Stabilization of cortical actin induces internalization of transient receptor potential 3 (Trp3)-associated caveolar Ca2+ signaling complex and loss of Ca2+ influx without disruption of Trp3-inositol trisphosphate receptor association

Ca2+ influx via plasma membrane Trp3 channels is proposed to be regulated by a reversible interaction with inositol trisphosphate receptor (IP,R) in the endoplasmic reticulum. Condensation of the cortical actin layer has been suggested to physically disrupt this interaction and inhibit Trp3-mediated Ca2+ influx. This study examines the effect of cytoskeletal reorganization on the localization and function of Trp3 and key Ca2+ signaling proteins. Calyculin-A treatment resulted in formation of condensed actin layer at the plasma membrane; internalization of Trp3, G alpha (q/11), phospholipase C beta, and caveolin-1; and attenuation of 1-oleoyl-2-acetyl-sn-glycerol- and ATP-stimulated Sr2+ influx. Importantly, Trp3 and IP3R-3 remained co-localized inside the cell and were co-immunoprecipitated. Jasplakinolide also induced internalization of Trp3 and caveolin-1. Pretreatment of cells with cytochalasin D or staurosporine did not affect Trp3 but prevented calyculin-A-induced effects. Based on these data, we suggest that Trp3 is assembled in a caveolar Ca2+ signaling complex with IP3R, SERCA, G alpha (q/11), phospholipase C beta, caveolin-1, and ezrin. Furthermore, our data demonstrate that conditions which stabilize cortical actin induce loss of Trp3 activity due to internalization of the Trp3-signaling complex, not disruption of IP3R-Trp3 interaction. This suggests that localization of the Trp3-associated signaling complex, rather than Trp3-IP3R coupling, depends on the status of the actin cytoskeleton.