Caveolin-1 alters Ca²⁺ signal duration through specific interaction with the Gαq family of G proteins

Caveolae are membrane domains having caveolin-1 (Cav1) as their main structural component. Here, we determined whether Cav1 affects Ca²⁺ signaling through the Gαq-phospholipase-Cβ (PLCβ) pathway using Fischer rat thyroid cells that lack Cav1 (FRTcav⁻) and a sister line that forms caveolae-like domains due to stable transfection with Cav1 (FRTcav⁺). In the resting state, we found that eCFP-Gβγ and Gαq-eYFP are similarly associated in both cell lines by Forster resonance energy transfer (FRET). Upon stimulation, the amount of FRET between Gαq-eYFP and eCFP-Gβγ remains high in FRTcav⁻ cells, but decreases almost completely in FRTcav⁺ cells, suggesting that Cav1 is increasing the separation between Gαq-Gβγ subunits. In FRTcav⁻ cells overexpressing PLCβ, a rapid recovery of Ca²⁺ is observed after stimulation. However, FRTcav⁺ cells show a sustained level of elevated Ca²⁺. FRET and colocalization show specific interactions between Gαq and Cav1 that increase upon stimulation. Fluorescence correlation spectroscopy studies show that the mobility of Gαq-eGFP is unaffected by activation in either cell type. The mobility of eGFP-Gβγ remains slow in FRTcav⁻ cells but increases in FRTcav⁺ cells. Together, our data suggest that, upon stimulation, Gαq(GTP) switches from having strong interactions with Gβγ to Cav1, thereby releasing Gβγ. This prolongs the recombination time for the heterotrimer, thus causing a sustained Ca²⁺ signal.