Biology subjects

Dasgupta, A.

Publications and source records attributed to Dasgupta, A..

At least 19 recordsLinked to original sources

bioRxiv2026

Stearic acid enhances membrane fluidization and peptidoglycan stiffness to promote the stability of Gram-positive bacteria

Saturated fatty acids such as stearic acid (SA) can exhibit both antimicrobial and growth-promoting effects on bacteria, depending on their concentration and chemical structure. However, the physical properties of the bacterial cell envelope in response to such molecules remain under-explored compared to their biochemical pathways. In this study, a comprehensive investigation is presented on the interaction of SA with the Gram-positive bacterium, Staphylococcus epider-midis (S. epi). SA alters bacterial growth, reflected in a higher maximum specific growth rate, a shorter lag phase, and an extended exponential phase, consistent with a prebiotic effect. Using fluorescence correlation spectroscopy and fluorescence lifetime imaging microscopy, we show that SA incorporation leads to significant fluidization of the lipid membrane, characterized by enhanced lateral diffusion and reduced membrane viscosity. Coarse-grained molecular dynamics (CG-MD) simulations demonstrate spontaneous insertion of SA into the membrane and a significant increase in mean-square displacement after insertion, supporting our experimental observations. Importantly, atomic force microscopy measurements show an increase in cell-envelope stiffness, reflected by a higher Youngs modulus which can be attributed to modulations in the glycan-peptide linkage density based on earlier studies that correlate stiffness changes to peptidoglycan (PG) crosslinking in Gram-positive strains [1]. These results provide direct evidence linking membrane fluidization induced by SA and increased cell wall stiffness due to transport modifications in the membrane mediated PG synthesis pathways to enhance bacterial cell viability.

biophysics↗

bioRxiv2026

Adaptive evolution of Topoisomerase II triggers reproductive isolation in Drosophila

A fundamental driver of biological diversification is the evolution of reproductive barriers between species. Instability and mis-regulation of repetitive DNA underlie numerous post-zygotic reproductive barriers, yet the molecular mechanisms are unknown. A long-studied genetic incompatibility between Drosophila melanogaster and D. simulans arises from mis-segregation of the D. melanogaster-specific 359bp DNA satellite in hybrid embryos. Here we report that the D. simulans version of the essential enzyme Topoisomerase II/Top2 causes this lethal incompatibility. Combining interspecies gene swaps with cell biology and genetics revealed that D. simulans-specific adaptive divergence of Top2 DNA-interacting domains prevents the resolution of 359bp-induced topological stress. Our findings demonstrate that species-specific DNA satellite topology requires species-specific molecular machinery and that even vital housekeeping genes can underlie reproductive isolation between closely related species.

evolutionary biology↗

bioRxiv2025

Probabilistic Multi-site MR Image Harmonization via Feature Preserving Conditional Generative Adversarial Networks

Brain magnetic resonance imaging (MRI) is pivotal in diagnosing and monitoring neurological disorders. However, despite their extensive applications, MR images have certain shortcomings. In particular, factors other than the anatomy of brain tissues influence the intensity distribution of voxels in MR images. These factors include hardware, software, magnetic field strength, and acquisition protocol. This inconsistency poses challenges in multi-site neuroimaging studies, where images are obtained from various devices with minimal control over acquisition parameters. Image harmonization algorithms aim to eliminate non-biological characteristics in MR images through various approaches, including converting images from multiple sites into a format resembling that of a designated target site. Among image harmonization methods, those relying on deep learning algorithms have gained significant attention recently. Nevertheless, certain aspects of deep learning-based image harmonization remain unexplored, notably the integration of probabilistic deep generative models to transform the distribution of MR images to a desired distribution. Inspired by this, we introduced a feature preserving conditional generative adversarial network (FP-cGAN) that converts images from multiple origins into the format of a target site while preserving anatomical features by imposing a novel regularizing constraint. We conduct our experiments on MR images from the SRPBS dataset, which comprises unpaired images in addition to paired (traveling subjects) images from multiple sites. We utilize the unpaired data for training our models and the paired data for evaluation. Furthermore, we compare our results with cycleGAN and histogram matching, two widely used image harmonization methods. Our experiments reveal that our approach surpasses the other techniques. Article HighlightsO_LIWe introduce an FP-cGAN model to harmonize multi-site MRI scans into a target site. C_LIO_LIA novel anatomical feature invariant constraint maintains tissue integrity. C_LIO_LIThe models are trained in an unpaired setting without requiring traveling subjects. C_LIO_LIWe evaluate harmonization performance using paired traveling subjects. C_LI

bioengineering↗

bioRxiv2025

Benzopyrene degrading Bacillus altitudinis AR19 isolated from Digboi oil refinery soil (India)

