Biology subjects

Sengupta, M.

Publications and source records attributed to Sengupta, M..

15 recordsLinked to original sources

bioRxiv2026

Transforming off-the-shelf personal glucose meter into a sustainable and decentralized label-free nucleic acid and NAAT detection platform

In todays world, point-of-care nucleic acid detection still remains extensively constrained and limited by the heavy dependence on centralized urban instrumentation facilities and complex assay workflows. Here, we elucidate a glucometer-based analytical platform that enables label-free detection of nucleic acids and the nucleic acid amplification products through a simple redox-mediated mechanism. The approach leverages the potassium ferricyanide (K3[Fe(CN)6])/ potassium ferrocyanide (K4[Fe(CN)6]), redox system, which is intrinsic to commercial glucometers, complementing with interactions between methylene blue (MB) and nucleic acids. These interactions transduce concentration differences in nucleic acids into quantifiable electrochemical signal readouts. Distinct varied signal outputs are observed between single-stranded and double-stranded DNA, enabling the direct detection as well as integration with nucleic acid amplification tests (NAATs), including polymerase chain reaction, rolling circle amplification, and loop-mediated isothermal amplification. Optimization of reaction parameters and conditions leads to enhancement of the overall signal discrimination and sensitivity across various assay formats. This innovation repurposes widely available off-the-shelf glucometers as a low-cost, portable nucleic acid detectors, thus eliminating the need for any specialized instrumentation. Our results enumerate and establish a generalized and scalable strategy for nucleic acid sensing. The platform thus supports sustainable and environmentally responsible point-of-care testing, thereby enabling improved accessibility and public health monitoring at resource-limited and remote settings.

biochemistry↗

bioRxiv2026

A PROPPIN links V-ATPase assembly to endocytic membrane dynamics in malaria parasites

Malaria parasites replicate inside red blood cells, degrading hemoglobin within a specialized digestive vacuole. Efficient hemoglobin processing is essential for parasite survival and influences antimalarial drug susceptibility. The vacuole constantly fuses with incoming hemoglobin-filled vesicles, yet the mechanisms that balance cargo influx with membrane homeostasis remain unclear. Here, using conditional reverse genetics, quantitative live-cell imaging, and 3D electron microscopy, we characterize the autophagy-related protein 18 of Plasmodium falciparum (PfATG18) as a key regulator of vacuolar membrane dynamics. Loss of PfATG18 caused vacuole fragmentation, accumulation of hemoglobin-filled vesicles, and parasite death. These defects were preceded by broad architectural destabilization of the parasites V-ATPase, a proton pump controlling organelle acidification and the vacuoles fusion-fission equilibrium. Direct interference with its membrane sector phenocopied PfATG18 deficiency. We found that PfATG18 does not interact directly with the proton pump but instead associates with a putative V-ATPase assembly factor and with complexes regulating phosphoinositide balance and vesicle trafficking. The breadth of these interactions indicates a multifaceted role at the vacuolar membrane and a regulatory influence on V-ATPase mediated through associated protein machinery. Although a point mutation in PfATG18 has been linked to artemisinin resistance, its complete knockout did not decrease sensitivity, but rather hypersensitized ring-stage parasites to dihydroartemisinin. Together, these findings establish PfATG18 as a central regulator of endocytic membrane homeostasis, essential for V-ATPase function and asexual parasite proliferation in the human blood.

microbiology↗

bioRxiv2025

Evaluating horseradish peroxidase-mimic DNAzyme transducer for glucometer readout: roles of various components and their optimization

The Development of decentralized disease detection elements is integral towards ensuring democratized access to healthcare. Personal glucometers, thanks to their low cost, ease of usage, and universal availability, are increasingly being identified as a central tenet of democratized disease detection. Towards this objective, it will first be critical to establish whether glucometers can be integrated with established molecular diagnosis workflows such as enzyme-linked absorbent assay (ELISA) or its analogous techniques such as enzyme-linked aptasorbent assays (ELASA). In ELISA or its analogous assays, horseradish peroxidase (HRP) or its mimics, such as hemin DNAzyme conjugated to biorecognition elements (e.g., antibody or aptamers), serve as the signal transducers. These transducers generate colorimetric or electrochemical signals measurable in centralized instruments such as ELISA readers or potentiostats. To become compatible with ELISA readouts in decentralized settings, the glucometers should therefore be able to quantify the activity of HRP mimics such as hemin DNAzyme. The current work thus explores the possibility of the use of glucometers as a general readout instrument for hemin DNAzyme transducer activity involving potassium ferrocyanide as a novel redox substrate. Using both absorbance and glucometer readout, we systematically investigated the role of buffer, pH, redox mediator, and H2O2 in achieving optimal readout conditions. It also probed the prospective DNAzyme operational range under these assay conditions. In addition, transferability of optimized assay parameters in other commercial glucometer brands, as well as preliminary suitability in HRP enzyme, was explored. Altogether, our study explores the possibility of using DNAzymes as transducers for glucometers as a democratized alternative to ELISA readers, multimode readers, or potentiostats towards equitable access to disease diagnosis.

