glu222, and G/R photoconversion is facilitated by ET to nearby electron acceptors. By contrast, mEos2 fails to produce the red emitting state in the presence of BQ; ET depletes the excited state configuration en route to the red-emitting fluorophore. These results show that ultrafast ET plays a pivotal role in multiple photoconversion mechanisms and provide a method to modulate the G/R photoconversion process.

}, keywords = {Benzoquinones, Electron Transport, Green Fluorescent Proteins, Light, Oxidation-Reduction}, issn = {2045-2322}, doi = {10.1038/srep01580}, author = {Saha, Ranajay and Verma, Pramod Kumar and Rakshit, Surajit and Saha, Suvrajit and Mayor, Satyajit and Pal, Samir Kumar} } @article {720, title = {Antiproliferative property of n-hexane and chloroform extracts of Anisomeles malabarica (L). R. Br. in HPV16-positive human cervical cancer cells.}, journal = {J Pharmacol Pharmacother}, volume = {3}, year = {2012}, month = {2012 Jan}, pages = {26-34}, abstract = {

OBJECTIVES: To find the efficacy of serial extracts of Anisomeles malabarica in inhibiting proliferation of and inducing apoptosis in human cervical cancer cells, SiHa and ME 180, that are HPV 16-positive.

MATERIALS AND METHODS: The whole plant was extracted in n-hexane, chloroform, ethyl acetate, n-butanol, methanol, and water. The cells were treated with the extracts at increasing concentrations to find the IC(50), adopting MTT ([3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide]) assay. Acridine orange (AO) and ethidium bromide (EB) and Hoechst 33258 staining were adopted to assess the mode of cell death, Annexin V-Cy3 staining to evaluate one of the early apoptotic features, JC-1 staining to assess the mitochondrial membrane depolarization, comet assay for DNA fragmentation, and cell cycle analysis for the distribution of cells after treatment.

RESULTS: n-Hexane and chloroform extracts were cytotoxic to the cervical cancer cells in dose- and duration-dependent manner. The cells that responded to the treatments revealed typical apoptotic features. Early features of apoptosis, phosphatidyl serine translocation and loss of mitochondrial trans-membrane potential, were observed in the treated cells, and comet assay revealed DNA damage. In the FACS analysis, the cells accumulated in the sub-G0/G1 phase of the cell cycle, except in n-hexane- and chloroform extract-treated SiHa cells at 24 h, which showed arrest in S- and G2/M phases.

CONCLUSIONS: n-Hexane and chloroform extracts of A. malabarica inhibit proliferation of and induce death in HPV16-positive cervical cancer cells, mostly by apoptosis and to some extent by necrosis.

}, issn = {0976-5018}, doi = {10.4103/0976-500X.92500}, author = {Preethy, Christo Paul and Padmapriya, Ramamoorthy and Periasamy, Vaiyapuri Subbarayan and Riyasdeen, Anvarbatcha and Srinag, Suresh and Krishnamurthy, Hanumanthappa and Alshatwi, Ali Abdullah and Akbarsha, Mohammad Abdulkader} } @article {717, title = {Developmental heterogeneity in DNA packaging patterns influences T-cell activation and transmigration.}, journal = {PLoS One}, volume = {7}, year = {2012}, month = {2012}, pages = {e43718}, abstract = {

