Biology subjects

Kelkar, M.

Publications and source records attributed to Kelkar, M..

6 recordsLinked to original sources

bioRxiv2025

Control of cellular cortical tension and shape by RhoGTPase signalling

Shape changes are ubiquitous in biology, from cytokinesis at the single cell scale to tissue-scale morphogenesis involving coordinated changes in hundreds of cells. In all cases, morphogenesis is powered by gradients in mechanical tension that arise downstream of signalling. Many pathways converge on RhoGTPases that modulate the cytoskeleton and cell contractility to control cell mechanics and, subsequently, shape. Despite their physiological importance, we lack a quantitative understanding of how changes in signalling alter cortical mechanics to drive cell shape change. Here, we use optogenetics to quantitatively characterise the relationship between the amount of RhoGEF localised to the membrane, the downstream myosin recruitment, and the subsequent mechanical changes. We then show that cortical myosin amount and cortical tension increase linearly with the amount of membranous RhoGEF signalling. Based on these data, we develop a predictive model of the temporal evolution of RhoGEF membrane localisation, cortical myosin enrichment, and cortical tension in response to a pulse of light. Using this model together with an active surface model of the cell cortex, we show that the cellular shape changes induced by localised optogenetic recruitment of RhoGEF signalling can be predicted, directly linking gradients in signalling to shape change. Significance statementShape changes are ubiquitous in biology, during division in single cells and in tissue during embryogenesis. In all cases, shape change is powered by gradients in mechanical tension that arise downstream of changes in biochemical signals. Despite their importance, we lack a quantitative understanding of how changes in signals alter cell mechanics to drive cell shape change. Here, we control the location and amount of biochemical signal using light to quantitatively characterise the relationship between signals and their resulting biological and mechanical changes. We show that mechanical change scales linearly with the amount of biochemical signal. Based on this, we develop a mathematical model that predicts cell mechanical and shape changes from the location and amount of biochemical signals.

biophysics↗

bioRxiv2024

Spatial and temporal signatures of cell competition revealed by K-function analysis

It is generally hypothesised that in MDCK wild-type versus ScrKD co-cultures mechanical competition leads to the physical elimination of ScrKD cells, while in MDCK wild-type versus RasV12 co-cultures biochemical competition results in the non-apoptotic extrusion of RasV12 cells. However, the exact spatiotemporal signatures of competitive interactions in these systems remains largely unknown. Utilising advanced single-cell image analysis, this research quantifies the spatiotemporal dynamics of these competitive interactions. Time-lapse microscopy, combined with cellular segmentation, tracking, cell-state classification, and K-function clustering analysis provide detailed insights into how wild-type cell proliferation relates to the elimination of mutant cells. We find striking differences in the underlying processes of each competition type. In the ScrKD competition, elimination seems driven by diffuse proliferative forces unrelated to the immediate elimination site. In contrast, RasV12 cell elimination is closely linked to local increases in wild-type cell mitoses prior to the RasV12 extrusion. Both competitions trigger an increase in local wild-type cell proliferation post-elimination, although the timing of these responses vary. This study not only sheds light on the diverse mechanisms of cell competition but also underscores the complexity of cellular interactions in tissue dynamics, providing new perspectives on cellular quality control and a new quantitative approach to characterise these interactions.

biophysics↗

bioRxiv2023

Learning dynamic image representations for self-supervised cell cycle annotation

Time-based comparisons of single-cell trajectories are challenging due to their intrinsic heterogeneity, autonomous decisions, dynamic transitions and unequal lengths. In this paper, we present a self-supervised framework combining an image autoencoder with dynamic time series analysis of latent feature space to represent, compare and annotate cell cycle phases across singlecell trajectories. In our fully data-driven approach, we map similarities between heterogeneous cell tracks and generate statistical representations of single-cell trajectory phase durations, onset and transitions. This work is a first effort to transform a sequence of learned image representations from cell cycle-specific reporters into an unsupervised sequence annotation.

cell biology↗

bioRxiv2022

Force propagation between epithelial cells depends on active coupling and mechano-structural polarization