Benzo[a]pyrene (B[a]P) is a five-ring polycyclic aromatic hydrocarbon (PAH) recognized as a potent carcinogen and persistent environmental pollutant, highly resistant to microbial degradation. In this study, a B[a]P-degrading and biosurfactant producing bacterium, Bacillus altitudinis AR19, was isolated from the Digboi oil refinery site in Assam, India. The isolate was screened using the spray plate technique, and its morphological features and cell viability were examined by scanning electron microscopy (SEM) and flow cytometry. Biosurfactant production was confirmed through drop collapse, oil spreading, and emulsification assays. Taxonomic identity was established using average nucleotide identity (ANI), and comparative pangenome analysis was performed among B. altitudinis (Taxonomy ID: 1178544), Bacillus pumilus (Taxonomy ID: 1408), Mycolicibacterium vanbaalenii (Taxonomy ID: 110539), and B. altitudinis AR19. The results highlight that several mechanisms including osmolyte accumulation, biosurfactant production, and mechanosensitive ion channels play key roles in enabling B. altitudinis AR19 to tolerate and degrade B[a]P. This study demonstrates the potential of B. altitudinis AR19 as a promising candidate for bioremediation of PAH-contaminated environments. Importance of the workThis manuscript describes the morphological and genomic characterization of Bacillus altitudinis AR19, a bacterium capable of degrading benzo[a]pyrene. Benzo[a]pyrene is a five-ring polycyclic aromatic hydrocarbon that is highly recalcitrant to bacterial remediation, making this work important for advancing bioremediation strategies.

microbiology↗

bioRxiv2025

Inference of VEGFR2 dimerization kinetics on the cell surface by integrating single-molecule imaging and mathematical modeling

Inter-receptor interactions play a key role in receptor signaling, which is the first step in cell signaling in response to external stimuli. In the case of Vascular Endothelial Growth Factor Receptor 2 (VEGFR2), dimerization is necessary for activation. VEGFR2 undergoes reversible interactions also in the absence of ligand. For a quantitative understanding of transmembrane signal transduction, it is necessary to quantify the interaction kinetics of VEGFR2 on the cell surface. Live-cell single-molecule imaging (SMI) has the powerful ability to capture receptor interaction events in their native cellular environment with high spatiotemporal resolution. However, it reveals these interactions for only the labeled subset, which is a small fraction of the full population of receptors. We have previously shown that mathematical modeling, combined with SMI data, offers a route to compensate for this lost information and infer the population-level receptor interaction kinetics from SMI data. Here, we applied this approach to VEGFR2, both wildtype full length VEGFR2 (FLR2) and a truncated mutant consisting of its extracellular and transmembrane domains (ECTM), which has been shown to exhibit reduced homotypic interactions in the absence of ligand. We developed a stochastic mathematical model mimicking VEGFR2 diffusion and interactions and determined the unknown model parameters through a combination of direct experimental measurements and stochastic model calibration. We found that a model of dimerization was sufficient to describe VEGFR2 interactions in the absence of ligand. While ECTM was primarily monomeric, FLR2 exhibited a substantial fraction of dimers. Our inference revealed that the difference between FLR2 and ECTM was primarily in the dimer association rate constant, which was about an order of magnitude lower for ECTM than for FLR2. To our knowledge, this is the first time that the interaction kinetics of VEGFR2 have been calculated in live cells. Author summaryInteractions between receptors on the cell surface play a large role in the initiation and modulation of cell signaling. For example, the receptor VEGFR2 in vascular endothelial cells must dimerize as a first step toward signaling. However, many interactions, including VEGFR2 dimerization, also take place in the absence of ligand stimulation. Therefore, elucidating when and where receptor interactions take place, and what their kinetics are, is critical for understanding receptor and cell signaling, especially on a quantitative level. Determining interaction kinetics is particularly challenging, given that membrane proteins do not express well in vitro, and are thus not amenable to traditional biophysical methods that quantify molecular interactions. In addition, the membrane and cellular environment is much more complex than an in vitro setting. To address these challenges, we combine single-molecule imaging, stochastic mathematical modeling, and rigorous model calibration to derive the dimerization kinetics of VEGFR2 (both full-length and a truncation mutant with reduced dimerization) on the surface of live cells. To our knowledge, this is the first time that the interaction kinetics of VEGFR2 have been calculated in live cells. Our approach is generalizable to many other cell surface proteins, opening an avenue for quantifying molecular interaction kinetics in situ.