Cellular differentiation programs are accompanied by large-scale changes in nuclear organization and gene expression. In this context, accompanying transitions in chromatin assembly that facilitates changes in gene expression and cell behavior in a developmental system are poorly understood. Here, we address this gap and map structural changes in chromatin organization during murine T-cell development, to describe an unusual heterogeneity in chromatin organization and associated functional correlates in T-cell lineage. Confocal imaging of DNA assembly in cells isolated from bone marrow, thymus and spleen reveal the emergence of heterogeneous patterns in DNA organization in mature T-cells following their exit from the thymus. The central DNA pattern dominated in immature precursor cells in the thymus whereas both central and peripheral DNA patterns were observed in na{\"\i}ve and memory cells in circulation. Na{\"\i}ve T-cells with central DNA patterns exhibited higher mechanical pliability in response to compressive loads in vitro and transmigration assays in vivo, and demonstrated accelerated expression of activation-induced marker CD69. T-cell activation was characterized by marked redistribution of DNA assembly to a central DNA pattern and increased nuclear size. Notably, heterogeneity in DNA patterns recovered in cells induced into quiescence in culture, suggesting an internal regulatory mechanism for chromatin reorganization. Taken together, our results uncover an important component of plasticity in nuclear organization, reflected in chromatin assembly, during T-cell development, differentiation and transmigration.

}, keywords = {Animals, Antigens, CD, Antigens, Differentiation, T-Lymphocyte, Bone Marrow Cells, Cell Lineage, Cell Movement, Cell Nucleus, Chromatin, DNA, Hematopoietic Stem Cells, Lectins, C-Type, Lymphocyte Activation, Mice, Microscopy, Confocal, Models, Biological, Models, Statistical, Sequence Analysis, DNA, Spleen, T-Lymphocytes}, issn = {1932-6203}, doi = {10.1371/journal.pone.0043718}, author = {Gupta, Soumya and Marcel, Nimi and Talwar, Shefali and Garg, Megha and R, Indulaxmi and Perumalsamy, Lakshmi R and Sarin, Apurva and Shivashankar, G V} } @article {714, title = {Distinct spatial and molecular features of notch pathway assembly in regulatory T cells.}, journal = {Sci Signal}, volume = {5}, year = {2012}, month = {2012 Jul 24}, pages = {ra53}, abstract = {

Variations in the spatial localization of signaling components and crosstalk among signaling cascades are mechanisms through which diversity in signaling networks is generated. The receptor Notch provides an example of regulation by spatial localization: In the canonical Notch signaling pathway, Notch is cleaved to produce the Notch intracellular domain (NICD, also known as NIC), which translocates to the nucleus to regulate gene expression. We describe a T cell receptor-dependent, non-nuclear distribution and function of the processed receptor Notch, which was associated with the improved survival of regulatory T cells (T(regs)) in vitro and in vivo and was compromised by T cell-specific deletion of Notch1. Unlike a nuclear-restricted mutant of NICD, mutant NICD that underwent nuclear export or was targeted to the plasma membrane protected Notch1(-/-) T(regs) from apoptosis induced by nutrient deprivation and oxidative stress. Notch signaling integrated with phosphatidylinositol 3-kinase signaling and mammalian target of rapamycin complex 2 (mTORC2) for this cell survival function. Biochemical and imaging approaches revealed a membrane-proximal complex containing NICD and the mTORC2 component Rictor, and this complex was stabilized by specific interactions with the Notch ligand Delta-like-1 and mediated the survival of T(regs). Together, our evidence for the spatial control of Notch and the crosstalk of Notch signaling with other pathways reveals coupling between the localization of Notch and diverse intracellular signaling pathways.

}, keywords = {Animals, Apoptosis, Blotting, Western, Carrier Proteins, Cell Line, Cell Membrane, Cell Survival, Flow Cytometry, Gene Knockout Techniques, Humans, Immunoprecipitation, Intercellular Signaling Peptides and Proteins, Mice, Mice, Inbred C57BL, Microscopy, Fluorescence, Phosphatidylinositol 3-Kinase, Rapamycin-Insensitive Companion of mTOR Protein, Receptor Cross-Talk, Receptor, Notch1, Signal Transduction, T-Lymphocytes, Regulatory, Trans-Activators, Transcription Factors}, issn = {1937-9145}, doi = {10.1126/scisignal.2002859}, author = {Perumalsamy, Lakshmi R and Marcel, Nimi and Kulkarni, Sneha and Radtke, Freddy and Sarin, Apurva} } @article {491, title = {Draft genome sequence of Rhodovulum sp. strain PH10, a phototrophic alphaproteobacterium isolated from a soil sample of mangrove of Namkhana, India. [Next Generation Genomics facility]}, journal = {J Bacteriol}, volume = {194}, year = {2012}, month = {2012 Nov}, pages = {6363}, abstract = {

