Biology subjects

Chakrabarti, S.

Publications and source records attributed to Chakrabarti, S..

At least 37 records · Page 2Linked to original sources

bioRxiv2024

TENM4 is an essential transduction component for touch and pain

Gentle touch is conveyed to the brain by fast-conducting sensory fibers. Mechanosensitive ion channels at the terminals of these neurons are thought to be gated by extracellular tethers that transmit force from the surrounding matrix to the channel complex, but the molecular identity of such tethers has remained unknown. Here, we identify Teneurin-4 (TENM4) as an essential extracellular linker protein for mechanotransduction in mechanoreceptors. Sensory neuron-specific deletion of Tenm4 in mice caused profound touch insensitivity, while acute and reversible proteolytic disassembly of TENM4 at sensory endings confirmed its direct role in force transduction. Ultrastructural analyses revealed TENM4 localization to filamentous structures at the neurite-laminin interface, defining it as a structural component of the mechanosensory tether. These findings identify TENM4 as a core element of fast somatic sensation and provide molecular insight into how extracellular forces are coupled to ion channel activation.

neuroscience↗

bioRxiv2024

LncRNA TAAL is a Modulator of Tie1-Mediated Vascular Function in Diabetic Retinopathy

Diabetic retinopathy (DR), a leading cause of vision impairment and blindness, is characterized by abnormal retinal vascular changes due to chronic hyperglycemia. The Tie-1 signaling pathway, essential for vascular growth and remodeling, has emerged as a key therapeutic target, though its molecular mechanisms and interactome remain largely unclear. Through a protein-centric approach, we identified a novel lncRNA and named it Tie1-associated angiogenic lncRNA (TAAL). TAAL lncRNA regulates endothelial cell migration, proliferation, tube formation, and permeability by modulating ER-calcium homeostasis and cytoskeleton dynamics. In zebrafish, taal modulation led to angiogenic defects, which were rescued by human TAAL orthologue. Our molecular studies further revealed that TAAL negatively regulates Tie1 protein via ubiquitin-mediated degradation. Notably, TAAL expression is upregulated in the blood of DR patients and downregulated in endothelial DR cell models. Overexpression of TAAL restored endothelial permeability and VE-cadherin surface expression. These findings establish TAAL as a novel regulator of Tie1 protein turnover, with potential therapeutic implications for diabetic retinopathy.

genetics↗

bioRxiv2024

Species-specific interactions with apex carnivores yield unique benefits and burdens for mesocarnivores

Mesocarnivores navigate a complex risk-reward continuum in ecosystems shared with their apex counterparts, balancing scavenging opportunities with risks of mortality. However, the risks to mesocarnivores in multi-carnivore systems are not uniform--often varying with specific apex-meso pairings. Using camera traps and GPS-telemetry, we examined space-use, temporal activity, fine-scale interactions, and scavenging behavior of apex (wolves, cougars) and mesocarnivores (red foxes, coyotes) in northern Yellowstone National Park, USA. Coyote space use was positively linked to wolf presence, while red fox space use was positively linked to cougar presence. Notably, photo-detection rates of mesocarnivores doubled within 24 hours of apex carnivore detections--except for coyotes following cougars. Coyotes were more frequent scavengers at wolf and cougar kills than red foxes, but they also suffered the highest mortality rates. Over 60% of wolf caused coyote mortalities were linked to wolf kill-sites, though wolves rarely consumed the coyotes themselves. In contrast, cougars hunted and consumed coyotes as prey, posing an additional risk of predation. These findings highlight how divergent hunting strategies and habitat preferences between canid and felid apex carnivores create separate risks for mesocarnivores. Our study offers new insights into the species-specific behavioral dynamics that can shape trophic interactions in multi-carnivore systems.

ecology↗

bioRxiv2024

Biased recruitment of H3.3 by HIRA is dictated by de-/acetylation and determines transcription memory and response