biochemistry↗

bioRxiv2024

Role of SCAL1 in Modulating Oxidative Stress, Cancer Stemness, Apoptotic Resistance in Tumorigenic Differentiation of Cigarette Smoke-Exposed BEAS-2B Cells

The smoke and cancer-associated lncRNA 1 (SCAL1) is an emergent biomarker in lung cancer. However, the precise role of SCAL1 as a mediator of tobacco smoke-induced lung carcinogenesis remains unclear. BEAS-2B cells were cultured and exposed to 20% cigarette smoke extract (CSE), followed by quantification of SCAL1. We evaluated the impact of SCAL1 on cell viability, ROS mitigation, cancer stemness, tumorigenic differentiation, cellular invasiveness, and apoptosis for different CSE incubation time points through SCAL1 expressional modulation using SCAL1-specific siRNAs and scrambled controls. We observed an upregulation of SCAL1 in cells exposed to CSE for 2, 4, and 6 hours, with the highest expression observed at 6 hours (p<0.001). Exposure of BEAS-2B cells to CSE showed the formation of focal adhesions and stress fibers resembling tunneling nanotubes. Intracellular ROS levels significantly increased upon CSE exposure compared to control cells (p<0.05). We found increased levels of anti-apoptotic and cancer stem (CSC) cell markers like BCL2, ALDH1A1, CD133, CD44, and TCTP and decreased levels of TP53 in CSE-exposed cells. Knockdown of SCAL1 using siRNA transfection reversed these effects at all time points. Additionally, we observed a significant decrease in the number of spheroid colonies in siSCAL1 (+) cells compared to siSCAL1 (-) cells (p<0.01) exposed to CSE. SCAL1 is pivotal in mediating cellular responses to cigarette smoke, leading to tumorigenic differentiation of BEAS-2B cells. Understanding the mechanisms could provide valuable insights into lung cancer pathogenesis and therapeutic approaches.

cancer biology↗

bioRxiv2024

An anomalous 3'-terminal phosphorothioated mismatch bypass activity and its application as a binary molecular switch

Phosphorothioated (PST) oligonucleotides are increasingly being used in RNA silencing, antisense, and biosensing applications. However, the possibilities and consequences of their desultory interactions with other possible nucleic acids and DNA polymerases inside the cell remain inadequately characterized. In this study, we report the discovery of an unusual terminal mismatch bypass activity involving 3'-PST containing DNA primers and certain strand displacement DNA polymerases. Using rolling circle DNA amplification, we have identified that strand displacement DNA polymerases such as phi29 and BST large fragment (LF) can bypass 3'-terminal PST mismatches upto 1 - 20 nt length. Next, we explore the length and sequence dependence of this unusual attribute, incubation in near-ambient and 60 - 65{degrees}C temperatures, and measures to blockade or modulate this mismatch bypass activity to create a binary fully nucleic acid-based and non-photocontrolled molecular switch (the first of its kind). After proposing possible underlying mechanisms for this activity, we discuss its potential consequences and applications.

biochemistry↗

bioRxiv2024

Highly synchronized inhibition from Purkinje cells entrains cerebellar output in zebrafish