biophysics↗

bioRxiv2025

KIF5B and Dynein regulate adhesion-dependent Golgi organization and microtubule acetylation

Cell-matrix adhesion regulates Golgi organization through Arf1-mediated dynein recruitment, maintaining its juxtanuclear localization. On loss of adhesion, Arf1 activation drops, causing loss of dynein, promoting differential disorganization of cis- vs trans-Golgi along microtubules. Golgi regulates microtubule nucleation and stability. In fibroblasts, acetylated tubulin levels drop on loss of adhesion, recovering on re-adhesion with time. Active Arf1 overexpression in preventing Golgi disorganization sustains microtubule acetylation, also seen in T24 bladder cancer cells. Active Arf1 binds KIF5B, recruiting it to the Golgi. KIF5B and dynein knockdown disorganize the Golgi as ministacks, with cis- and trans-Golgi. Dynein knockdown disrupts MTOC positioning, causing ministacks to disperse, preventing Golgi reorganization upon re-adhesion. Dispersed ministacks interestingly maintain microtubule acetylation in adherent and non-adherent cells. The joint KIF5B-dynein knockdown causes the Golgi to lose its ribbon morphology, becoming compact while keeping cis- and trans-Golgi together. This also causes a change in spreading, aspect ratio and migration of knockdown cells, which could be regulated by their Golgi phenotype. In evaluating adhesion-dependent Golgi organization, we reveal the Arf1-KIF5B-dynein crosstalk to regulate Golgi-dependent tubulin acetylation and cell function. SummaryKIF5B and dynein are vital microtubule-associated motors that drive organelle positioning and organization. Adhesion-dependent Arf1 activation mediates KIF5B and dyneins recruitment to the Golgi, regulating its organization and position. This, in turn, regulates microtubule acetylation levels, localization, and cellular functions.

cell biology↗

bioRxiv2025

Integrins and tetraspanins mediate CD36-actin coupling and regulate CD36 organization and signaling

The actin cortex plays a large role in regulating the dynamic organization of cell surface receptors, which in turn regulates their signaling. However, many receptors have short intracellular domains and no known link to cortical actin. In this work, we identified the {beta}1-integrin subunit and several tetraspanins - CD9, CD81 and CD151 - as part of the hitherto unknown molecular link between the surface receptor CD36 and cortical actin. We found that CD36 in vascular endothelial cells is recruited into complexes/nanodomains containing these proteins, with stronger recruitment near the cell edge. Perturbing this recruitment via the mutation G12V in the N-terminal transmembrane domain of CD36 alters the dynamic organization of CD36 on the vascular endothelial cell surface and weakens its coupling to cortical actin dynamics. Moreover, perturbing this recruitment abolishes thrombospondin-1-induced CD36 signaling through the Src family kinase Fyn. Given their many interactions with other transmembrane proteins, tetraspanins and integrins may provide a ubiquitous mechanism for plasma membrane-cortical actin coupling.

cell biology↗

bioRxiv2025

In vitro and in vivo synergy of Vancomycin and β-lactams against drug-resistant Mycobacterium tuberculosis and Non-tuberculous mycobacteria