Chromatin plasticity and epigenetic memory, fundamental for eukaryotic biology, are determined by differential/regulated de novo deposition or recycling of pre-existing histones, which in turn dictate transcriptional programs. Recruitment of the histone-H3 variant, H3.3, mediated by the HIRA chaperone complex, is both causally and consequentially associated with transcription. Despite decades of work, endogenous regulatory mechanisms that differentiate between de novo deposition and recycling activities of HIRA are still unknown. Here, we have investigated the pivotal role of HIRA de-/acetylation in regulating its function. Our results unequivocally establish function separation effects of acetyl and deacetyl mimic mutations of lysine-600, vis-a-vis de novo deposition or recycling of H3.3, respectively. Importantly, we demonstrate that HIRA deacetylation-dependent biased H3.3 recycling determines transcriptional output, possibly through preferential enrichment of H3.3-K36me3. Besides unraveling tunable regulatory mechanism that governs HIRA function, we illustrate a causal link between the chaperone activity, biased recruitment of pre-existing histones, and gene expression.

molecular biology↗

bioRxiv2024

Androgen receptor plays critical role in regulating cervical cancer cell migration

Cervical cancer (CC) is the second most common cancer among women in India and the fourth worldwide. While major genes and pathways have been studied, further research is needed to identify candidates for targeted therapy in metastatic disease. This study used a network biology approach to identify key genes in disease progression. Stage-specific cervical cancer protein-protein interaction networks (PPIN) were constructed by overlaying stage-specific, patient-derived transcriptomics data onto a human protein-protein interaction network (HPPIN). Graph-theory-based network analysis identified important interacting proteins (IIPs) with maximum connectivity, high centrality scores, and significant global and local network perturbation scores. Among the identified IIPs, the Androgen receptor (AR) emerged as one of the crucial yet understudied regulator in cervical cancer. Patient samples and in vitro experiments showed significant downregulation in cervical cancer. Ligand-dependent overexpression of AR reduced cancer cell migration while failed to induce apoptosis in CC cell lines. Downregulation of mesenchymal markers and restoration of epithelial markers suggested ARs potential in reversing invasive properties of cervical cancer cells. AR overexpression upregulated its downstream target PTEN and restored GSK3{beta} activity by interfering with AKT phosphorylation, probably leading to degradation of mesenchymal markers. Further studies showed AR reduced cell motility by hindering focal adhesion formation and Actin filament assembly. An increased G-Actin ratio suggested AR disrupted cytoskeletal dynamics through the RhoA/ROCK1/LIMK1/CFL1 pathway, impeding cervical cancer cell spread.

cancer biology↗

bioRxiv2024

Functional implications of the interaction of the SARS-CoV-2 Nucleocapsid protein with factors involved in nonsense-mediated mRNA decay

The RNA genome of the SARS-CoV-2 virus encodes for four structural proteins, 16 non- structural proteins and nine putative accessory factors. A high throughput analysis of interactions between human and SARS-CoV-2 proteins identified multiple interactions of the structural Nucleocapsid (N) protein with RNA processing factors. The N-protein, which is responsible for packaging of the viral genomic RNA was found to interact with two RNA helicases, UPF1 and MOV10 that are involved in nonsense-mediated mRNA decay (NMD). Using a combination of biochemical and biophysical methods, we investigated the interaction of the SARS-CoV-2 N-protein with NMD factors at a molecular level. Our studies led us to identify the core NMD factor, UPF2, as an interactor of N. The viral N-protein engages UPF2 in multipartite interactions and can negate the stimulatory effect of UPF2 on UPF1 catalytic activity. N also inhibits UPF1 ATPase and unwinding activities by competing in binding to the RNA substrate. We further investigate the functional implications of inhibition of UPF1 catalytic activity by N in mammalian cells. The interplay of SARS-CoV-2 N with human UPF1 and UPF2 does not affect decay of host cell NMD targets but might play a role in stabilizing the viral RNA genome.

biochemistry↗

bioRxiv2024

Mutational and evolutionary dynamics of non-structural and spike proteins from variants of concern (VOC) ofSARS-CoV-2 in India