We report the 4.8-Mb draft genome of Rhodovulum sp. strain PH10, a phototrophic bacterium belonging to class Alphaproteobacteria, isolated from a soil sample collected from the mangrove forest of Namkhana in India. This genome is the first from the genus Rhodovulum and will lead to a better understanding of the genes/pathways involved in activities like phototrophic growth and nitrogen fixation in this group of bacteria.

}, keywords = {Genome, Bacterial, India, Molecular Sequence Data, Rhodovulum, Soil Microbiology, Wetlands}, issn = {1098-5530}, doi = {10.1128/JB.01695-12}, author = {Khatri, Indu and Korpole, Suresh and Subramanian, Srikrishna and Pinnaka, Anil Kumar} } @article {495, title = {Draft genome sequence of Staphylococcus aureus 118 (ST772), a major disease clone from India. [Next Generation Genomics facility]}, journal = {J Bacteriol}, volume = {194}, year = {2012}, month = {2012 Jul}, pages = {3727-8}, abstract = {

We report the draft genome sequence of an ST772 Staphylococcus aureus disease isolate carrying staphylococcal cassette chromosome mec (SCCmec) type V from a pyomyositis patient. Our de novo short read assembly is \~{}2.8 Mb and encodes a unique Panton-Valentine leukocidin (PVL) phage with structural genes similar to those of ϕ7247PVL and novel lysogenic genes at the N termini.

}, keywords = {Cloning, Molecular, Genome, Bacterial, India, Molecular Sequence Data, Pyomyositis, Staphylococcal Infections, Staphylococcus aureus}, issn = {1098-5530}, doi = {10.1128/JB.00480-12}, author = {Prabhakara, Sushma and Khedkar, Supriya and Loganathan, Ramya Malarini and Chandana, S and Gowda, Malali and Arakere, Gayathri and Seshasayee, Aswin Sai Narain} } @article {494, title = {Draft genome sequence of Staphylococcus aureus ST672, an emerging disease clone from India. [Next Generation Genomics facility]}, journal = {J Bacteriol}, volume = {194}, year = {2012}, month = {2012 Dec}, pages = {6946-7}, abstract = {

We report the draft genome sequence of methicillin-resistant Staphylococcus aureus (MRSA) strain ST672, an emerging disease clone in India, from a septicemia patient. The genome size is about 2.82 Mb with 2,485 open reading frames (ORFs). The staphylococcal cassette chromosome mec (SCCmec) element (type V) and immune evasion cluster appear to be different from those of strain ST772 on preliminary examination.

}, keywords = {Bacteremia, Bacterial Proteins, Bacterial Typing Techniques, Base Sequence, DNA, Bacterial, Genome, Bacterial, Humans, Methicillin-Resistant Staphylococcus aureus, Molecular Sequence Data, Open Reading Frames, Penicillin-Binding Proteins, Sequence Analysis, DNA, Staphylococcal Infections}, issn = {1098-5530}, doi = {10.1128/JB.01868-12}, author = {Khedkar, Supriya and Prabhakara, Sushma and Loganathan, Ramya Malarini and S, Chandana and Gowda, Malali and Arakere, Gayathri and Seshasayee, Aswin Sai Narain} } @article {490, title = {Genome sequence of the halotolerant bacterium Imtechella halotolerans K1T. [Next Generation Genomics facility]}, journal = {J Bacteriol}, volume = {194}, year = {2012}, month = {2012 Jul}, pages = {3731}, abstract = {

We report the 3.087-Mb genome sequence of Imtechella halotolerans K1(T), isolated from an estuarine water sample collected from Kochi, Kerala, India. Strain K1 was recently reported as a novel genus of the family Flavobacteriaceae.