The unabated increase in antimicrobial resistance has underlined the importance of identifying novel drug combinations which eliminate infections more potently and likely reduce the emergence of resistance. In this context, we have identified Vancomycin and many {beta}-lactams as being potently active against drug-resistant Mycobacterium tuberculosis and non-tuberculous mycobacteria including M. abscessus, emerging as pathogens of concern owing to their inherent drug resistance profile. In this study, we have identified combinations of Vancomycin, a glycopeptide and {beta}-lactams, especially Ceftriaxone, Ceftazidime and Meropenem, in the presence or absence of Sulbactam, a {beta}- lactamase inhibitor, as possessing potent antimicrobial activity against several drug-resistant mycobacterial strains. The combination of Vancomycin and {beta}-lactams exhibited potent bactericidal activity and reduced the bacterial load better than either drug alone. The molecular basis of synergy was mediated by increase in permeability of mycobacterial cell as demonstrated by ethidium bromide assay. In the murine model of mycobacterial infection, synergistic combination of Vancomycin and {beta}-lactams outperformed clinically utilized drugs including Isoniazid, Rifampicin and Ethambutol against M. tuberculosis and Amikacin, Clarithromycin against M. abscessus. The combinations caused a significant reduction in bacterial load in various organs in M. tuberculosis and M. abscessus infected mice. Thus, the synergistic combination of Vancomycin and {beta}-lactams could potentially be utilized for treatment of recalcitrant mycobacterial infection especially those caused due to drug-resistant pathogens.

microbiology↗

bioRxiv2025

Unsupervised Detection of Rare Events in Liquid Biopsy Assays

The use of liquid biopsies in the detection, diagnosis and treatment monitoring of different types of cancers and other diseases often requires identifying and enumerating instances of analytes that are rare. Most current techniques that aim to computationally isolate these rare instances or events first learn the signature of the event, and then scan the appropriate biological assay for this signature. While such techniques have proven to be very useful, they are limited because they must first establish what signature to look for, and only then identify events that are consistent with this signature. In contrast to this, in this study, we present an automated approach that does not require the knowledge of the signature of the rare event. It works by breaking the assay into a sequence of components, learning the probability distribution of these components, and then isolating those that are rare. This is done with the help of deep generative algorithms in an unsupervised manner, meaning without a-priori knowledge of the rare event associated with an analyte. In this study, this approach is applied to immunofluorescence microscopy images of peripheral blood, where it is shown that it successfully isolates biologically relevant events in blood from normal donors spiked with cancer-related cells and in blood from patients with late-stage breast cancer.

bioinformatics↗

bioRxiv2024

Essential genes encoded by the mating-type locus of the human fungal pathogen Cryptococcus neoformans

Fungal sexual reproduction is controlled by the mating-type (MAT) locus. In contrast to a majority of species in the phylum Basidiomycota that have tetrapolar mating-type systems, the opportunistic human pathogen Cryptococcus neoformans employs a bipolar mating-type system, with two mating types (a and ) determined by a single MAT locus that is unusually large ([~]120 kb) and contains more than 20 genes. While several MAT genes are associated with mating and sexual development, others control conserved cellular processes (e.g. cargo transport and protein synthesis), of which five (MYO2, PRT1, RPL22, RPL39, and RPO41) have been hypothesized to be essential. In this study, through genetic analysis involving sporulation of heterozygous diploid deletion mutants, as well as in some cases construction and analyses of conditional expression alleles of these genes, we confirmed that with the exception of MYO2, both alleles of the other four MAT genes are indeed essential for cell viability. We further showed that while MYO2 is not essential, its function is critical for infectious spore production, faithful cytokinesis, adaptation for growth at high temperature, and pathogenicity in vivo. Our results demonstrate the presence of essential genes in the MAT locus that are divergent between cells of opposite mating types. We discuss possible mechanisms to maintain functional alleles of these essential genes in a rapidly-evolving genomic region in the context of fungal sexual reproduction and mating-type evolution. IMPORTANCESexual reproduction is essential for long-term evolutionary success. Fungal cell type identity is governed by the MAT locus, which is typically rapidly evolving and highly divergent between different mating types. In this study, we show that the a and alleles of four genes encoded in the MAT locus of the opportunistic human fungal pathogen C. neoformans are essential. We demonstrate that a fifth gene, MYO2, which had been predicted to be essential, is in fact dispensable for cell viability. However, a functional MYO2 allele is important for cytokinesis and fungal pathogenicity. Our study highlights the need for careful genetic analyses in determining essential genes, which is complementary to high-throughput approaches. Additionally, the presence of essential genes in the MAT locus of C. neoformans provides insights into the function, maintenance, and evolution of these fast-evolving genomic regions.