Monitoring the genetic diversity and emerging mutations of SARS-CoV-2 still remains to be crucial in India. This study extensively analyzes the lineage dynamics, mutation screening, structural analysis, and phylodynamics of SARS-CoV-2 variants of concern (VOC) in India from October 2020 to September 2023. Predominant variants identified include alpha, beta, delta, and omicron, with delta and omicron making up 76.05% of sequenced genomes. The B.1.617.2 lineage of delta was the major contributor to COVID-19 cases before the rapid rise of omicron. Mutation screening of non-structural proteins (NSPs) and spike revealed distinct profiles for each VOC. Co-mutation patterns/networks of the most frequently observed mutations specific for each VOC were also identified and subsequently structural and energetic alteration of the co-mutants were analyzed using rigorous molecular dynamics simulations. Furthermore, comparative analysis of phylogenetic trees based on genomic and mutational data revealed that nsp1, nsp3, nsp4, nsp13, and nsp14 exhibit strongest association with the increased mutation load across the genome of SARS-CoV-2 in Indian population. Our comparative phylogenetic study also revealed that mutability patterns of nsp14 and spike have highest similarity supporting critical role of nsp14 for SARS-CoV-2 infectivity and persistence. This research provides a comprehensive overview of SARS-CoV-2 evolution in India.

bioinformatics↗

bioRxiv2024

The impact of ERUPR on mitochondrial integrity mediated by PDK4

ER and mitochondrial stress are often interconnected and considered as major contributors to aging as well as neurodegeneration. Co-ordinated induction of ERUPR and mitoUPR has been observed in diabetes and pulmonary disorders. However, in the context of aging and neurodegeneration, regulation of this intra-organellar crosstalk has remained relatively elusive. Here, we demonstrate that pyruvate dehydrogenase kinase 4 (PDK4), a mitochondrial protein accumulates at the ER-mitochondrial contact sites (MAMs) during ER stress. Classically, PDK4 is known to phosphorylate PDHA1 (pyruvate dehydrogenase E1 subunit alpha 1), and plays a significant role in regulating the oxidative phosphorylation driven ATP production. In this study, we propose a non-canonical function of PDK4; we show that it acts as a connecting link between ERUPR and mitoUPR with significance in aging and neurodegeneration. Transcriptomics analyses show increased PDK4 levels upon drug induced ER stress. We detect elevated PDK4 levels in brain lysates of Alzheimers disease (AD) patients, as well as in in vivo and ex vivo AD models. Additionally, exogenous expression of PDK4 was found to refine ER-mitochondrial communication, significantly altering mitochondrial morphology and function. Further, we also observe defective autophagic clearance of mitochondria under such conditions. It is prudent to suggest that elevated PDK4 levels could be one of the key factors connecting ERUPR with mitoUPR, a phenotypic contributor in aging and at least some neurodegenerative diseases.

molecular biology↗

bioRxiv2024

Comparative vitreous proteomic profiling of proliferative diabetic retinopathy and diabetic with no-retinopathy subjects implicates impaired autophagy in DR pathogenesis

PurposeDiabetic retinopathy (DR) is a neurovascular complication of diabetes (DM) causing the loss of neurons (ganglion cells) in the retina. The disease etiology and potential pathogenic mechanisms in this disease remains unclear. In the present study, we aimed to further understand the key and novel pathogenic mechanisms involved in DR pathogenesis by taking cues from our global proteomics data. MethodologyThe study was approved by the institutional review board (IRB) of LVPEI, Hyderabad, India. Vitreous humour samples (PDR; n=3, DM; n=3, Control; n=3) were collected from patients undergoing vitrectomy and subjected to LC-MS-MS analysis. The acquired raw data were searched against the human vitreous proteome and was further analysed by various bioinformatic and proteomic tools. Western blotting and IHC was performed to validate crucial pathways. Blood samples from patients (DM, PDR & NPDR) and controls (n=50); cadaveric retinas from diabetic and non-diabetic donors (n=10) and epiretinal membranes (ERM, n=10) from DR cases and controls were collected and RNA was isolated. Quantitative expression of genes involved in autophagy were performed. CT was compared across different categories and significance estimated using a student t-test. ResultA total of 1079 proteins were identified with 16 completely novel proteins in eye proteome. Top pathways identified were autophagy, inflammation, LXR/RXR activation (lipid metabolism), ROS generation by macrophages, apoptosis and protein degradation. Regulatory proteins identified were associated with cell death, phagocytic activation, angiogenesis and apoptosis. Autophagy inducers such as ROS was found to be accumulated in the DR vitreous. TREM2, microglial receptor was identified as a novel protein in PDR vitreous. The expression of TREM2, an autophagy-associated gene was significantly (p-value = 0.05) upregulated in all categories as compared to control (NDM and/or NDM/No-DR). TREM2 protein also seemed to colocalise with microglial marker F4/80 in retinal tissues and intense expression was observed near the blood vessels in case of PDR retina. Other autophagy-associated markers were also differentially regulated in DR as compared to controls. ConclusionThis study emphasises on the strong role of autophagy pathways and its associated genes in the development of DR.