}, keywords = {Gene Expression Regulation, Bacterial, Genome, Bacterial, Gram-Negative Aerobic Bacteria, Molecular Sequence Data}, issn = {1098-5530}, doi = {10.1128/JB.00506-12}, author = {Kumar, Shailesh and Vikram, Surendra and Subramanian, Srikrishna and Raghava, Gajendra Pal Singh and Pinnaka, Anil Kumar} } @article {489, title = {Genome sequence of the marine bacterium Marinilabilia salmonicolor JCM 21150T. [Next Generation Genomics facility]}, journal = {J Bacteriol}, volume = {194}, year = {2012}, month = {2012 Jul}, pages = {3746}, abstract = {

We report the 4.98-Mb genome sequence of Marinilabilia salmonicolor JCM 21150(T), which was isolated from marine mud in the year 1961. The draft genome of strain Marinilabilia salmonicolor JCM 21150(T) contains 4,982,627 bp with a G+C content of 41.92\% and 4,227 protein coding genes, 52 tRNAs, and 3 rRNAs.

}, keywords = {Bacteria, Gene Expression Regulation, Bacterial, Genome, Bacterial, Molecular Sequence Data}, issn = {1098-5530}, doi = {10.1128/JB.00649-12}, author = {Kumar, Shailesh and Subramanian, Srikrishna and Raghava, Gajendra Pal Singh and Pinnaka, Anil Kumar} } @article {712, title = {An inhibitor of nonhomologous end-joining abrogates double-strand break repair and impedes cancer progression.}, journal = {Cell}, volume = {151}, year = {2012}, month = {2012 Dec 21}, pages = {1474-87}, abstract = {

DNA Ligase IV is responsible for sealing of double-strand breaks (DSBs) during nonhomologous end-joining (NHEJ). Inhibiting Ligase IV could result in amassing of DSBs, thereby serving as a strategy toward treatment of cancer. Here, we identify a molecule, SCR7 that inhibits joining of DSBs in cell-free repair system. SCR7 blocks Ligase IV-mediated joining by interfering with its DNA binding but not that of T4 DNA Ligase or Ligase I. SCR7 inhibits NHEJ in a Ligase IV-dependent manner within cells,\ and activates the intrinsic apoptotic pathway. More importantly, SCR7 impedes tumor progression in mouse models and when coadministered with DSB-inducing therapeutic modalities enhances their sensitivity significantly. This inhibitor to target NHEJ offers a strategy toward the treatment of cancer and improvement of existing regimens.

}, keywords = {Amino Acid Sequence, Animals, Cell Line, Tumor, Disease Models, Animal, DNA Breaks, Double-Stranded, DNA End-Joining Repair, DNA Ligase ATP, DNA Ligases, Drug Design, Drug Resistance, Neoplasm, Humans, Lymphocytes, Lymphoma, Male, Mice, Mice, Inbred BALB C, Mice, Nude, Models, Molecular, Molecular Sequence Data, Neoplasms, Protein Structure, Tertiary, Pyrimidines, Radiation Tolerance, Rats, Schiff Bases, Sequence Alignment}, issn = {1097-4172}, doi = {10.1016/j.cell.2012.11.054}, author = {Srivastava, Mrinal and Nambiar, Mridula and Sharma, Sheetal and Karki, Subhas S and Goldsmith, G and Hegde, Mahesh and Kumar, Sujeet and Pandey, Monica and Singh, Ram K and Ray, Pritha and Natarajan, Renuka and Kelkar, Madhura and De, Abhijit and Choudhary, Bibha and Raghavan, Sathees C} } @article {482, title = {A protein complex network of Drosophila melanogaster. [Drosophila facility]}, journal = {Cell}, volume = {147}, year = {2011}, month = {2011 Oct 28}, pages = {690-703}, abstract = {

