摘要 : Caveolae are membrane domains having caveolin-1 (Cav1) as their main structural component. Here, we determined whether Cav1 affects Ca2+ signaling through the G alpha(q)-phospholipase-C beta (PLC beta) pathway using Fischer rat th... 展开 Caveolae are membrane domains having caveolin-1 (Cav1) as their main structural component. Here, we determined whether Cav1 affects Ca2+ signaling through the G alpha(q)-phospholipase-C beta (PLC beta) 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-G beta gamma and G alpha(q)-eYFP are similarly associated in both cell lines by Forster resonance energy transfer (FRET). Upon stimulation, the amount of FRET between G alpha(q)-eYFP and eCFP-G beta gamma remains high in FRTcav- cells, but decreases almost completely in FRTcav+ cells, suggesting that Cav1 is increasing the separation between G alpha q-G beta gamma subunits. In FRTcav(-) cells overexpressing PLC beta, 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 G alpha(q) and Cav1 that increase upon stimulation. Fluorescence correlation spectroscopy studies show that the mobility of G alpha(q)-eGFP is unaffected by activation in either cell type. The mobility of eGFP-G beta gamma remains slow in FRTcav(-) cells but increases in FRTcav(+) cells. Together, our data suggest that, upon stimulation, G alpha(q)(GTP) switches from having strong interactions with G beta gamma to Cav1, thereby releasing G beta gamma. This prolongs the recombination time for the heterotrimer, thus causing a sustained Ca2+ signal. 收起