Biology subjects

Ghoshal, A.

Publications and source records attributed to Ghoshal, A..

12 recordsLinked to original sources

bioRxiv2025

Discovery and structure-activity relationship analysis of 2-Pyridyl Thienopyrimidine derivatives as promising therapeutic candidates for the treatment of Buruli ulcer

Mycobacterium ulcerans, the bacterium causing Buruli ulcer (BU), can potentially develop resistance to existing antibiotics (rifampicin - clarithromycin/ moxifloxacin), underscoring the need for new antimycobacterial treatments. This study screened the Pathogen Box from Medicines for Malaria Venture (MMV) to identify M. ulcerans inhibitors. Four hit compounds were found, including the 2-(6-methylpyridin-2-yl)-N-(pyrimidin-4-yl)thieno[3,2-d]pyrimidin-4-amine MMV688122 as a novel anti-M. ulcerans chemotype. Synthesis of structural analogues of MMV688122 allowed the identification of 2-(4-methylpyridin-2-yl)-N-(pyrimidin-4-yl)thieno[3,2-d]pyrimidin-4-amine MMV1578877 as the most potent, with submicromolar activity. Importantly, this analogue was non-cytotoxic up to 100 {micro}M in human fibroblasts. Structure-activity relationship (SAR) studies indicated the crucial role of the methylpyridin-2-yl group in inhibiting M. ulcerans and the possibility to replace the thienopyrimidine core by a quinazoline. While MMV1578877 showed better metabolic stability than MMV688122, further improvement and testing in real-world M. ulcerans clinical isolates are still required. Further metabolite identification and SAR data should guide the optimization of this novel chemotype to enable in vivo testing. Author SummaryBuruli ulcer is a neglected tropical disease that causes severe skin ulcers and long-term disability, mostly affecting people in remote African communities. Current treatments rely on antibiotics that are not always effective and may lead to resistance. In this study, we searched for new drug candidates by testing a library of compounds provided by the Medicines for Malaria Venture (MMV). We discovered a promising new chemical family that can kill the bacteria responsible for Buruli ulcer in the lab. One compound, in particular, showed strong activity without harming human cells. This compound also showed better stability and effectiveness. These findings bring us closer to developing a new, safer, and more effective treatment for Buruli ulcer.

pharmacology and toxicology↗

bioRxiv2025

Species Dependent Metabolism of a Covalent nsP2 Protease Inhibitor with in Vivo Anti-alphaviral Activity

RA-0002034 (1) is a potent covalent inhibitor targeting the alphavirus nsP2 cysteine protease. The species-dependent pharmacokinetics and metabolism of 1 were investigated to evaluate its therapeutic potential. Pharmacokinetic profiling revealed rapid clearance in mice, predominantly mediated by glutathione S-transferase (GST)-catalyzed conjugation. This metabolic liability contrasted with slower clearance observed in human hepatocytes and preclinical species such as rats, dogs, and monkeys. Cross-species studies confirmed the dominance of GST-driven metabolism in mice, whereas oxidative pathways were more pronounced in dogs. Despite rapid systemic clearance, 1 achieved antiviral efficacy in mice, reducing CHIKV viral loads in multiple tissues. Initial estimations of human hepatic clearance and half-life extrapolated from animal data indicate that b.i.d. dosing of 1 will be possible to maintain concentrations sufficient for antiviral activity in humans. These cross-species pharmacokinetic and metabolism studies support the continued evaluation of 1 as a promising anti-alphaviral therapeutic. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=61 SRC="FIGDIR/small/632788v1_ufig1.gif" ALT="Figure 1"> View larger version (14K): org.highwire.dtl.DTLVardef@8c1558org.highwire.dtl.DTLVardef@cd7e21org.highwire.dtl.DTLVardef@113e88aorg.highwire.dtl.DTLVardef@7c7dc_HPS_FORMAT_FIGEXP M_FIG C_FIG

pharmacology and toxicology↗

bioRxiv2024

Deregulation of epigenetic factor, KDM6A, is involved during Candida albicans mediated delayed wound healing

Deregulation of wound healing process leads to chronic non-healing wounds. Opportunistic fungal pathogens like Candida albicans is known to colonize open wounds leading to delayed, insufficient, or disorganized healing. In our study, we investigate the regulatory mechanisms orchestrating this C. albicans mediated delay of wound healing and underscore a critical role for the transcription factor c-AMP Response Element Binding Protein (CREB) in regulating pro-inflammatory cytokines and genes implicated in wound healing. Parallelly, we found the epigenetic factor, KDM6A (Ubiquitously Transcribed tetratricopeptide repeat on chromosome X, also known as UTX) to be downregulated upon infection and modulating expression of metalloproteinases during C. albicans governed wound healing impairment. Mechanistically, we observed enhanced recruitment of EZH2, a methyl transferase, over the promoters of KDM6A during infection, hence, governing its expression. Our findings, therefore, underscore a role of KDM6A and CREB in C. albicans mediated delayed wound healing.

