Biology subjects

Dasgupta, S.

Publications and source records attributed to Dasgupta, S..

34 records · Page 2Linked to original sources

bioRxiv2024

Molecular insights into the oligosaccharide binding, recognition and hydrolysis by a fungal exo-beta-(1,3)-glucanase

Carbohydrate hydrolysing enzymes assume special industrial and commercial interest as a source for yielding fermentable glucose especially for the biofuel industry. Among these enzymes, the exo-{beta}-(1,3) glucanases are promising for industrial use as they hydrolyze sugars such as laminarin, a major constituent of the algal cell wall. Exploring the structure and function of these enzymes is of particular interest for the improvement of their functional properties for industrial use. We report the structural and biochemical characterizations of Aspergillus oryzae exo-{beta}-(1,3) glucanase (AoBgl). We have expressed, purified, and performed biochemical characterizations of the recombinant AoBgl. Purified AoBgl is found to hydrolyse {beta}-(1,3)-glycosidic linkages present in the oligosaccharide laminaritriose and the polysaccharide, laminarin effectively while retaining >50% activity at glucose concentrations of around 1.5M. We have determined three high-resolution structures of AoBgl: (a) apo form at 1.75 [A], (b) complexed form with bound disaccharide at 1.73 [A] and (c) glucose-bound form at 1.20 [A]. Sequence analysis and structural comparison indicate that AoBgl belongs to the GH5 sugar hydrolase family. The sugar-bound structures reveal the mode of substrate binding and interactions at the active site of AoBgl. Further, molecular dynamics simulation and mutational studies indicate that AoBgl can effectively bind trisaccharides and higher oligosaccharides. Our biochemical and structural data provide detailed molecular insights into the active site of this GH5 enzyme and would be helpful in the rational engineering of glycosyl hydrolases belonging to similar families for industrial use.

biochemistry↗

bioRxiv2024

A Postdoctoral Training Program in Bioimage Analysis

We herein describe a postdoctoral training program designed to train biologists with microscopy experience in bioimage analysis. We detail the rationale behind the program, the various components of the training program, and outcomes in terms of works produced and the career effects on past participants. We analyze the results of an anonymous survey distributed to past and present participants, indicating overall high value of all 12 rated aspects of the program, but significant heterogeneity in which aspects were most important to each participant. Finally, we propose this model as a template for other programs which may want to train experts in professional skill sets, and discuss the important considerations when running such a program. We believe that such programs can have extremely positive impact for both the trainees themselves and the broader scientific community.

cell biology↗

bioRxiv2024

Single cell sequencing of zebrafish kidney marrows reveals AHR2-dependent endogenous regulation of hematopoiesis

The aryl hydrocarbon receptor (AHR) is a ligand-dependent transcription factor that mediates a wide range of biological and toxicological responses. While largely studied in ligand-activated toxicant responses, AHR also plays important roles in endogenous physiological processes. We leveraged single cell sequencing and an AHR2 knockout zebrafish line to investigate the role of AHR2 in regulating hematopoiesis (production and differentiation of red and white blood cells from hematopoietic stem cells). Our objectives were to determine if absence of AHR2-1) alters proportions of immune cell populations and/or 2) impacts gene expression within individual immune cell types. We dissected kidney marrow (organ of hematopoiesis in zebrafish) from adult wildtype and AHR2 knockout zebrafish (N=3/genotype), isolated single cells and sequenced [~] 5000 cells/sample (10X Genomics). We identified 14 cell clusters representing the expected major blood (erythrocytes, thrombocytes), immune (B cells, macrophages, lymphoid cells, granulocytes, etc), progenitors and kidney cell populations. We focused our analyses only on the progenitor and mature immune cell types. While there were no genotype-specific differences in proportion of individual cell types, gene expression differences were observed within several cell types. For known genes, such as rrm2, changes were up to 2000-fold, signifying their importance in AHR2-hematopoesis interaction. Several of the known genes are also identified as markers of carcinoma cells for an array of cancer types. However, many of the dysregulated genes are poorly annotated, limiting our ability to examine biological processes and pathways dysregulated on AHR2 mutation. Nevertheless, our study indicates that AHR2 plays an important endogenous role in hematopoiesis. Future work will focus on better characterizing anatomy of dysregulated genes and their functions in hematopoiesis.