Cerebellar function, known to be important for motor learning and motor coordination, is mediated by efferent neurons that project to diverse motor areas. To understand cerebellar function, it is imperative to study how these efferent neurons integrate inputs from the principal neurons of the cerebellar cortex, the inhibitory Purkinje neurons (PNs). In zebrafish, PNs are bistable and we show here that bistability influences spike synchrony among PNs. Bistability also alters spike correlation with motor bouts. We asked how PN population synchrony influences Eurydendroid cells (ECs), which are postsynaptic targets of PNs and are the cerebellar efferent cells in zebrafish. Using optogenetics, we artificially modulated population synchrony of PNs over millisecond time scales and showed that under conditions of high synchrony, EC firing is briefly suppressed and entrained by PN spiking. However, the magnitude of such modulation is relatively small and indicates a strong combined influence of other synaptic inputs on EC spiking. Key PointsO_LICerebellar Purkinje neurons (PN) in larval zebrafish alter simple spike correlations with each other based on cellular state. C_LIO_LIThey also alter simple spike correlations with motor bouts as a function of state. C_LIO_LIWe altered PN population synchrony in a graded manner using optogenetics. C_LIO_LIPN targets are cerebellar efferent neurons, which in teleosts are called eurydendroid cells. C_LIO_LIWhen PN population is firing with high synchrony, eurydendroid cells are entrained better than when the PN input is asynchronous. C_LIO_LIThis can explain how PNs use bistability to modulate their influence on cerebellar output and ultimately, motor behavior. C_LI

neuroscience↗

bioRxiv2023

A phenotypically robust model of Spinal and Bulbar Muscular Atrophy in Drosophila

Spinal and bulbar muscular atrophy (SBMA) is an X-linked disorder that affects males who inherit the androgen receptor (AR) gene with an abnormal CAG triplet repeat expansion. The resulting protein contains an elongated polyglutamine (polyQ) tract and causes motor neuron degeneration in an androgen-dependent manner. The precise molecular sequelae of SBMA are unclear. To assist with its investigation and the identification of therapeutic options, we report here a new model of SBMA in Drosophila melanogaster. We generated transgenic flies that express the full-length, human AR with a wild-type or pathogenic polyQ repeat. Each transgene is inserted into the same "safe harbor" site on the third chromosome of the fly as a single copy and in the same orientation. Expression of pathogenic AR, but not of its wild-type variant, in neurons or muscles leads to consistent, progressive defects in longevity and motility that are concomitant with polyQ-expanded AR protein aggregation and reduced complexity in neuromuscular junctions. Additional assays show adult fly eye abnormalities associated with the pathogenic AR species. The detrimental effects of pathogenic AR are accentuated by feeding flies the androgen, dihydrotestosterone. This new, robust SBMA model can be a valuable tool towards future investigations of this incurable disease.

neuroscience↗

bioRxiv2022

A machine-learning tool to identify bistable states from calcium imaging data

Mapping neuronal activation using calcium imaging in vivo during behavioral tasks has advanced our understanding of nervous system function. In almost all of these studies, calcium imaging is used to infer spike probabilities since action potentials activate voltage-gated calcium channels and increase intracellular calcium levels. However, neurons not only fire action potentials, but also convey information via intrinsic dynamics such as by generating bistable membrane potential states. While a number of tools for spike inference have been developed and are currently being used, no tool exists for converting calcium imaging signals to maps of cellular state in bistable neurons. Purkinje neurons (PNs) in the larval zebrafish cerebellum exhibit membrane potential bistability, firing either tonically or in bursts. Several studies have implicated the role of a population code in cerebellar function, with bistability adding an extra layer of complexity to this code. In this manuscript we develop a tool, CaMLSort which uses convolutional recurrent neural networks to classify calcium imaging traces as arising from either tonic or bursting cells. We validate this classifier using a number of different methods and find that it performs well on simulated event rasters as well as real biological data that it had not previously seen. Moreover, we find that CaMLsort generalizes to other bistable neurons, such as dopaminergic neurons in the ventral tegmental area of mice. Thus, this tool offers a new way of analyzing calcium imaging data from bistable neurons to understand how they participate in network computation and natural behaviors. Key Points SummaryCalcium imaging - the gold standard of inferring neuronal activity - does not report cellular state in neurons that are bistable, such as Purkinje neurons in the cerebellum of larval zebrafish. We model the relationship between Purkinje neuron electrical activity and its corresponding calcium signal to compile a dataset of state-labelled simulated calcium signals. We apply machine-learning methods to this dataset to develop a tool that can classify the state of a Purkinje neuron using only its calcium signal, which works well on real data even though it was trained only on simulated data. CaMLsort also generalizes well to bistable neurons in a different brain region (ventral tegmental area) in a different model organism (mouse). This tool offers a new way of analyzing calcium imaging data from populations of bistable neurons, thereby facilitating our understanding of how these neurons carry out their functions in a circuit.