immunology↗

bioRxiv2024

Structure Activity of β-Amidomethyl Vinyl Sulfones as Covalent Inhibitors of Chikungunya nsP2 Cysteine Protease with Anti-alphavirus Activity

Despite their widespread impact on human health there are no approved drugs for combating alphavirus infections. The heterocyclic {beta}-aminomethyl vinyl sulfone RA-0002034 (1a) is a potent irreversible covalent inhibitor of the alphavirus nsP2 cysteine protease with broad spectrum antiviral activity. Analogs of 1a that varied each of three regions of the molecule were synthesized to establish structure-activity relationships for inhibition of Chikungunya (CHIKV) nsP2 protease and viral replication. The covalent warhead was highly sensitive to modifications of the sulfone or vinyl substituents. However, numerous alterations to the core 5-membered heterocycle and its aryl substituent were well tolerated and several analogs were identified that enhanced CHIKV nsP2 binding. For example, the 4-cyanopyrazole analog 8d exhibited a kinact/Ki ratio >10,000 M-1s-1. 3-Arylisoxazole was identified an isosteric replacement for the 5-membered heterocycle, which circumvented the intramolecular cyclization that complicated the synthesis of pyrazole-based inhibitors like 1a. The accumulated structure-activity data was used to build a ligand-based model of the enzyme active site, which can be used to guide the design of covalent nsP2 protease inhibitors as potential therapeutics against alphaviruses.

cell biology↗

bioRxiv2024

Discovery of a cell-active chikungunya virus nsP2 protease inhibitor using a covalent fragment-based screening approach

Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that has been responsible for numerous large-scale outbreaks in the last twenty years. Currently, there are no FDA-approved therapeutics for any alphavirus infection. CHIKV non-structural protein 2 (nsP2), which contains a cysteine protease domain, is essential for viral replication, making it an attractive target for a drug discovery campaign. Here, we optimized a CHIKV nsP2 protease (nsP2pro) biochemical assay for the screening of a 6,120-compound cysteine-directed covalent fragment library. Using a 50% inhibition threshold, we identified 153 hits (2.5% hit rate). In dose-response follow up, RA-0002034, a covalent fragment that contains a vinyl sulfone warhead, inhibited CHIKV nsP2pro with an IC50 of 58 {+/-} 17 nM, and further analysis with time-dependent inhibition studies yielded a kinact/KI of 6.4 x 103 M-1s-1. LC-MS/MS analysis determined that RA-0002034 covalently modified the catalytic cysteine in a site-specific manner. Additionally, RA-0002034 showed no significant off-target reactivity against a panel of cysteine proteases. In addition to the potent biochemical inhibition of CHIKV nsP2pro activity and exceptional selectivity, RA-0002034 was tested in cellular models of alphavirus infection and effectively inhibited viral replication of both CHIKV and related alphaviruses. This study highlights the discovery and characterization of the chemical probe RA-0002034 as a promising hit compound from covalent fragment-based screening for development toward a CHIKV or pan-alphavirus therapeutic. Significance StatementChikungunya virus is one of the most prominent and widespread alphaviruses and has caused explosive outbreaks of arthritic disease. Currently, there are no FDA-approved drugs to treat disease caused by chikungunya virus or any other alphavirus-caused infection. Here, we report the discovery of a covalent small molecule inhibitor of chikungunya virus nsP2 protease activity and viral replication of four diverse alphaviruses. This finding highlights the utility of covalent fragment screening for inhibitor discovery and represents a starting point towards the development of alphavirus therapeutics targeting nsP2 protease.

biochemistry↗

bioRxiv2023

Synergistic phenotypic shifts during domestication promote plankton-to-biofilm transition in purple sulfur bacterium Chromatium okenii