immunology↗

bioRxiv2024

Experimental type 1 diabetes metabolically rejuvenates CD8+ T cells for improved control of tumor growth through an IGF1-IGF1R axis

AbstractEpidemiological studies suggest that patients with pre-existing type 1 diabetes (T1D) have a decreased risk of developing melanoma, prostate cancer, and breast cancer, although the underlying mechanism remains to be elucidated. In translational modelling, we observed that streptozotocin (STZ) induced T1D mice exhibited restricted melanoma and carcinoma (mammary, lung and colon) growth in association with extended overall survival. Tumor-infiltrating CD8+ T cells were found to be responsible for tumor growth restriction. Tumor infiltrating CD8+ T cells but not tumor cells themselves exhibited higher glycolytic and cytotoxic activities in T1D hosts. Such improved anti-tumor T cell function was linked to selective upregulated expression of insulin-like growth factor 1, insulin-like growth factor 1 receptor, and phospho-mTOR in CD8+ T cells in the TME. T1D patient derived CD8+ T cells displayed superior activation in vitro after tumor antigen stimulation vs. non-diabetic CD8+ T cells. Activation of T1D patient derived CD8+ T cells was sensitive to targeted antagonism of IGF1R and mTOR, supporting the operational involvement of the IGF1R-mTOR signaling axis. Our results suggest that selective activation of the intrinsic IGF1R-mTOR signaling axis in CD8+ T cells represents a preferred endpoint to achieving more effective immunotherapy outcomes and improved cancer patient management. SignificanceExperimental type 1 diabetes decelerates tumor growth through metabolic activation of cytotoxic T cells dependent on an IGF1R-mTOR signaling pathway. CD8+IGF1R+IGF1+ T cells play a crucial role in T1D dependent tumor control.

cancer biology↗

bioRxiv2024

Flame retardant tetrabromobisphenol A (TBBPA) disrupts histone acetylation during zebrafish maternal-to-zygotic transition

3,3,5.5-Tetrabromobisphenol A (TBBPA) is a widely used brominated flame-retardant utilized in the production of electronic devices and plastic paints. The objective of this study is to use zebrafish as a model and determine the effects of TBBPA exposure on early embryogenesis. We initiated TBBPA exposures (0, 10, 20 and 40M) at 0.75 h post fertilization (hpf) and monitored early developmental events such as cleavage, blastula and epiboly that encompass maternal-to-zygotic transition (MZT) and zygotic genome activation (ZGA). Our data revealed that TBBPA exposures induced onset of developmental delays by 3 hpf (blastula). By 5.5 hpf (epiboly), TBBPA-exposed (10-20 M) embryos showed concentration-dependent developmental lag by up to 3 stages or 100% mortality at 40 M. Embryos exposed to sublethal TBBPA concentrations from 0.75-6 hpf and raised in clean water to 120 hpf showed altered larval photomotor response (LPR), suggesting a compromised developmental health. To examine the genetic basis of TBBPA-induced delays, we conducted mRNA-sequencing on embryos exposed to 0 or 40 M TBBPA from 0.75 hpf to 2, 3.5 or 4.5 hpf. Read count data showed that while TBBPA exposures had no overall impacts on maternal or maternal-zygotic genes, collective read counts for zygotically activated genes were lower in TBBPA treatment at 4.5 hpf compared to time-matched controls, suggesting that TBBPA delays ZGA. Gene ontology assessments for both time- and stage-matched differentially expressed genes revealed TBBPA-induced inhibition of chromatin assembly- a process regulated by histone modifications. Since acetylation is the primary histone modification system operant during early ZGA, we immunostained embryos with an H3K27Ac antibody and demonstrated reduced acetylation in TBBPA-exposed embryos. Leveraging in silico molecular docking studies and in vitro assays, we also showed that TBBPA potentially binds to P300- a protein that catalyzes acetylation- and inhibits P300 activity. Finally, we co-exposed embryos to 20 M TBBPA and 50 M n-(4-chloro-3-trifluoromethyl-phenyl)-2-ethoxy-6-pentadecyl-benzamide (CTPB) -a histone acetyltransferase activator that promotes histone acetylation- and showed that TBBPA-CTPB co or pre-exposures significantly reversed TBBPA-only developmental delays, suggesting that TBBPA-induced phenotypes are indeed driven by repression of histone acetylation. Collectively, our work demonstrates that TBBPA disrupts ZGA and early developmental morphology, potentially by inhibiting histone acetylation. Future studies will focus on mechanisms of TBBPA-induced chromatin modifications.