neuroscience↗

bioRxiv2022

V2b neurons act via multiple targets in spinal motor networks

Within the spinal cord, interneurons shape motor neuron activity. These interneurons can project over long distances in the longitudinal axis, but systematic mapping of their connectivity has been limited. In this study, using larval zebrafish, we mapped local and long-range connectivity of a cardinal spinal population, the Gata3+ V2b class. V2b neurons are inhibitory and project ipsilateral, descending axons. We show that V2b neurons are active during fictive swimming, slightly leading the motor burst. Via optogenetic mapping of output in the rostrocaudal axis, we demonstrate that V2b neurons robustly inhibit motor neurons and other major spinal interneuron classes, including V2a, V1, commissural neurons and other V2b neurons. V2b inhibition is patterned along the rostrocaudal axis, providing long-range inhibition to motor and V2a neurons but more localized innervation of the V1 class. Furthermore, these results provide the first demonstration of reciprocal V1/V2b inhibition in axial circuits, potentially representing an ancestral motif of the limb control network.

neuroscience↗

bioRxiv2022

Transcriptomics based prediction of survival and response to therapy in malignant mesothelioma

Malignant mesothelioma is an aggressive cancer with limited treatment options and poor prognosis. Better understanding of mesothelioma genomics and transcriptomics could advance novel therapies. We performed whole-exome and RNA-sequencing of germline and tumors of 122 patients with pleural, peritoneal, and tunica-vaginalis mesothelioma. We identify a 48 gene prognostic signature that is highly predictive of mesothelioma patient survival including CCNB1, whose expression is highly predictive of patient survival on its own. Using a synthetic-lethality (SL) based pipeline for analyzing the patients transcriptomic data, we identified SL-based signatures predictive of response to an anti-PD1 immune checkpoint inhibitor and combination therapies with pemetrexed. These SL-profiles successfully predict the overall patient-response observed across targeted, immuno- and chemotherapies in 11 independent mesothelioma clinical trials spanning 7 different treatments. These findings lay a basis for future studies aimed specifically at testing the ability of these SL profiles to serve as treatment biomarkers in mesothelioma.

cancer biology↗

bioRxiv2021

Modelling spinal locomotor circuits for movements in developing zebrafish

Many spinal circuits dedicated to locomotor control have been identified in the developing zebrafish. How these circuits operate together to generate the various swimming movements during development remains to be clarified. In this study, we iteratively built models of developing zebrafish spinal circuits coupled to simplified musculoskeletal models that reproduce coiling and swimming movements. The neurons of the models were based upon morphologically or genetically identified populations in the developing zebrafish spinal cord. We simulated intact spinal circuits as well as circuits with silenced neurons or altered synaptic transmission to better understand the role of specific spinal neurons. Analysis of firing patterns and phase relationships helped identify possible mechanisms underlying the locomotor movements of developing zebrafish. Notably, our simulations demonstrated how the site and the operation of rhythm generation could transition between coiling and swimming. The simulations also underlined the importance of contralateral excitation to multiple tail beats. They allowed us to estimate the sensitivity of spinal locomotor networks to motor command amplitude, synaptic weights, length of ascending and descending axons, and firing behaviour. These models will serve as valuable tools to test and further understand the operation of spinal circuits for locomotion.

neuroscience↗

bioRxiv2021

Spinal V1 neurons inhibit motor targets locally and sensory targets distally to coordinate locomotion

Rostro-caudal coordination of spinal motor output is essential for locomotion. Most spinal interneurons project axons longitudinally to govern locomotor output, yet their connectivity along this axis remains unclear. In this study, we use larval zebrafish to map synaptic outputs of a major inhibitory population, V1 (Eng1+) neurons, which are implicated in dual sensory and motor functions. We find that V1 neurons exhibit long axons extending rostrally and exclusively ipsilaterally for an average of 6 spinal segments; however, they do not connect uniformly with their post-synaptic targets along the entire length of their axon. Locally, V1 neurons inhibit motor neurons (both fast and slow) and other premotor targets including V2a, V2b and commissural pre-motor neurons. In contrast, V1 neurons make robust inhibitory contacts throughout the rostral extent of their axonal projections onto a dorsal horn sensory population, the Commissural Primary Ascending neurons (CoPAs). In a computational model of the ipsilateral spinal network, we show that this pattern of short range V1 inhibition to motor and premotor neurons is crucial for coordinated rostro-caudal propagation of the locomotor wave. We conclude that spinal network architecture in the longitudinal axis can vary dramatically, with differentially targeted local and distal connections, yielding important consequences for function.