Cell-generated forces play a major role in coordinating the large-scale behavior of cell assemblies, in particular during development, wound healing and cancer. Mechanical signals propagate faster than biochemical signals, but can have similar effects, especially in epithelial tissues with strong cell-cell adhesion. However, a quantitative description of the transmission chain from force generation in a sender cell, force propagation across cell-cell boundaries, and the concomitant response of receiver cells is missing. For a quantitative analysis of this important situation, here we propose a minimal model system of two epithelial cells on an H-pattern ("cell doublet"). After optogenetically activating RhoA, a major regulator of cell contractility, in the sender cell, we measure the mechanical response of the receiver cell by traction force and monolayer stress microscopies. In general, we find that the receiver cells shows an active response so that the cell doublet forms a coherent unit. However, force propagation and response of the receiver cell also strongly depends on the mechano-structural polarization in the cell assembly, which is controlled by cell-matrix adhesion to the adhesive micropattern. We find that the response of the receiver cell is stronger when the mechano-structural polarization axis is oriented perpendicular to the direction of force propagation, reminiscent of the Poisson effect in passive materials. We finally show that the same effects are at work in small tissues. Our work demonstrates that cellular organization and active mechanical response of a tissue is key to maintain signal strength and leads to the emergence of elasticity, which means that signals are not dissipated like in a viscous system, but can propagate over large distances.

biophysics↗

bioRxiv2022

Spindle reorientation in response to mechanical stress is an emergent property of the spindle positioning mechanisms

Proper orientation of the mitotic spindle plays a crucial role in embryos, during tissue development, and in adults, where it functions to dissipate mechanical stress to maintain tissue integrity and homeostasis. While mitotic spindles have been shown to reorient in response to external mechanical stresses, the subcellular cues that mediate spindle reorientation remain unclear. Here, we have used a combination of optogenetics and computational modelling to better understand how mitotic spindles respond to inhomogeneous tension within the actomyosin cortex. Strikingly, we find that the optogenetic activation of RhoA only influences spindle orientation when it is induced at both poles of the cell. Under these conditions, the sudden local increase in cortical tension induced by RhoA activation reduces pulling forces exerted by cortical regulators on astral microtubules. This leads to a perturbation of the torque balance exerted on the spindle, which causes it to rotate. Thus, spindle rotation in response to mechanical stress is an emergent phenomenon arising from the interaction between the spindle positioning machinery and the cell cortex.

cell biology↗

bioRxiv2022

Tension at intercellular junctions is necessary for accurate orientation of cell division in the epithelium plane

The direction in which a cell divides is set by the orientation of its mitotic spindle and is important for determining cell fate, controlling tissue shape and maintaining tissue architecture. Division perpendicular to the plane of the substrate can promote tissue stratification during development or wound healing, but also metastasis when orientation is aberrant. Much is known about the molecular mechanisms involved in setting the spindle orientation. However, less is known about the contribution of mechanical factors, such as tissue tension, in ensuring spindle orientation in the plane of the epithelium, despite epithelia being continuously subjected to mechanical stresses. Here, we used suspended epithelial monolayers devoid of extracellular matrix and subjected to varying levels of tissue tension to study the orientation of division relative to the tissue plane. We found that decreasing tissue tension by compressing the monolayers or by inhibiting myosin contractility leads to a higher frequency of out-of-plane divisions. Reciprocally, accurate in-plane division can be restored by increasing tissue tension by increasing cell contractility or by tissue stretching. By considering the full three-dimensional geometry of the epithelium, we show that spindles are sensitive to tissue tension, independently of cell shape, through its impact on the tension at subcellular surfaces. Overall, our data suggest that accurate spindle orientation in the plane of the epithelium necessitates the presence of a sufficiently large tension at intercellular junctions. Significance statementIn growing epithelia, divisions are typically oriented in the plane of the tissue to drive expansion. In some organs, divisions are then re-oriented so that they occur perpendicular to the epithelium plane to drive tissue stratification and cell differentiation. When uncontrolled, this switch in orientation can lead to defects in tissue organisation and, in cancer, likely contribute to metastasis. While much is known about the molecular mechanisms controlling mitotic spindle orientation, less is known about the role of mechanical factors. Here we use mechanical and chemical perturbations to show that mechanics plays a role in controlling the plane of division. Overall, our data suggest that the orientation of spindles in the epithelium plane requires sufficient tension across intercellular junctions.

biophysics↗