pharmacology and toxicology↗

bioRxiv2024

Addressing anemia severity in antimony-resistant Leishmania donovani infection at the nexus of oxidative outburst and iron transit

Despite the withdrawal of pentavalent-antimonials in treating Visceral leishmaniasis for more than a decade, recent clinical isolates of Leishmania donovani (LD) exhibit unresponsiveness towards pentavalent-antimony (LD-R). This antimony-unresponsiveness points towards a genetic adaptation that underpins LD-Rs evolutionary persistence and superiority over sensitive counterparts. This study highlights LDs response to antimony exposure in terms of increased potential of scavenging host-derived iron within its parasitophorous vacuoles (PV). LD-R employs a strategy to both produce and rapidly scavenge host-iron in a ROS-dependant manner, and selectively reshuffle iron exporter, Ferroportin, around its PV. Higher iron utilization leads to subsequent iron-insufficiency, compensated by increased erythrophagocytosis facilitated by the breakdown of SIRP, orchestrated by a complex interplay of two proteases, Furin and ADAM10. Understanding these mechanisms is crucial for managing LD-R infections and their associated complications, like anemia, and may also provide valuable insights into understanding resistance developed in other pathogens that rely on host iron.

microbiology↗

bioRxiv2022

A neural theory for counting memories

"Ive never smelled anything like this." "Ive seen you once before." "Ive heard this song many times." Keeping track of the number of times different stimuli have been experienced is a critical computation for behavior. This computation occurs ubiquitously across sensory modalities, and naturally without reward or punishment. However, the neural circuitry that mediates this computation remains unknown. Here, we propose a theoretical two-layer neural circuit that can store counts of stimulus occurrence frequencies. This circuit implements a data structure, called a count sketch, that is commonly used in computer science to maintain item frequencies in streaming data. Our first model implements the count sketch data structure using Hebbian synapses and outputs stimulus-specific frequencies. Our second model uses anti-Hebbian plasticity and only tracks frequencies within four count categories ("1-2-3-many"), which we suggest makes a better trade-off between the number of categories that need to be distinguished and the potential ethological value of those categories. Using real-world datasets, we show how both models can closely track the frequencies of different stimuli experienced, while being robust to noise, thus expanding the traditional novelty-familiarity memory axis from binary to continuous. Finally, we show that an implementation of the "1-2-3-many" count sketch -- including network architecture, synaptic plasticity rule, and output neuron that encodes count categories -- exists in a novelty detection circuit in the insect mushroom body, and we argue that similar circuit motifs also appear in mammals, suggesting that basic memory counting machinery may be broadly conserved.

neuroscience↗

bioRxiv2022

Role of vegetation characteristics on the distribution of three hornbill species in and around Pakke Tiger Reserve, Arunachal Pradesh, India

The relationship between various vegetation characteristics and the relative abundance of three hornbill species [Great Pied Hornbill (Buceros bicornis), Wreathed Hornbill (Rhyticeros undulatus) and Oriental Pied Hornbill (Anthracoceros albirostris)] was studied in and around Pakke Tiger Reserve, Arunachal Pradesh. We walked transects (n=11; 22 walks) in three study sites to detect hornbills. Vegetation sampling was done using circular plots (n=33; 10 m radius) at every 400m interval along each transect. Encounter rate (1.5{+/-} 0.188/km) of Great Pied Hornbill (Buceros bicornis) was highest in the protected and undisturbed forest area where food and roosting tree density were also high (114/ha). Oriental Pied Hornbill was common in both the sites within Pakke Tiger reserve near riverine forests (0.75{+/-}0.25/km) and also in the dense undisturbed forest (0.875{+/-}0.226/km). Multivariate analysis revealed that tree density, presence of fruiting trees (utilized by hornbills), canopy cover, and tree diversity in a particular area are the major factors responsible for the assemblage of more than one species of hornbills. The study shows that protection of the forest patches to keep the diversity and density of the tree species intact is crucial for the survival and distribution of the hornbills in the landscape.