neuroscience↗

bioRxiv2020

CRTC1-MAML2 Establishes a PGC1α-IGF1 Circuit that Confers Vulnerability to PPARγ Inhibition

Mucoepidermoid carcinoma (MEC) is a life-threatening salivary gland cancer that is driven primarily by the transcriptional co-activator fusion CRTC1-MAML2. The mechanisms by which the chimeric CRTC1-MAML2 oncoprotein rewires gene expression programs that promote tumorigenesis remain poorly understood. Here, we show that CRTC1-MAML2 induces transcriptional activation of the non-canonical PGC-1 splice variant PGC-14, which regulates PPAR{gamma}-dependent IGF-1 expression. This mitogenic transcriptional circuitry is consistent across cell lines and primary tumors. CRTC1-MAML2 positive tumors are dominated by IGF-1 pathway activation and small molecule drug screens reveal that tumor cells harboring the fusion gene are selectively sensitive to IGF-1R inhibition. Furthermore, this dependence on autocrine regulation of IGF-1 transcription renders MEC cells susceptible to PPAR{gamma}inhibition with inverse agonists. These results yield insights into the aberrant co-regulatory functions of CRTC1-MAML2 and identify a specific vulnerability that can be exploited for precision therapy.

cancer biology↗

bioRxiv2019

Spinal V2b neurons reveal a role for ipsilateral inhibition in speed control

The spinal cord contains a diverse array of interneurons that govern motor output. Traditionally, models of spinal circuits have emphasized the role of inhibition in enforcing reciprocal alternation between left and right sides or flexors and extensors. However, recent work has shown that inhibition also increases coincident with excitation during contraction. Here, using larval zebrafish, we investigate the V2b (Gata3+) class of neurons, which contribute to flexor-extensor alternation but are otherwise poorly understood. Using newly generated transgenic lines we define two stable subclasses with distinct neurotransmitter and morphological properties. These two V2b subclasses make direct synapses onto motor neurons with differential targeting to slower and faster circuits. In vivo, optogenetic suppression of V2b activity leads to increases in locomotor speed. We conclude that V2b neurons exert speed-specific influence over axial motor circuits throughout the rostrocaudal axis. Together, these results indicate a new role for ipsilateral inhibition in speed control.

neuroscience↗

bioRxiv2016

Loss of the habenula neuromodulator Kisspeptin1 disrupts learning

Learning how to actively avoid a predictable aversive stimulus involves two steps: recognizing the cue that predicts upcoming punishment, and learning a behavioral response that will lead to avoidance. In zebrafish, ventral habenula (vHb) neurons have been proposed to participate in both steps by encoding the expected aversiveness of a stimulus. vHb neurons increase their firing rate as expectation of punishment grows, but reduce their activity as avoidance learning occurs. How the change in vHb activity occurs is not known. Here, we ask whether the neuromodulator kisspeptin1, which is expressed in the ventral habenula, could be involved. Kiss1 mutants were generated with Cas9 using guide RNAs targeted to the signal sequence. Mutants, which have a stop codon upstream of the kisspeptin1 peptide, have a deficiency in learning to avoid a shock that is predicted by light. Electrophysiology indicates that kisspeptin1 has a concentration-dependent effect on vHb neurons: depolarizing at low concentrations and hyperpolarizing at high concentrations. These data suggest that as the fish learns to cope with a threat, kisspeptin1 may differentially modulate vHb neurons. This implies that learning a behavioral strategy to overcome a stressor is accompanied by physiological change in habenula neurons.\n\nSignificance statementLearning to deal with adversity can positively affect ones ability to cope with challenges in the immediate future. Control thus causes short-term change in the brain. Here, we show that the neuromodulator kisspeptin1 is required to learn to avoid a punishment. Expression and electrophysiological recordings suggest that this molecule functions by controlling the ventral habenula, a region of the brain that mediates fear by regulating serotonin release. Kisspeptin1 may be a potential player in resilience developed as a result of control, extending previous findings that it can reduce fear.

animal behavior and cognition↗