The ability to isolate microorganisms from natural environments to pure cultures under optimized laboratory settings has markedly improved our understanding of microbial ecology. Laboratory-induced artificial growth conditions often diverge from those in natural ecosystems, forcing wild isolates into selective pressures which are distinct compared to those in nature. Consequently, fresh isolates undergo diverse eco-physiological adaptations mediated by modification of key phenotypic traits. For motile microorganisms, we still lack a biophysical understanding of the relevant traits which emerge during domestication, and possible mechanistic interrelations between them which could ultimately drive short-to-long term microbial adaptation under laboratory conditions. Here, using microfluidics, atomic force microscopy (AFM), quantitative imaging, and mathematical modelling, we study phenotypic adaptation of natural isolates of Chromatium okenii, a motile phototrophic purple sulfur bacterium (PSB) common to meromictic settings, grown under ecologically-relevant laboratory conditions over multiple generations. Our results indicate that the naturally planktonic C. okenii populations leverage synergistic shifts in cell-surface adhesive interactions, together with changes in their cell morphology, mass density, and distribution of intracellular sulfur globules, to supress their swimming traits, ultimately switching to a sessile lifeform under laboratory conditions. A computational model of cell mechanics confirms the role of the synergistic phenotypic shifts in suppressing the planktonic lifeform. Over longer domestication periods ([~]10 generations), the switch from planktonic to sessile lifeform is driven by loss of flagella and enhanced adhesion. By investigating key phenotypic traits across different physiological stages of lab-grown C. okenii, we uncover a progressive loss of motility via synergistic phenotypic shifts during the early stages of domestication, which is followed by concomitant deflagellation and enhanced surface attachment that ultimately drive the transition of motile sulphur bacteria to a sessile biofilm state. Our results establish a mechanistic link between suppression of motility and surface attachment via synergistic phenotypic changes, underscoring the emergence of adaptive fitness under felicitous laboratory conditions that comes at a cost of lost ecophysiological traits tailored for natural environments. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=85 SRC="FIGDIR/small/563228v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@fcb6b0org.highwire.dtl.DTLVardef@13e18d0org.highwire.dtl.DTLVardef@1cf3ee6org.highwire.dtl.DTLVardef@12a837b_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2023

Phytoplankton tune local pH to actively modulate circadian swimming behavior

Diel vertical migration (DVM), the diurnal exodus of motile phytoplankton between the light- and nutrient-rich aquatic regions, is governed by endogenous biological clocks. Many species exhibit irregular DVM patterns wherein out-of-phase gravitactic swimming-relative to that expected due to the endogenous rhythm-is observed. How cells achieve and control this irregular swimming behavior, and its impact on biological fitness remain poorly understood. Combining local environmental monitoring with behavioral and physiological analyses of motile bloom-forming Heterosigma akashiwo cells, we report that phytoplankton species modulate their DVM pattern by progressively tuning local pH, yielding physiologically equivalent yet behaviorally distinct gravitactic sub-populations which remain separated vertically within a visibly homogeneous cell distribution. Individual and population-scale tracking of the isolated top and bottom sub-populations revealed similar gravitactic (swimming speed and stability) and physiological traits (growth rate and maximum photosynthetic yield), suggesting that the sub-populations emerge due to mutual co-existence. Exposing the top (bottom) sub-population to the spent media of the bottom (top) counterpart recreates the emergent vertical distribution, while no such phenomenon was observed when the sub-populations were exposed to their own spent media. A model of swimming mechanics based on the quantitative analysis of cell morphologies confirms that the emergent sub-populations represent distinct swimming stabilities, resulting from morphological transformations after the cells are exposed to the spent media. Together with the corresponding night-time dataset, we present an integrated picture of the circadian swimming, wherein active chemo-regulation of the local environment underpins motility variations for potential ecological advantages via intraspecific division of labor over the day-night cycle. This chemo-regulated migratory trait offers mechanistic insights into the irregular diel migration, relevant particularly for modelling phytoplankton transport, fitness and adaptation as globally ocean waters see a persistent drop in the mean pH. One sentence summaryActive regulation of local pH diversifies the diel vertical migration of motile phytoplankton.

biophysics↗

bioRxiv2021

Active reconfiguration of cytoplasmic lipid droplets governs migration of nutrient-limited phytoplankton

As open oceans continue to warm, modified currents and enhanced stratification exacerbate nitrogen and phosphorus limitation, constraining primary production. The ability to migrate vertically bestows motile phytoplankton a crucial-albeit energetically expensive-advantage toward vertically redistributing for optimal growth, uptake and resource storage in nutrient-limited water columns. However, this traditional view discounts the possibility that the phytoplankton migration strategy may be actively selected by the storage dynamics when nutrients turn limiting. Here we report that storage and migration in phytoplankton are coupled traits, whereby motile species harness energy storing lipid droplets (LDs) to biomechanically regulate migration in nutrient limited settings. LDs grow and translocate-directionally-within the cytoplasm to accumulate below the cell nucleus, tuning the speed, trajectory and stability of swimming cells. Nutrient reincorporation reverses the LD translocation, restoring the homeostatic migratory traits measured in population-scale millifluidic experiments. Combining intracellular LD tracking and quantitative morphological analysis of red-tide forming alga, Heterosigma akashiwo, along with a model of cell mechanics, we discover that the size and spatial localization of growing LDs govern the ballisticity and orientational stability of migration. The strain-specific shifts in migration which we identify here are amenable to a selective emergence of mixotrophy in nutrient-limited phytoplankton. We rationalize these distinct behavioral acclimatization in an ecological context, relying on concomitant tracking of the photophysiology and reactive oxygen species (ROS) levels, and propose a dissipative energy budget for motile phytoplankton alleviating nutrient limitation. The emergent resource acquisition strategies, enabled by distinct strain-specific migratory acclimatizing mechanisms, highlight the active role of the reconfigurable cytoplasmic LDs in guiding vertical movement. By uncovering the mechanistic coupling between dynamics of intracellular changes to physiologically-governed migration strategies, this work offers a tractable framework to delineate diverse strategies which phytoplankton may harness to maximize fitness and resource pool in nutrient-limited open oceans of the future. One sentence summaryPhytoplankton harness reconfigurable lipid droplets to biomechanically tune migratory strategies in dynamic nutrient landscapes.

biophysics↗

bioRxiv2021

Zinc2+ ion inhibits SARS-CoV-2 main protease and viral replication in vitro.

