SUMMARY Shape is an indicator of cell health. But how is the information in shape decoded? We hypothesize that decoding occurs by modulation of signaling through changes in plasma membrane curvature. Using analytical approaches and numerical simulations, we studied how elongation of cell shape affects plasma membrane signaling. Mathematical analyses reveal transient accumulation of activated receptors at regions of higher curvature with increasing cell eccentricity. This distribution of activated receptors is periodic, following the Mathieu function, and it arises from local imbalance between reaction and diffusion of soluble ligands and receptors in the plane of the membrane. Numerical simulations show that transient microdomains of activated receptors amplify signals to downstream protein kinases. For growth factor receptor pathways, increasing cell eccentricity elevates the levels of activated cytoplasmic Src and nuclear MAPK1,2. These predictions were experimentally validated by changing cellular eccentricity, showing that shape is a locus of retrievable information storage in cells.
Less than half of patients suffering from major depressive disorder, a leading cause of disability worldwide, achieve remission with current antidepressants, making it imperative to develop more effective treatment. A new therapeutic direction is emerging from the increased understanding of natural resilience as an active stress-coping process. It is known that potassium (K+) channels in the ventral tegmental area (VTA) are an active mediator of resilience. However, no druggable targets have been identified to potentiate active resilience mechanisms. In the chronic social defeat stress model of depression, we report that KCNQ-type K+ channel openers, including FDA-approved drug retigabine (ezogabine), show antidepressant efficacy. We demonstrate that overexpression of KCNQ channels in the VTA dopaminergic neurons and either local infusion or systemic administration of retigabine normalized neuronal hyperactivity and depressive behaviours. These findings identify KCNQ as a target for conceptually novel antidepressants that function through the potentiation of active resilience mechanisms.
Nephrotoxicity is a critical adverse event that leads to discontinuation of kinase inhibitor (KI) treatment. Here we show, through meta-analyses of FDA Adverse Event Reporting System, that dasatinib is associated with high risk for glomerular toxicity that is uncoupled from hypertension, suggesting a direct link between dasatinib and podocytes. We further investigate the cellular effects of dasatinib and other comparable KIs with varying risks of nephrotoxicity. Dasatinib treated podocytes show significant changes in focal adhesions, actin cytoskeleton, and morphology that are not observed with other KIs. We use phosphoproteomics and kinome profiling to identify the molecular mechanisms of dasatinib-induced injury to the actin cytoskeleton, and atomic force microscopy to quantify impairment to cellular biomechanics. Furthermore, chronic administration of dasatinib in mice causes reversible glomerular dysfunction, loss of stress fibers, and foot process effacement. We conclude that dasatinib induces nephrotoxicity through altered podocyte actin cytoskeleton, leading to injurious cellular biomechanics.
Caveolae are membrane domains that may influence cell signaling by sequestering specific proteins such as G-protein-coupled receptors (GPCRs). While previous reports largely show that Gαq subunits, but not other G-proteins, interact strongly with the caveolae protein, Caveolin-1 (Cav1), the inclusion of GPCRs in caveolae is controversial. Here, we have used fluorescence methods to determine the effect of caveolae on the physical and functional properties of two GPCRs that have been reported to reside in caveolae, bradykinin receptor type 2 (B2R), which is coupled to Gαq, and the μ-opioid receptor (μOR), which is coupled to Gαi. While caveolae do not affect cAMP signals mediated by μOR, they prolong Ca2+ signals mediated by B2R. In A10 cells that endogenously express B2R and Cav1, downregulation of Cav1 ablates the prolonged recovery seen upon bradykinin stimulation in accord with the idea that the presence of caveolae prolongs Gαq activation. Immunofluorescence and Förster resonance energy transfer (FRET) studies show that a significant fraction of B2R resides at or close to caveolae domains while none or very little μOR resides in caveolae domains. The level of FRET between B2R and caveolae is reduced by downregulation of Gαq or by addition of a peptide that interferes with Gαq–Caveolin-1 interactions, suggesting that Gαq promotes localization of B2R to caveolae domains. Our results lead to the suggestion that Gαq can localize its associated receptors to caveolae domains to enhance their signals.
The shape of a cell within tissues can represent the history of chemical and physical signals that it encounters, but can information from cell shape regulate cellular phenotype independently? Using optimal control theory to constrain reaction-diffusion schemes that are dependent on different surface-to-volume relationships, we find that information from cell shape can be resolved from mechanical signals. We used microfabricated 3-D biomimetic chips to validate predictions that shape-sensing occurs in a tension-independent manner through integrin β3 signaling pathway in human kidney podocytes and smooth muscle cells. Differential proteomics and functional ablation assays indicate that integrin β3 is critical in transduction of shape signals through ezrin–radixin–moesin (ERM) family. We used experimentally determined diffusion coefficients and experimentally validated simulations to show that shape sensing is an emergent cellular property enabled by multiple molecular characteristics of integrin β3. We conclude that 3-D cell shape information, transduced through tension-independent mechanisms, can regulate phenotype.
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