ecology↗

bioRxiv2021

Global analysis of human-to-mouse intercellular RNA transfer in cell culture

Full-length mRNAs can transfer between adjacent mammalian cells via direct cell-to-cell connections called tunneling nanotubes (TNTs). However, the extent of mRNA transfer at the transcriptome-wide level (the transferome) is unknown. Here, we analyzed whole transcriptome mRNA and lncRNA transfer between heterogeneous human-mouse cell populations in in vitro co-culture using RNA-sequencing. Our data indicate that mRNA transfer is non-selective, prevalent across the human transcriptome, and that the amount of transfer to mouse embryonic fibroblasts (MEFs) strongly correlates with the endogenous level of gene expression in donor human breast cancer cells (MCF7). These results were validated by both quantitative RT-PCR and in situ hybridization, and analysis shows that typically <1% of endogenous mRNAs and lncRNAs undergo transfer. Non-selective expression-dependent RNA transfer was further validated using synthetic RNA reporters. Notably, significant differential changes in the native MEF transcriptome were observed in response to co-culture, including the upregulation of multiple cancer and cancer-associated fibroblast-related genes and pathways. Together, these results lead us to suggest that TNT-mediated RNA transfer could be a phenomenon of physiological importance under both normal and pathogenic conditions. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=183 SRC="FIGDIR/small/470233v2_ufig1.gif" ALT="Figure 1"> View larger version (44K): org.highwire.dtl.DTLVardef@13a7176org.highwire.dtl.DTLVardef@1ef1c75org.highwire.dtl.DTLVardef@1355eeforg.highwire.dtl.DTLVardef@ebc275_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2021

Non-enzymatic assembly of active chimeric ribozymes from aminoacylated RNA oligonucleotides

Aminoacylated tRNAs, which harbor a covalent linkage between amino acids and RNA, are a universally conserved feature of life. Because they are essential substrates for ribosomal translation, aminoacylated oligonucleotides must have been present in the RNA World prior to the evolution of the ribosome. One possibility we are exploring is that the aminoacyl ester linkage served another function before being recruited for ribosomal protein synthesis. The nonenzymatic assembly of ribozymes from short RNA oligomers under realistic conditions remains a key challenge in demonstrating a plausible pathway from prebiotic chemistry to the RNA World. Here, we show that aminoacylated RNAs can undergo template-directed assembly into chimeric amino acid-RNA polymers that are active ribozymes. We demonstrate that such chimeric polymers can retain the enzymatic function of their all-RNA counterparts by generating chimeric hammerhead, RNA ligase, and aminoacyl transferase ribozymes. Amino acids with diverse side chains form linkages that are well tolerated within the RNA backbone, potentially bringing novel functionalities to ribozyme catalysis. Our work suggests that aminoacylation chemistry may have played a role in primordial ribozyme assembly. Increasing the efficiency of this process provides an evolutionary rationale for the emergence of sequence and amino acid specific aminoacyl-RNA synthetase ribozymes, which could then have generated the substrates for ribosomal protein synthesis. Significance StatementThe emergence of the primordial ribosome from the RNA World would have required access to aminoacylated RNA substrates. The spontaneous generation of such substrates without enzymes is inefficient, and it remains unclear how they could be selected for in a prebiotic milieu. In our study we identify a role for aminoacylated RNA in ribozyme assembly, a longstanding problem in the origin of life research. We show that aminoacylated RNAs, but not unmodified RNAs, rapidly assemble into chimeric amino acid-bridged ribozymes that retain their native enzymatic activity. Our work therefore addresses two key challenges within the origin-of-life field: we demonstrate assembly of functional ribozymes and we identify a potential evolutionary benefit for RNA aminoacylation that is independent of coded peptide translation.

