Biology subjects

Suleiman, H. Y.

Publications and source records attributed to Suleiman, H. Y..

7 recordsLinked to original sources

bioRxiv2026

Progressive Actomyosin Remodeling Defining a Structured Podocyte Injury Response in Nephrotic Syndrome

Nephrotic syndrome, the leading cause of chronic kidney disease worldwide, is associated with injury and eventual loss of podocytes, the specialized epithelial cells that form the final layer of the kidney's filtration apparatus. The progression from injury to loss is largely irreversible. However, some forms of nephrotic syndrome undergo remission, suggesting that injured podocytes may retain a capacity for repair, the structural basis of which remains unknown. Podocytes respond to injury by the stereotypical morphological change known as foot process effacement (FPE), which can be visualized using transmission electron microscopy. Using modified ultrastructure expansion microscopy (U-ExM) protocol combined with super-resolution microscopy, we discovered that injured podocytes exhibit a three-stage progression of FPE maturation, with kinetics determined by the degree of actomyosin contractility. Through longitudinal study of adriamycin-induced nephropathy in mice, we identified three sequential stages of actomyosin reorganization within the sarcomere-like structures (SLSs) that define the injury response: focal infiltration of myosin IIA into synaptopodin-rich foot processes (Stage 1), disordered intermingling of synaptopodin/myosin IIA (Stage 2), and the formation of highly organized periodic contractile arrays oriented perpendicular to the direction of capillary blood flow (Stage 3). Longitudinal quantification revealed predictable temporal dynamics, with mature Stage 3 predominating by day 7-9 post-injury. To determine whether this sequence is conserved in human disease, we analyzed kidney biopsies from patients with minimal change disease, focal segmental glomerulosclerosis, membranous nephropathy, and diabetic nephropathy. The same three stages were present across all four diseases, and their distribution tracked disease course: acute podocytopathies exhibited heterogeneous early-stage patterns, whereas chronic injuries converged on uniform, mature SLS organization. These findings demonstrate that podocyte injury is a graded, potentially reversible process rather than a binary switch. They identify structured actomyosin remodeling as a conserved wound response across mechanistically distinct kidney diseases and suggest that therapeutic strategies targeting specific stages of this progression could alter the trajectory of diverse glomerular diseases.

cell biology↗

bioRxiv2026

Synaptopodin enables directional mechanoadaptation of integrin-based adhesions

The attachment of cells to their substrate through adhesion complexes is fundamental to tissue architecture and function. These adhesions are inherently optimized to resist shear forces parallel to the substrate, yet certain specialized cells must also withstand substantial perpendicular forces. How cells adapt their adhesion machinery to resist forces in different directions has remained unclear. In the kidney, podocytes experience perpendicular forces from pressurized filtrate flow while maintaining attachment to the glomerular basement membrane through integrin-based adhesions. Here we show that synaptopodin converts adhesions from shear-resistant to perpendicular force-resistant structures through coordinated reorganization of the actin cytoskeleton and adhesion complexes. Using an inertial force application system, we demonstrate that synaptopodin triggers force-dependent redistribution of {beta}1-integrin to the cell periphery specifically in response to perpendicular loading, while synaptopodin-deficient cells lack this directional adaptation and detach. This mechanism operates in multiple cell types and is physiologically essential: synaptopodin-null mice subjected to elevated glomerular pressure develop significant proteinuria and podocyte foot process effacement. These findings reveal a molecular basis for directional mechanoadaptation, whereby a single protein enables cells to reconfigure their adhesion architecture in response to the direction of applied force.

biophysics↗

bioRxiv2026

Orthogonal Force Balance Between Contractility and Shear Stress Governs Podocyte Dynamics