genomics↗

bioRxiv2024

Transcriptional noise sets fundamental limits to spatially resolved cell-state decoding of the circadian clock.

The rhythmic circadian clock generates temporal variation of critical physiological processes in tissues, making accurate measurement of clock phase or tissue time a fundamentally important problem. Recent advances in single-sample time inference provides a potentially powerful alternative to traditional time-series based approaches. Single-sample techniques typically leverage population-level RNA rhythms, but the feasibility of single-cell phase detection remains an open question. Combining multiplexed smFISH to simultaneously measure up to 6 mouse fibroblast genes, with a novel inference algorithm using Gaussian Processes, here we demonstrate that even when technical drop-outs are minimized, transcriptional noise in core-clock genes precludes single-cell phase inference. Simulations predict that above 50 clock-like genes would make single-cell phase inference possible. Remarkably however, just 3 core-clock genes are sufficient if RNA levels of [~] 70 cells are averaged. Further, we demonstrate how averaging allows detecting spatially-resolved, heterogeneous clock phases in desynchronized cells. Our work provides a conceptual framework for achieving high-resolution phase detection with a minimal set of core-clock genes, with implications for probing the origins of clock dysfunction, otherwise unresolvable using population-based approaches.

cell biology↗

bioRxiv2024

CircRNA_012164/MicroRNA-9-5p axis mediates cardiac fibrosis in diabetic cardiomyopathy

Noncoding RNAs play a part in many chronic diseases and interact with each other to regulate gene expression. MicroRNA-9-5p (miR9) has been thought to be a potential inhibitor of diabetic cardiomyopathy. Here we examined the role of miR9 in regulating cardiac fibrosis in the context of diabetic cardiomyopathy. We further expanded our studies through investigation of a regulatory circularRNA, circRNA_012164, on the action of miR9. We showed at both the in vivo and in vitro level that glucose induced downregulation of miR9 and upregulation of circRNA_012164 resulted in the subsequent upregulation of downstream fibrotic genes. Further, knockdown of circRNA_012164 shows protective effects in cardiac endothelial cells and reverses increased transcription of genes associated with fibrosis and fibroblast proliferation through a regulatory axis with miR9. This study presents a novel regulatory axis involving noncoding RNA that is evidently important in the development of cardiac fibrosis in diabetic cardiomyopathy.

pathology↗

bioRxiv2024

Combining lineage correlations and a small molecule inhibitor to detect circadian control of the cell cycle.

Chronotherapy has emerged as an exciting possibility for improving treatment regimens in cancer. A strong influence of the circadian clock on the cell cycle is a crucial requirement for successful chronotherapy. However, though a number of molecular interactions have been discovered between these two oscillators, it remains unclear whether these interactions are sufficient to generate emergent control of cellular proliferation. In this work, we computationally explore a strategy to detect clock control over the cell cycle, by computing lineage correlations in cell cycle times in the presence and absence of the clock inhibitor KL001. Using phenomenological models, we and others have previously suggested that the cousin-mother inequality - a phenomenon where cousin cells show stronger cell cycle time correlations than mother-daughter pairs, could be leveraged to probe circadian effects on cellular proliferation. Using stochastic simulations calibrated to match HCT116 colon cancer proliferation datasets, we demonstrate that the established gene-networks giving rise to the cell cycle and circadian oscillations are sufficient to generate the cousin-mother inequality. In the presence of KL001 which stabilizes CRY1, our models predict greater than 50% decrease in the cousin-mother inequality, but counter-intuitively, very little change in population growth rates. Our results predict a range of underlying cell cycle times where the cousin-mother inequality should be observed, and consequently suggest the exciting possibility of combining measurements of lineage correlations with KL001 as a probe of circadian clock - cell cycle interactions.

