Matrix Rigidity‐Dependent Regulation of Ca(2+) at Plasma Membrane Microdomains by FAK Visualized by Fluorescence Resonance Energy Transfer

The dynamic regulation of signal transduction at plasma membrane microdomains remains poorly understood due to limitations in current experimental approaches. Genetically encoded biosensors based on fluorescent resonance energy transfer (FRET) can provide high spatiotemporal resolution for imaging cell signaling networks. Here, distinctive regulation of focal adhesion kinase (FAK) and Ca(2+) signals are visualized at different membrane microdomains by FRET using membrane‐targeting biosensors. It is shown that rigidity‐dependent FAK and Ca(2+) signals in human mesenchymal stem cells (hMSCs) are selectively activated at detergent‐resistant membrane (DRM or rafts) microdomains during the cell–matrix adhesion process, with minimal activities at non‐DRM domains. The rigidity‐dependent Ca(2+) signal at the DRM microdomains is downregulated by either FAK inhibition or lipid raft disruption, suggesting that FAK and lipid raft integrity mediate the in situ Ca(2+) activation. It is further revealed that transient receptor potential subfamily M7 (TRPM7) participates in the mobilization of Ca(2+) signals within DRM regions. Thus, the findings provide insights into the underlying mechanisms that regulate Ca(2+) and FAK signals in hMSCs under different mechanical microenvironments.