Determining the composition of protein complexes is an essential step toward understanding the cell as an integrated system. Using coaffinity purification coupled to mass spectrometry analysis, we examined protein associations involving nearly 5,000 individual, FLAG-HA epitope-tagged Drosophila proteins. Stringent analysis of these data, based on a statistical framework designed to define individual protein-protein interactions, led to the generation of a Drosophila protein interaction map (DPiM) encompassing 556 protein complexes. The high quality of the DPiM and its usefulness as a paradigm for metazoan proteomes are apparent from the recovery of many known complexes, significant enrichment for shared functional attributes, and validation in human cells. The DPiM defines potential novel members for several important protein complexes and assigns functional links to 586 protein-coding genes lacking previous experimental annotation. The DPiM represents, to our knowledge, the largest metazoan protein complex map and provides a valuable resource for analysis of protein complex evolution.

}, keywords = {Animals, Drosophila melanogaster, Drosophila Proteins, Proteasome Endopeptidase Complex, Protein Interaction Mapping, Proteomics, SNARE Proteins}, issn = {1097-4172}, doi = {10.1016/j.cell.2011.08.047}, author = {Guruharsha, K G and Rual, Jean-Fran{\c c}ois and Zhai, Bo and Mintseris, Julian and Vaidya, Pujita and Vaidya, Namita and Beekman, Chapman and Wong, Christina and Rhee, David Y and Cenaj, Odise and McKillip, Emily and Shah, Saumini and Stapleton, Mark and Wan, Kenneth H and Yu, Charles and Parsa, Bayan and Carlson, Joseph W and Chen, Xiao and Kapadia, Bhaveen and VijayRaghavan, K and Gygi, Steven P and Celniker, Susan E and Obar, Robert A and Artavanis-Tsakonas, Spyros} } @article {483, title = {Inositol 1,4,5-trisphosphate receptor and dSTIM function in Drosophila insulin-producing neurons regulates systemic intracellular calcium homeostasis and flight. [Drosophila facility]}, journal = {J Neurosci}, volume = {30}, year = {2010}, month = {2010 Jan 27}, pages = {1301-13}, abstract = {

Calcium (Ca(2+)) signaling is known to regulate the development, maintenance and modulation of activity in neuronal circuits that underlie organismal behavior. In Drosophila, intracellular Ca(2+) signaling by the inositol 1,4,5-trisphosphate receptor and the store-operated channel (dOrai) regulates the formation and function of neuronal circuits that control flight. Here, we show that restoring InsP(3)R activity in insulin-producing neurons of flightless InsP(3)R mutants (itpr) during pupal development can rescue systemic flight ability. Expression of the store operated Ca(2+) entry (SOCE) regulator dSTIM in insulin-producing neurons also suppresses compromised flight ability of InsP(3)R mutants suggesting that SOCE can compensate for impaired InsP(3)R function. Despite restricted expression of wild-type InsP(3)R and dSTIM in insulin-producing neurons, a global restoration of SOCE and store Ca(2+) is observed in primary neuronal cultures from the itpr mutant. These results suggest that restoring InsP(3)R-mediated Ca(2+) release and SOCE in a limited subset of neuromodulatory cells can influence systemic behaviors such as flight by regulating intracellular Ca(2+) homeostasis in a large population of neurons through a non-cell-autonomous mechanism.

}, keywords = {Animals, Calcium, Calcium Signaling, Cell Membrane, Cells, Cultured, Central Nervous System, Drosophila, Drosophila Proteins, Flight, Animal, Homeostasis, Inositol 1,4,5-Trisphosphate Receptors, Insulin, Intracellular Fluid, Membrane Proteins, Mutation, Neural Pathways, Neurons, Pupa, Stromal Interaction Molecule 1}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.3668-09.2010}, author = {Agrawal, Neha and Venkiteswaran, Gayatri and Sadaf, Sufia and Padmanabhan, Nisha and Banerjee, Santanu and Hasan, Gaiti} }