A GPRC5B-Fyn signaling axis regulates VRAC activity by targeting LRRC8A.

The mechanisms by which physiological signals gate the volume-regulated anion channel (VRAC) remain poorly understood. Here, we identify a GPRC5B-Fyn signaling axis that directly controls VRAC activation by targeting the essential subunit LRRC8A. We found that the orphan G protein-coupled receptor GPRC5B binds LRRC8A and recruits the Src family kinase Fyn, which in turn suppresses hypotonicity-evoked VRAC activation. Disrupting Fyn binding to GPRC5B or substituting a predicted phosphorylation site on LRRC8A (Y362) abolishes this inhibition. Across primary astrocytes and reconstituted systems, GPRC5B prevents the intersubunit conformational rearrangements associated with VRAC opening without altering basal channel expression. In astrocytes, the MLC protein MLC1 partially sequesters GPRC5B, thereby reducing the GPRC5B-LRRC8A interaction, providing a mechanistic link to VRAC dysfunction in megalencephalic leukoencephalopathy. These results reveal a receptor-kinase-channel pathway that gates VRAC through Y362-dependent modulation of LRRC8A, establishing a molecular framework for VRAC regulation and its disruption in disease.