biochemistry↗

bioRxiv2021

Assessment of dependency and consumption pattern of different forest products by the forest fringe villages of Shivalik Himalaya, Uttarakhand, India

Forest are essential for human beings for the enormous services it gives for livelihood and subsistence in the developing countries. We estimated the consumption and extraction levels of three major forest products viz., timber, fuelwood, and fodder in 20 forest fringe villages of the Timli forest range of Uttarakhand, India. We used a questionnaire-based household-level survey to collect information on the household economy and dependence of 380 households selected through stratified random sampling. We estimated that 69% of the overall yearly timber consumption of 20 villages, which comes around 2750 cubic meters (cum), was extracted from the nearby forest. The average timber consumption was 0.52 {+/-} 0.22 cum household-1 year-1. We estimated the total annual fuelwood and fodder consumption to be 298913.89, and 204475 Quintal (Qt). The average fuelwood and fodder consumption were estimated to be 417.6 Quintal household-1 year-1 and 49 {+/-} 9.1 Qt. household-1 year-1. We did general linear regression analysis to assess major biophysical and socio-economic determinants of villages and households for dependency on timber, fuelwood, and fodder. We found that the population of the village, distance from forest, distance from market, and annual average income are the major determining factors for timber, fodder, and fuelwood demand of the villages. Extraction of timber and non-timber forest products was the primary cause of depletion of forest biomass and forest carbon emission. Conservation effective management strategies in collaboration with all the stakeholder departments are needed to conserve forest resources with minimum extraction pressure from forest fringe villages of the study area. HighlightsO_LIQuestionnaire based household level survey was done to assess extraction and consumption patter of fuelwood, fodder and timber in Shivalik range of Uttarakhand, India. C_LIO_LIFuelwood and Timber consumption was found high compared to other studies in the low altitude areas. C_LIO_LI69% of total timber and 62% of the total fodder requirement met from the adjacent forest areas. C_LIO_LIPopulation of the village, distance from the forest and nearby market place, and annual average income were the major determinants of dependency of different forest products. C_LI

ecology↗

bioRxiv2021

Role of Anthropogenic drivers in altering the forest community structure in a prime tiger habitat in central India

Tropical forests are complex systems with heterogenous community assemblages often threatened under conservation conflicts. Herbivory and disturbances affect the diversity and species assemblages within forest patches having different disturbance regimes. We studied the change in plant community composition and structure under a disturbance gradient in the tropical dry deciduous forest of the corridor area between Nagzira-Navegaon Tiger reserve of central India. We tested the hypothesis that the plant community will change along the proximity gradient from the human settlement depending on the anthropogenic stress. We sampled 183 nested quadrat plots to collect data on species abundance and various disturbance parameters. Density, diversity, and Importance Value Index were calculated from the collected data on species abundance and girth at breast height (GBH) of individual tree species. We did multivariate analysis to assess the changes in species assemblage along the disturbance gradients. We found 76% dissimilarity between the plant communities in the three disturbance gradients from near to far from the villages perpetrated by the difference in mean abundance of species like Tectona grandis, Terminalia sp, and Largerstroemia parviflora. The anthropogenic factors significantly influence the density and diversity of tree species and regeneration classes. We found the abundance of regeneration class increased along the distance from the villages. The study intensifies the need for proper management and conservative approach to preserve the minimum diversity of the forest patches for its structural and functional contiguity as a corridor in the central Indias highly susceptible and intricate corridor framework.

ecology↗

bioRxiv2021

Stratification Of Chemotherapy-Treated Stage III Colorectal Cancer Patients Using Multiplexed Imaging And Single Cell Analysis Of T Cell Populations