biophysics↗

bioRxiv2024

Localization and function of key axonemal microtubule inner proteins and dynein docking complex members reveal extensive diversity among vertebrate motile cilia

Vertebrate motile cilia are broadly classified as (9+2) or (9+0), based on the presence or absence of the central pair apparatus, respectively. Cryogenic electron microscopy (cryo-EM) analyses of (9+2) cilia have uncovered an elaborate axonemal protein composition; whether these features are relevant to (9+0) cilia remain unclear. We previously demonstrated that Cfap53, a key microtubule inner protein (MIP) as well as centriolar-satellites component, is essential for motility of (9+0), but not (9+2) cilia. Here, we show that in (9+2) cilia, Cfap53 functions redundantly with a paralogous MIP, Mns1. Mns1 localizes to ciliary axonemes, and combined loss of both proteins in zebrafish and mice, caused severe loss of outer dynein arms (ODAs) of (9+2) cilia, significantly affecting their motility. Moreover, using immunoprecipitation, we demonstrate that while Mns1 can self-associate and interact with Cfap53, Cfap53 is unable to self-associate. Finally, we show that multiple additional dynein interacting proteins, such as the ODA docking complex (ODA-DC) members, show strikingly distinct localization patterns between various motile cilia-types. Our findings clarify how paralogous MIPs, Cfap53 and Mns1, function in regulating motility of (9+2) versus (9+0) cilia, and establish that localization pattern of other key motility proteins also differ between these cilia-types, further emphasizing extensive structural variations among these organelles.

developmental biology↗

bioRxiv2023

Discerning state estimation and sensory gating, two presumptive predictive signals in mouse barrel cortex

The brain is assumed to contain distinctive predictive systems ranging from sensorimotor to cognitive functions. Here we report the successful functional delineation of two different presumptive predictive systems on the neuronal level, state estimation (SE) and sensory gating (SG), which both attenuate sensory flow during movement. Studying neuronal sensorimotor responses throughout the depth of primary somatosensory cortex in mice trained on a whisker reach task, SE appeared as a learned attenuation of tactile signal flow, due to a suppressive predictive signal, precisely at the time of an experimental sensory consequence. In contrast, SG was observable during a much longer interval after onset of the motor command. Both phenomena are internal, presumptively predictive signals, as blockade of the reafference did not affect them. We speculate that SG may be related to cognitive signals monitoring goals of movements, while SE likely is the expression of the classical notion of the reafference principle.

neuroscience↗

bioRxiv2023

Digging deeper into pain - an ethological behavior assay correlating well-being in mice with human pain experience.

The pressing need for safer, more efficacious analgesics is felt worldwide. Pre-clinical tests in animal models of painful conditions represent one of the earliest checkpoints novel therapeutics must negotiate before consideration for human use. Traditionally, the pain status of laboratory animals has been inferred from evoked nociceptive assays which measure their responses to noxious stimuli. The disconnect between how pain is tested in laboratory animals and how it is experienced by humans may in part explain the shortcomings of current pain medications and highlights a need for refinement. Here, we survey human chronic pain patients who assert that everyday aspects of life, such as cleaning and leaving the house, are affected by their on-going level of pain. Accordingly, we test the impact of painful conditions on an ethological behavior of mice, digging. Stable digging behavior was observed over time in naive mice of both sexes. By contrast, deficits in digging were seen following acute knee inflammation. The analgesia conferred by meloxicam and gabapentin was compared in the monosodium iodoacetate knee osteoarthritis model, meloxicam more effectively ameliorating digging deficits, in line with human patients finding meloxicam more effective. Lastly, in a visceral pain model, the decrease in digging behavior correlated with the extent of disease. Ultimately, we make a case for adopting ethological assays, such as digging, in studies of pain in laboratory animals, which we believe to be more representative of the human experience of pain and thus valuable in assessing clinical potential of novel analgesics in animals.

neuroscience↗

bioRxiv2023

The Lion's Share: Implications of carnivore diet on endangered herbivores in Tsavo