Maintenance of tissue barriers under mechanical stress represents a fundamental biological challenge across organ systems. In the kidney, podocyte cells withstand highly variable hemodynamic forces while preserving a tensioned, nanostructured filtration barrier. Dysregulation of this barrier leads to significant pathology, but the mechanical principles underlying homeostasis of cells against flow of filtrates have not yet been identified. Here, we uncover a counterintuitive mechanical homeostasis mechanism whereby podocyte attachment depends on a dynamic balance between external fluid shear stress and internal cellular contractility. Integrated biomechanical modeling and experiment reveal a previously unrecognized mechanosensing circuit that optimizes integrin distribution at foot process peripheries. Our mathematical framework for cell-matrix adhesion stability reveals, surprisingly, that reducing blood pressure can worsen outcomes when cell contractility is impaired, contrary to clinical belief that lowering blood pressure universally benefits cellular adhesion and kidney function. We validated this principle through mouse models with manipulated blood pressure and myosin inhibition, demonstrating that concurrent reduction of both shear stress and contractility worsens podocyte injury and proteinuria. Super-resolution microscopy confirms our predicted integrin redistribution patterns under these mechanical perturbations. These findings establish a fundamental mechanobiological principle applicable beyond nephrology, and suggest potential treatment pathways targeting non-equilibrium steady states.

cell biology↗

bioRxiv2025

WNK1 kinase activity is required for maintenance of podocyte structure

The filtration-function of glomeruli requires slit diaphragms formed by interdigitating podocyte foot processes, which are actin-based membrane protrusions. Dysregulation of mechanisms that maintain these membrane extensions lead to foot process effacement, proteinuria, and progression to chronic kidney disease. Building on our previous work that showed WNK1 kinase activity is necessary for the maintenance of normal biomechanical properties of glomeruli and podocyte foot process architecture, we tested the hypothesis that WNK1 kinase activity affects the structure of podocyte foot processes through modulation of actomyosin activity and focal adhesion complexes. Using a WNK1 kinase specific inhibitor, we determined by immunofluorescence microscopy of nascent focal adhesions, podocyte membrane spreading/extensions, and NMII paralog localization and extent of activation calculated from quantification of phosphorylated myosin, that all were sensitive to WNK1 kinase activity. Moreover, biochemical evidence of WNK1 kinase activity-dependent signalosomes supports a role for WNK1 in the maintenance of podocyte foot processes, and sarcomere-like structures (SLSs) that are induced in models of podocyte injury. Using primary and immortalized podocyte cell lines developed from control and Col4a3-/-Alport Syndrome model mice, we measured WNK1 kinase activity-dependent improvement in properties of injured podocytes in vitro. Physiological relevance of WNK1 kinase activity-dependent structural maintenance of podocyte foot processes was confirmed by significant acute proteinuria measured in response to WNK1 inhibition in vivo. Collectively, the results provide evidence that WNK1 kinase signalosome activity that includes formation of nascent focal adhesions and regulation of NMII localization and activity at membrane protrusions and extensions, are necessary for physiological maintenance of slit diaphragms. SignificanceTerminally differentiated podocytes are arborized cells with interdigitating foot processes that form the renal filtration barrier. Loss of foot process structural integrity causes progressive proteinuria, which can lead to irreversible renal injury, but the mechanisms that maintain foot process structure are incompletely understood. We report evidence that WNK1 kinase activity is required for maintenance of normal glomerular filtration in vivo, and this is mediated in part through WNK1 activity-dependent modulation of non-muscle myosin II activity, and formation of nascent focal adhesions that are necessary for lamellipodial extensions. Using glomeruli and podocyte cell lines developed from an Alport Syndrome podocyte injury model, we show that aspects of abnormal podocyte structure associated with chronic kidney disease can be suppressed through increased WNK1 activation.

cell biology↗

bioRxiv2024

Ultrastructure expansion microscopy (U-ExM) of mouse and human kidneys for analysis of subcellular structures