Colorectal cancer (CRC) has one of the highest cancer incidences and mortality rates. In stage III, postoperative chemotherapy benefits <20% of patients, while more than 50% will develop distant metastases. Predictive biomarkers for identification of patients with increased risk for disease recurrence are currently lacking, with progress in biomarker discovery hindered by the diseases inherent heterogeneity. The immune profile of colorectal tumors has previously been found to have prognostic value. The aims of this study were to evaluate immune signatures in the tumor microenvironment (TME) using an in situ multiplexed immunofluorescence imaging and single cell analysis technology (Cell DIVE). Tissue microarrays (TMAs) with up to three 1mm diameter cores per patient were prepared from 117 stage III CRC patients treated with adjuvant fluoropyrimidine/oxaliplatin chemotherapy. Single sections underwent multilplexed immunofluorescence with Cy3- and Cy5-conjugated antibodies for immune cell markers (CD45, CD3, CD4, CD8, FOXP3, PD1) and cell segmentation markers (DAPI, pan-cytokeratin, AE1, NaKATPase and S6). We applied a probabilistic multi-class, multi-label classification algorithm based on multi-parametric models to build statistical models of protein expression to classify immune cells. Expert annotations of immune cell markers were made on a range of images, and Support Vector Machines (SVM) were used to derive a statistical model for cell classification. Images were also manually scored independently by a Pathologist as high, moderate or low, for stromal and total immune cell content. Excellent agreement was found between manual and total automated scores (p<0.0001). Higher levels of multi-marker classified regulatory T cells (CD3+CD4+FOXP3+PD1-) were significantly associated with disease-free survival (DFS) and overall-survival (OS) (p=0.049 and 0.032), compared to FOXP3 alone. Our results also showed that PD1- Tregs rather than PD1+ Tregs were associated with improved survival. Overall, compared to single markers, multi-marker classification provided more accurate quantitation of immune cells with greater potential for predicting patient outcomes.

cancer biology↗

bioRxiv2018

Physics of lumen growth

We model the dynamics of formation of intercellular secretory lumens. Using conservation laws, we quantitatively study the balance between paracellular leaks and the build-up of osmotic pressure in the lumen. Our model predicts a critical pumping threshold to expand stable lumens. Consistently with experimental observations in bile canaliculi, the model also describes a transition between a monotonous and oscillatory regime during luminogenesis as a function of ion and water transport parameters. We finally discuss the possible importance of regulation of paracellular leaks in intercellular tubulogenesis.

biophysics↗

bioRxiv2018

Extensive genomic diversity among Mycobacterium marinum strains revealed by whole genome sequencing

Mycobacterium marinum is the causative agent for the tuberculosis-like disease mycobacteriosis in fish and skin lesions in humans. Ubiquitous in its geographical distribution, M. marinum is known to occupy diverse fish as hosts. However, information about its genomic diversity is limited. Here, we provide the genome sequences for 15 M. marinum strains isolated from infected humans and fish. Comparative genomic analysis of these and four available genomes of the M. marinum strains M, E11, MB2 and Europe reveal high genomic diversity among the strains, leading to the conclusion that M. marinum should be divided into two different clusters, the \"M\"- and the \"Aronson\"-type. We suggest that these two clusters should be considered, if not two separate species, at least two M. marinum subspecies. Our data also show that the M. marinum pan-genome for both groups is open and expanding and we provide data showing high number of mutational hotspots in M. marinum relative to other mycobacteria such as Mycobacterium tuberculosis. This high genomic diversity might be related to that M. marinum occupy different ecological niches.

genomics↗

bioRxiv2017

A neural algorithm for a fundamental computing problem

Similarity search, such as identifying similar images in a database or similar documents on the Web, is a fundamental computing problem faced by many large-scale information retrieval systems. We discovered that the flys olfac-tory circuit solves this problem using a novel variant of a traditional computer science algorithm (called locality-sensitive hashing). The flys circuit assigns similar neural activity patterns to similar input stimuli (odors), so that behav-iors learned from one odor can be applied when a similar odor is experienced. The flys algorithm, however, uses three new computational ingredients that depart from traditional approaches. We show that these ingredients can be translated to improve the performance of similarity search compared to tra-ditional algorithms when evaluated on several benchmark datasets. Overall, this perspective helps illuminate the logic supporting an important sensory function (olfaction), and it provides a conceptually new algorithm for solving a fundamental computational problem.

neuroscience↗