Zinc deficiency is linked to poor prognosis in COVID-19 patients while clinical trials with Zinc demonstrate better clinical outcome. The molecular target and mechanistic details of anti-coronaviral activity of Zinc remain obscure. We show that ionic Zinc not only inhibits SARS-CoV-2 main protease (Mpro) with nanomolar affinity, but also viral replication. We present the first crystal structure of Mpro-Zn2+ complex at 1.9 [A] and provide the structural basis of viral replication inhibition. We show that Zn2+ coordinates with the catalytic dyad at the enzyme active site along with two previously unknown water molecules in a tetrahedral geometry to form a stable inhibited Mpro-Zn2+ complex. Further, natural ionophore quercetin increases the anti-viral potency of Zn2+. As the catalytic dyad is highly conserved across SARS-CoV, MERS-CoV and all variants of SARS-CoV-2, Zn2+ mediated inhibition of Mpro may have wider implications.

biochemistry↗

bioRxiv2021

Obesity-instructed TREM2high macrophages identified by comparative analysis of diabetic mouse and human kidney at single cell resolution

Mouse models are a tool for studying the mechanisms underlying complex diseases; however, differences between species pose a significant challenge for translating findings to patients. Here, we used single-cell transcriptomics and orthogonal validation approaches to provide cross-species taxonomies, identifying shared broad cell classes and unique granular cellular states, between mouse and human kidney. We generated cell atlases of the diabetic and obese kidney using two different mouse models, a high-fat diet (HFD) model and a genetic model (BTBR ob/ob), at multiple time points along disease progression. Importantly, we identified a previously unrecognized, expanding Trem2high macrophage population in kidneys of HFD mice that matched human TREM2high macrophages in obese patients. Taken together, our cross-species comparison highlights shared immune and metabolic cell-state changes.

genomics↗

bioRxiv2020

Fright and Fight: Role of predation and competition on mate search tactics of wild male zebrafish

Mate search tactics and association preferences among organisms in natural habitats can be dynamic and are determined by inherent trait preferences as well as the cost-benefit trade-offs associated with each mating decision. Two of the prime factors regulating mating decisions are the presence of competing conspecifics and predatory threats, both of which have important fitness consequences for the individual. We studied the influence of these two factors separately in mate search tactics and association preferences among zebrafish males. Male zebrafish were presented with a choice of two patches, consisting of different number of females, of which one patch was also associated with a predatory threat. We found that males made a preferential choice for the patch with more number of females only when the numerical difference between choices are starkly different, irrespective of the predatory threat associated with the patch. This points towards the role of numerical cognition in assessing cost-benefit tradeoffs in male zebrafish. We also studied the association preference of males in a multi-choice setup, consisting of four separate mixed-sex groups of zebrafish varying in densities. Our results showed that while test males preferred to visit the male-biased patches more often, they spent more time near female-biased patches or patches with equal sex ratio patches indicating the role of complex interplay of social cues in determining the associative behavior of males to a patch. This study, thus, sheds further light on the interactive roles of social cues and cognitive abilities in mate association patterns in this species.

animal behavior and cognition↗

bioRxiv2020

What drives male association tendencies in wild zebrafish? Role of female and vegetation densities

Mating strategies in species is context-dependent and driven by several ecological and demographic factors. In natural habitats, a multitude of ecological factors interact and these eventually determine mating preferences and mate choice decisions among species. While zebrafish (Danio rerio) is a widely studied biological model organism, our understanding of their mating preferences, strategies and their underlying ecological drivers are still limited. Here, we explore the role of ecological factors such as spatial variation (or patchiness) in mate density and associated vegetation cover in determining mate association tactics among males in a wild zebrafish population. We employed a multi-choice experimental design for a better representation of ecologically relevant scenarios. Our results revealed that when presented with patches with increasing female densities, males displayed only a marginal increase in preference for higher female density patches. However, when female density varied concomitantly with variation in vegetation cover, the males associated more with higher foliage density patches irrespective of the female density in that patch. Our findings throw light on the complex interaction between these two most basic ecological factors in determining mate search strategies and mate associations among these group-living fish species.

animal behavior and cognition↗