genetics↗

bioRxiv2024

Human-mouse proteomics reveals the shared pathways in Alzheimer's disease and delayed protein turnover in the amyloidome

Murine models of Alzheimers disease (AD) are crucial for elucidating disease mechanisms but have limitations in fully representing AD molecular complexities. We comprehensively profiled age-dependent brain proteome and phosphoproteome (n > 10,000 for both) across multiple mouse models of amyloidosis. We identified shared pathways by integrating with human metadata, and prioritized novel components by multi-omics analysis. Collectively, two commonly used models (5xFAD and APP-KI) replicate 30% of the human protein alterations; additional genetic incorporation of tau and splicing pathologies increases this similarity to 42%. We dissected the proteome-transcriptome inconsistency in AD and 5xFAD mouse brains, revealing that inconsistent proteins are enriched within amyloid plaque microenvironment (amyloidome). Determining the 5xFAD proteome turnover demonstrates that amyloid formation delays the degradation of amyloidome components, including A{beta}-binding proteins and autophagy/lysosomal proteins. Our proteomic strategy defines shared AD pathways, identify potential new targets, and underscores that protein turnover contributes to proteome-transcriptome discrepancies during AD progression.

biochemistry↗

bioRxiv2024

An Extensive Atlas of Proteome and Phosphoproteome Turnover Across Mouse Tissues and Brain Regions

Understanding how proteins in different mammalian tissues are regulated is central to biology. Protein abundance, turnover, and post-translational modifications like phosphorylation, are key factors that determine tissue-specific proteome properties. However, these properties are challenging to study across tissues and remain poorly understood. Here, we present Turnover-PPT, a comprehensive resource mapping the abundance and lifetime of 11,000 proteins and 40,000 phosphosites across eight mouse tissues and various brain regions, using advanced proteomics and stable isotope labeling. We revealed tissue-specific short- and long-lived proteins, strong correlations between interacting protein lifetimes, and distinct impacts of phosphorylation on protein turnover. Notably, we discovered that peroxisomes are regulated by protein turnover across tissues, and that phosphorylation regulates the stability of neurodegeneration-related proteins, such as Tau and -synuclein. Thus, Turnover-PPT provides new fundamental insights into protein stability, tissue dynamic proteotypes, and the role of protein phosphorylation, and is accessible via an interactive web-based portal at https://yslproteomics.shinyapps.io/tissuePPT.

systems biology↗

bioRxiv2023

Efficacy of Drpitor1a, a Dynamin-Related Protein 1 inhibitor, in Pulmonary Arterial Hypertension

RationaleDynamin-related protein 1 (Drp1), a large GTPase, mediates mitochondrial fission. Increased Drp1-mediated fission permits accelerated mitosis, contributing to hyperproliferation of pulmonary artery smooth muscle cells (PASMC), which characterizes pulmonary arterial hypertension (PAH). We developed a Drp1 inhibitor, Drpitor1a, and tested its ability to regress PAH. ObjectivesAssess Drpitor1as efficacy and toxicity in: a)normal and PAH human PASMC (hPASMC); b)normal rats versus rats with established monocrotaline (MCT)-induced PAH. MethodsDrpitor1as effects on recombinant and endogenous Drp1-GTPase activity, mitochondrial fission, and cell proliferation were studied in hPASMCs (normal=3; PAH=5). Drpitor1as pharmacokinetics and tissue concentrations were measured (n=3 rats/sex). In a pilot study (n=3-4/sex/dose), Drpitor1a (1mg/kg/48-hours, intravenous) reduced adverse PA remodeling only in females. Consequently, we compared Drpitor1a to vehicle in normal (n=6 versus 8) and MCT-PAH (n=9 and 11) females, respectively. Drpitor1a treatment began 17-days post-MCT with echocardiography and cardiac catheterization performed 28-29 days post-MCT. ResultsDrpitor1a inhibited recombinant and endogenous Drp1 GTPase activity, which was increased in PAH hPASMC. Drpitor1a inhibited mitochondrial fission and proliferation and induced apoptosis, in PAH hPASMC but not normal hPASMC. Drpitor1a tissue levels were higher in female versus male RVs. In MCT-PAH females, Drpitor1a regressed PA obstruction, lowered pulmonary vascular resistance, and improved RV function, without hematologic, renal, or hepatic toxicity. ConclusionsDrpitor1a inhibits Drp1 GTPase, reduces mitochondrial fission, and inhibits cell proliferation in PAH hPASMC. Drpitor1a caused no toxicity in MCT-PAH and had no significant effect on normal rats or hPASMCs. Drpitor1a is a potential PAH therapeutic which displays an interesting therapeutic sexual dimorphism.

