Caveolae Modulate the Activity of LRRC8-Mediated VRAC by the Structural Membrane Protein Caveolin-1

Abstract

The volume-regulated anion channel (VRAC) plays a critical role in cell volume regulation and other fundamental physiological processes. However, the mechanism of how VRAC is activated and modulated has not been completely clarified. Caveolin-1 (Cav-1), as an important ion channel binding protein, forms complexes with channel proteins and exchangers to regulate channel activity and function. The purpose of this study was to explore the importance and value of Cav-1 in cardiac VRAC activation and regulation. In the study, we proved that the membrane protein LRRC8A was detected in the same caveolae-enriched fractions, as the same as Caveolin-1 in ventricular myocytes. The intracellular Cl⁻ concentration increased and the cell volume decreased dramatically after caveolae being destroyed in cardiomyocytes. Moreover, we found that I_Cl,vol decreased not only in LRRC8A silencing cardiomyocytes but also in Cav-1 silencing cardiomyocytes, which indicated that caveolin-1 may affect the function of VRAC. Then we further explore the physical relationship between LRRC8A and Cav-1 in cell membrane. We observed that the fluorescence label of LRRC8A was overlapping with Cav-1 in the cell plasma membrane and caveolin-1 co-immunoprecipitated with LRRC8A, which demonstrated that Cav-1 is the basis of VRAC channel activation by acting on LRRC8A. The whole study provides further evidence of the relevance of Cav-1 on the activation and modulation of endothelial LRRC8A-mediated VRAC.