Biology subjects

Karmakar, R.

Publications and source records attributed to Karmakar, R..

5 recordsLinked to original sources

bioRxiv2025

Normalized Raman Imaging for Studies of Tissue Physiology of the Kidney

Conventional histological relies on fixation, embedding, sectioning, and staining methods that distort cellular architecture, extract lipids, and introduces variability, limiting reproducibility. We present Normalized Stimulated Raman Imaging (NoRI) that enables quantitative, label-free protein and lipid measurements at high-spatial resolution. NoRI computationally corrects protein, lipid, and water Raman signals, allowing quantitative biomass measurements while preserving tissue architecture, facilitating analysis by convolutional neural networks. Applied to mouse kidney, NoRI accurately classified tubule types (F1-[harmonic mean of precision and recall against manual annotation]=0.93), anatomical regions (F1=0.91), and biological sex (F1=0.97), revealing greater cytoplasmic lipid (+6.9mg/mL; p=0.028) and nuclear protein (+26.3mg/mL; p<0.001) in female tubules. In acute kidney injury, NoRI captured dynamic lipid and protein organization and quantified brush border remodelling and lipid droplet changes over 25 days. Spatial lipid quantification was central for feature classification in AKI (F1=1.0). These results establish NoRI as a reproducible, quantitative method for diagnostics, histopathology, and feature discovery.

physiology↗

bioRxiv2025

Kidney Tissue Characterization using Normalized Raman Imaging and Segment-Anything

Normalized Raman Imaging (NoRI) enables high-resolution, label-free quantification of protein and lipid concentrations in biological tissues. Because NoRI provides rich molecular information, the analysis of its large, multi-channel datasets turns into a significant computational bottleneck. In this work, we introduce a novel, modular computational pipeline for automated segmentation and quantification of kidney tissue structures imaged with NoRI. The pipeline integrates classical image processing with state-of-the-art machine learning tools, including the Segment Anything Model (SAM) and ilastik, to segment key anatomical and biochemical features--such as tubules, nuclei, brush borders, and lumens. A custom contrast-enhancement strategy was developed to create a third SAM input channel from NoRI data, leading to a substantial improvement in segmentation performance (F1 score: 0.9226). Our framework enables accurate cytoplasm, resolved quantification of protein and lipid concentrations and reveals distinct biochemical signatures across renal tubule subtypes and experimental conditions. This method offers a robust, scalable foundation for quantitative tissue analysis and enhances the utility of NoRI imaging for biomedical research.

bioinformatics↗

bioRxiv2024

Membrane binding properties of the cytoskeletal protein bactofilin

Bactofilins are a widespread family of cytoskeletal proteins with important roles in bacterial morpho-genesis, chromosome organization and motility. They polymerize in a nucleotide-independent manner, forming non-polar filaments that are typically associated with the cytoplasmic membrane. Membrane binding was suggested to be mediated by a short N-terminal peptide, but the underlying mechanism and the conservation of this interaction determinant among bacteria remain unclear. Here, we use the bacto-filin homolog BacA of the stalked bacterium Caulobacter crescentus as a model to analyze the membrane-binding behavior of bactofilins. Based on site-directed mutagenesis of the N-terminal region, we identify the full membrane-targeting sequence of BacA (MFSKQAKS) and pinpoint amino acid residues that are critical for its function in vivo and in vitro. Molecular dynamics simulations then provide detailed insight into the molecular mechanism underlying the membrane affinity of this peptide. Collectively, these analyses reveal a delicate interplay between the water exclusion of hydrophobic N-terminal residues, the arrangement of the peptide within the membrane and the electrostatic attraction between positively charged groups in the peptide and negative charges in the phospholipid molecules. A comprehensive bio-informatic analysis shows that the composition and properties of the membrane-targeting sequence of BacA are conserved in numerous bactofilin homologs from diverse bacterial phyla. Importantly, our findings reveal cooperative effects between the membrane-binding and polymerization activities of BacA. Moreover, they demonstrate that both of these activities critically contribute to the recruitment of the BacA client protein PbpC, a membrane-bound cell wall synthase that uses a conserved peptide in its N-terminal cytoplasmic tail to interact with BacA assemblies. Finally, we show that PbpC can functionally replace the endogenous membrane-targeting sequence of BacA when provided at elevated levels in trans, indicating that client proteins can make a significant contribution to the membrane association of bacto-filin polymers. Together, these results unravel the mechanistic underpinnings of membrane binding by bactofilin homologs, thereby illuminating a previously obscure but important aspect in the biology of this cytoskeletal protein family.

microbiology↗

bioRxiv2023

Eukaryotic chemotaxis under periodic stimulation shows temporal gradient dependence

When cells of the social amoeba Dictyostelium discoideum are starved of nutrients they start to synthesize and secrete the chemical messenger and chemoattractant cyclic Adenosine Mono Phosphate (cAMP). This signal is relayed by other cells, resulting in the establishment of periodic waves. The cells aggregate through chemotaxis towards the center of these waves. We investigated the chemotactic response of individual cells to repeated exposure to waves of cAMP generated by a microfluidic device. When the period of the waves is short, the chemotactic ability of the cells was found to increase upon exposure to more waves, suggesting the development of a longer-term memory. This effect was not significant for longer wave periods. We show that the experimental results are consistent with a model that includes a slowly rising and decaying component that is activated by the temporal gradient of cAMP concentration. The observed enhancement in chemotaxis is relevant to populations in the wild: once sustained, periodic waves of the chemoattractant are established, it is beneficial to cells to improve their chemotactic ability in order to reach the aggregation center sooner.

biophysics↗

bioRxiv2021

BoolSim, a Graphical Interface for Open Access Boolean Network Simulations and Use in Guard Cell CO2 Signaling

Signaling networks are at the heart of almost all biological processes. Most of these networks contain a large number of components and often the connections between these components are either not known, or the rate equations that govern the dynamics of soluble signaling components are not quantified. This uncertainty in network topology and parameters can make it challenging to formulate detailed mathematical models. Boolean networks, in which all components are either on or off, have emerged as viable alternatives to more detailed mathematical models but can be difficult to implement. Therefore, open source format of such models for community use is desirable. Here we present BoolSim, a freely available graphical user interface (GUI) that allows users to easily construct and analyze Boolean networks. BoolSim can be applied to any Boolean network. We demonstrate BoolSims application using a previously published network for abscisic acid-driven stomatal closure in Arabidopsis. We also show how BoolSim can be used to generate testable predictions by extending the network to include CO2 regulation of stomatal movements. Predictions of the model were experimentally tested and the model was iteratively modified based on experiments showing that ABA closes stomata even at near zero CO2 concentrations (1.5 ppm CO2). One Sentence SummaryThis study presents an open-source, graphical interface for the simulation of Boolean networks and applies it to an abscisic acid signaling network in guard cells, extended to include input from CO2.

plant biology↗