Ultrastructure expansion microscopy (U-ExM) involves the physical magnification of specimens embedded in hydrogels, which allows for super-resolution imaging of subcellular structures using a conventional diffraction-limited microscope. Methods for expansion microscopy exist for several organisms, organs, and cell types, and used to analyze cellular organelles and substructures in nanoscale resolution. Here, we describe a simple step-by-step U-ExM protocol for the expansion, immunostaining, imaging, and analysis of cytoskeletal and organellar structures in kidney tissue. We detail the critical modified steps to optimize isotropic kidney tissue expansion, and preservation of the renal cell structures of interest. We demonstrate the utility of the approach using several markers of renal cell types, centrioles, cilia, the extracellular matrix, and other cytoskeletal elements. Finally, we show that the approach works well on mouse and human kidney samples that were preserved using different fixation and storage conditions. Overall, this protocol provides a simple and cost-effective approach to analyze both pre-clinical and clinical renal samples in high detail, using conventional lab supplies and standard widefield or confocal microscopy.

cell biology↗

bioRxiv2021

An ex vivo culture model of kidney podocyte injury reveals mechanosensitive, synaptopodin-templating, sarcomere-like structures.

Chronic kidney diseases are widespread and incurable. The biophysical mechanisms underlying them are unclear, in part because material systems for reconstituting the microenvironment of the relevant kidney cells are limited. A critical question is how kidney podocytes (glomerular epithelial cells) regenerate the foot processes of the filtration apparatus following injury. Recently identified sarcomere-like structures (SLSs) with periodically spaced myosin IIA (a contractile protein) and synaptopodin (an actin-associated protein) appear in injured podocytes in vivo. We hypothesized that SLSs template synaptopodin in the initial stages of recovery, and tested this hypothesis by developing an ex vivo culture system that models both kidney physiology and pathophysiology. SLSs were observed in vitro for the first time as podocytes migrated out of harvested kidney glomeruli onto micropatterns of physiologically relevant proteins. SLSs emerged over two days, and cells formed foot process-like extensions from these periodically spaced proteins. SLS distributions and morphology were sensitive to actomyosin inhibitors, substrate stiffness, and extracellular matrix proteins associated with pathology. These results indicate a role for mechanobiological factors in podocyte recovery from injury, and suggest SLSs as a target for therapeutic intervention.

cell biology↗

bioRxiv2021

3D Visualization of the Podocyte Actin Network using Integrated Membrane Extraction, Electron Microscopy, and Deep Learning

Although actin stress fibers are abundant in cultured cells, little is known about these structures in vivo. In podocytes of the kidney glomerulus, much evidence suggests that mechanobiological mechanisms underlie injury, with changes to actin stress fiber structures potentially responsible for pathological changes to cell morphology. However, this hypothesis is difficult to rigorously test in vivo due to challenges with visualization. We therefore developed the first visualization technique capable of resolving the three-dimensional (3D) podocyte actin network with unprecedented detail in healthy and injured podocytes, and applied this technique to reveal the changes in the actin network that occur upon podocyte injury. Using isolated glomeruli from healthy mice as well as from three different mouse injury models (Cd2ap-/-, Lamb2-/- and the Col4a3-/- model of Alport syndrome), we applied our novel imaging technique that integrates membrane-extraction, focused ion bean scanning electron microscopy (FIB-SEM), and deep learning image segmentation. In healthy glomeruli, we observed actin cables that link the interdigitating podocyte foot processes to newly described actin structures located at the periphery of the cell body. The actin cables within the foot processes formed a continuous, mesh-like, electron dense sheet that incorporated the slit diaphragms required for kidney filtration. After injury, the actin network was markedly different, having lost its organization and presenting instead as a disorganized assemblage of actin condensates juxtaposed to the glomerular basement membrane. The new visualization method enabled us, for the first time, to observe the detailed 3D organization of actin networks in both healthy and injured podocytes. Shared features of actin condensations across all three injury models further suggested common mechanobiological pathways that govern changes to podocyte morphology after injury.

cell biology↗