pharmacology and toxicology↗

bioRxiv2023

In Silico Therapeutic Intervention on Cytokine Storm in COVID-19

The recent global COVID-19 outbreak, attributed by the World Health Organization to the rapid spread of the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), underscores the need for an extensive exploration of virological intricacies, fundamental pathophysiology, and immune responses. This investigation is vital to unearth potential therapeutic avenues and preventive strategies. Our study delves into the intricate interaction between SARS-CoV-2 and the immune system, coupled with exploring therapeutic interventions to counteract dysfunctional immune responses like the cytokine storm (CS), a driver of disease progression. Understanding these immunological dimensions informs the design of precise multiepitopetargeted peptide vaccines using advanced immunoinformatics and equips us with tools to confront the cytokine storm. Employing a control theory-based approach, we scrutinize the perturbed behavior of key proteins associated with cytokine storm during COVID-19 infection. Our findings support ACE2 activation as a potential drug target for CS control and confirm AT1R inhibition as an alternative strategy. Leveraging deep learning, we identify potential drugs to individually target ACE2 and AT1R, with Lomefloxacin and Fostamatinib emerging as standout options due to their close interaction with ACE2. Their stability within the protein-drug complex suggests superior efficacy among many drugs from our deep-learning analysis. Moreover, there is a significant scope for optimization in fine-tuning protein-drug interactions. Strong binding alone may not be the sole determining factor for potential drugs; precise adjustments are essential. The application of advanced computational power offers novel solutions, circumventing time-consuming lab work. In scenarios necessitating both ACE2 and AT1R targeting, optimal drug combinations can be derived from our analysis of drug-drug interactions, as detailed in the manuscript.

bioinformatics↗

bioRxiv2023

A phosphorylation-deficient ribosomal protein eS6 is largely functional in Arabidopsis thaliana, rescuing mutant defects from global translation and gene expression to photosynthesis and growth

The eukaryote-specific ribosomal protein of the small subunit eS6 is phosphorylated through the Target of rapamycin (TOR) kinase pathway. Although this phosphorylation event responds dynamically to environmental conditions and has been studied for over 50 years, its biochemical and physiological significance remains controversial and poorly understood. Here we report data from Arabidopsis thaliana, which indicate that plants expressing only a largely phospho-deficient isoform of eS6 grow essentially normally under laboratory conditions. The eS6A (RPS6A) paralog of eS6 functionally rescued double mutations in both rps6a and rps6b genes when expressed at approximately twice the wild-type dosage. A mutant isoform of eS6A lacking the major six phosphorylatable serine and threonine residues in its carboxyl-terminal tail also rescued the lethality, rosette growth, and polyribosome loading of the double mutant. It also complemented many mutant phenotypes of rps6 that were newly characterized here, including photosynthetic efficiency, and the vast majority of gene expression defects that were measured by transcriptomics and proteomics. However, compared to plants rescued with a phospho-enabled version of eS6A, the phospho-deficient seedlings retained a mild pointed-leaf phenotype, root growth was reduced, and certain cell cycle related mRNAs and ribosome biogenesis proteins were misexpressed. The residual defects of the phospho-deficient seedlings could be understood as an incomplete rescue of the rps6 mutant defects, with little or no evidence for gain-of-function defects. As expected, the phospho-deficient eS6A also rescued the rps6a and rps6b single mutants; however, phosphorylation of the eS6B paralog remained lower than predicted, further underscoring that plants can tolerate phospho-deficiency of eS6 well. Our data also yield new insights into how plants cope with mutations in essential, duplicated ribosomal protein isoforms.

plant biology↗

bioRxiv2023

Salmonella Typhimurium effector SseI regulates host peroxisomal dynamics to acquire lysosomal cholesterol for better intracellular growth

