A TRPV4-dependent calcium signaling axis regulates lamellipodial actin architecture to promote cell migration.

Cell migration is crucial for development and tissue homeostasis, while its dysregulation leads to severe pathologies. Cell migration is driven by the extension of actin-based lamellipodial protrusions powered by actin polymerization, which is tightly regulated by signaling pathways, including Rho GTPases and calcium (Ca(2+)) signaling. While the importance of Ca(2+) signaling in lamellipodial protrusions has been established, the molecular mechanisms linking Ca(2+) to lamellipodia assembly are unknown. Here, we identify a novel Ca(2+) signaling axis involving the mechano-gated channel transient receptor potential vanilloid 4 (TRPV4), which regulates lamellipodial protrusions in various cell types. Using Ca(2+) and Förster resonance energy transfer (FRET) imaging, we demonstrate that TRPV4-mediated Ca(2+) influx upregulates RhoA activity within lamellipodia, which then facilitates formin-mediated actin assembly. Mechanistically, we identify Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) and tumor endothelial marker 4 (TEM4) as key mediators relaying the TRPV4-mediated Ca(2+) signal to RhoA. These data define a molecular pathway by which Ca(2+) influx regulates small GTPase activity within a specific cellular domain-lamellipodia-and demonstrate its critical role in organizing the actin machinery and promoting cell migration in diverse biological contexts.