Predation by mammalian carnivores can have cascading, regulatory effects across ecological communities. An understanding of predator diet can therefore provide crucial information regarding their ecology and conservation, as well as their impacts on prey populations. Using scats collected between 2019 and 2023, coupled with estimates of prey abundance from aerial surveys, we characterized lion prey-consumption and preference in Tsavo, Kenya. A lion-specific biomass model applied to prey frequencies in scats revealed that more than 85% of lion diet consisted of large ungulates weighing over 150 kg. While large ungulates were also preferred prey items in terms of their availability, we found a disproportionately high consumption and preference of lions for the endangered hirola and Grevys zebra-- species that were introduced in Tsavo as part of ex-situ conservation programs. Hirola and Grevys zebra populations have historically remained small in Tsavo despite strong recovery efforts, and our results likely indicate a disproportionate impact of lion predation on these small but crucial populations. Preferential predation, coupled with high availability of alternative prey, may trap hirola and Grevys zebra within a predator-pit. Our findings have strong implications for understanding lion diet, optimal foraging, and the potential effects predators have on endangered prey species in a landscape of critical conservation importance.

ecology↗

bioRxiv2023

Touch sensation requires the mechanically-gated ion channel Elkin1.

The slightest touch to the skin initiates tactile perception that is almost immediate1. The extraordinary speed of touch perception is enabled by mechanically-activated ion channels, the opening of which excites the endings of sensory neurons innervating the skin to initiate sensation. Here we identify a new mechanically-activated ion channel, Elkin12, that, when ablated in mice, leads to a profound behavioural touch insensitivity. Touch insensitivity in Elkin1-/- mice was caused by a loss of mechanically-activated currents (MA-currents) in around half of all sensory neurons that are activated by light touch (low threshold mechanoreceptors, LTMRs). Reintroduction of Elkin1 into sensory neurons from Elkin1-/- mice acutely restored MA-currents. Piezo23-6 is an established mechanosensitive ion channel required for touch sensation. In mice genetic ablation of Piezo2 renders many, but not all, LTMRs insensitive to mechanical force4,5,7. Here we show that Elkin1 underpins PIEZO2-independent touch sensation. Additionally, we find that Elkin1 is present in many nociceptive sensory neurons which detect potentially damaging and painful mechanical force. These nociceptors depend on Elkin1 for effectively communicating information on sustained noxious mechanical forces. We further identified molecular and functional interactions between the known mechanotransduction protein Stoml38,9 and Elkin1 ion channels. Our data identify Elkin1 as a novel core component of touch transduction in mammals. The specific sensory deficits exhibited by Elkin1-/- mice make Elkin1 a highly desirable target that could be harnessed to treat somatic sensory disorders including pain.

neuroscience↗

bioRxiv2023

Eating an elephant, one bite at a time: predator interactions at carrion bonanzas

Resource specific competition between predators has typically been studied from their interactions at meso-herbivore carcasses, because such carcasses are abundant. Mega-carcasses like those of elephants are rare but unparalleled in the extent of carrion biomass they offer and the long durations they can persist. These rare resource bonanzas can thus provide unique opportunities to understand sympatric species interactions within likely relaxed competitive scenarios. Using remote cameras that were operational 24-h a day, we monitored two elephant carcasses in Tsavo, Kenya, from when they were discovered until they were completely consumed or became inaccessible. While we found high temporal overlaps in activity patterns between all predators, the terrestrial predator guild (lion/leopard/spotted hyena) was not observed to feed simultaneously, suggesting strong interference competition. Based on photo-analysis and video-evidence of exclusion from a carcass, interference competition within the terrestrial predator guild favored lions over hyenas, and hyenas over leopards. The carcass at the terrestrial-aquatic interface showed more simultaneous feeding bouts between predators (crocodile/spotted hyena), indicating either facilitation and/or higher coexistence between predators that typically occupy different niches. We also observed a hippopotamus scavenging from an elephant carcass, thereby documenting a rare instance of a megaherbivore feeding on a megaherbivore. Our results highlight the importance of monitoring such carcasses through remote cameras, which can significantly add to our existing understanding of food webs and carrion ecology.

ecology↗