Intracellular Salmonella resides and multiplies in cholesterol-rich specialized compartment called Salmonella-containing vacuoles (SCVs) and avoids fusion with acidic lysosomes. Given, lysosomes are primary organelle that redistributes LDL derived cholesterol to other organelles; we questioned how lysosomal cholesterol can be transported to SCV. We demonstrate here that peroxisomes are recruited to SCVs in human primary macrophages, epithelial cells and functions as pro-bacterial organelles. Further, this interaction is assisted by SseI, a Salmonella effector protein containing mammalian peroxisome targeting sequence. SseI localizes to peroxisome, interacts and activates a host Ras GTPase, ARF-1 on the peroxisome membrane. Activation of ARF-1 leads to recruitment of phosphatidylinsolitol-5- phosphate-4 kinase to generate phosphatidylinsolitol-4-5-bisphosphate on peroxisomes. Accordingly, the{Delta} sseI strain showed reduced virulence in cell lines and during mice infection. Taken together, our work identified a fascinating mechanism by which a pathogen targets host organelles via its secretory effectors and exploits host metabolic intermediates for its intracellular proliferation.

cell biology↗

bioRxiv2023

A thin-film optogenetic visual prosthesis

Retinitis pigmentosa and macular degeneration lead to photoreceptor death and loss of visual perception. Despite recent progress, restorative technologies for photoreceptor degeneration remain largely unavailable. Here, we describe a novel optogenetic visual prosthesis (FlexLED) based on a combination of a thin-film retinal display and optogenetic activation of retinal ganglion cells (RGCs). The FlexLED implant is a 30 {micro}m thin, flexible, wireless {micro}LED display with 8,192 pixels, each with an emission area of 66 {micro}m2. The display is affixed to the retinal surface, and the electronics package is mounted under the conjunctiva in the form factor of a conventional glaucoma drainage implant. In a rabbit model of photoreceptor degeneration, optical stimulation of the retina using the FlexLED elicits activity in visual cortex. This technology is readily scalable to hundreds of thousands of pixels, providing a route towards an implantable optogenetic visual prosthesis capable of generating vision by stimulating RGCs at near-cellular resolution.

neuroscience↗

bioRxiv2023

Multiscale imaging and quantitative analysis of plasma membrane protein-cortical actin interplay

The spatiotemporal organization of cell surface receptors is important for cell signaling. Cortical actin (CA), the subset of the actin cytoskeleton subjacent to the plasma membrane (PM), plays a large role in cell surface receptor organization. This was however shown largely through actin perturbation experiments, which raise concerns of nonspecific effects and preclude quantification of actin architecture and dynamics under unperturbed conditions. These limitations make it challenging to predict how changes in CA properties can affect receptor organization. To derive direct relationships between the architecture and dynamics of CA and the spatiotemporal organization of PM proteins, including cell surface receptors, we developed a multiscale imaging and computational analysis framework based on the integration of single-molecule imaging (SMI) of PM proteins and fluorescent speckle microscopy (FSM) of CA (combined: SMI-FSM) in the same live cell. SMI-FSM revealed differential relationships between PM proteins and CA based on the PM proteins actin binding ability, diffusion type and local CA density. It also highlighted the complexity of cell wide actin perturbation, where we found that global changes in actin properties caused by perturbation were not necessarily reflected in the CA properties near PM proteins, and the changes in PM protein properties upon perturbation varied based on the local CA environment. Given the widespread use of SMI as a method to study the spatiotemporal organization of PM proteins and the versatility of SMI-FSM, we expect it to be widely applicable to enable future investigation of the influence of CA architecture and dynamics on different PM proteins, especially in the context of actin-dependent cellular processes, such as cell migration. SignificancePlasma membrane protein organization, an important factor for shaping cellular behaviors, is influenced by cortical actin, the subset of the actin cytoskeleton near the plasma membrane. Yet it is challenging to directly and quantitatively probe this influence. Here, we developed an imaging and analysis approach that combines single-molecule imaging, fluorescent speckle microscopy and computational statistical analysis to characterize and correlate the spatiotemporal organization of plasma membrane proteins and cortical actin. Our approach revealed different relationships between different proteins and cortical actin, and highlighted the complexity of interpreting cell wide actin perturbation experiments. We expect this approach to be widely used to study the influence of cortical actin on different plasma membrane components, especially in actin-dependent processes.

biophysics↗