Molecular Determinants of TRPC Channels Gating by STIM1

Abstract

We investigated if STIM1 SOAR, which has a coiled‐coil (CC) domain, interacts with the TRPC channel N‐ (NT) and/or C‐terminal (CT) CCs by determining how mutations in the TRPC CCs affect their interaction with SOAR and their gating by STIM1. We found that SOAR binds directly to TRPC1, TRPC4 and TRPC5, but not to TRPC3 and TRPC6. Mutations of 3 residues in the NT CCs of all TRPCs eliminate their surface expression and reduce binding of TRPC1 and TRPC5 to STIM1, while increasing binding of TRPC3 and TRPC6 to STIM1. In contrast, most single mutations in the NT CC of TRPC3 enhance binding to STIM1 with no appreciable effect on surface expression. Notably, I807S in the CT CC of TRPC3 has no effect on its interaction with STIM1, but eliminates the increased interaction observed by mutations of the NT CC. Accordingly, sensitivity of TRPC3 and TRPC3(L245S) to STIM1 knockdown by siRNA and scavenging STIM1 by the channel dead Orai1(R91W) show that a greater inhibition of STIM1 function is required to inhibit the function of TRPC3(L245S) compared to WT TRPC3. Lastly, TRPC3(I807S) eliminates regulation of TRPC3 and TRPC3(L245S) by STIM1. These findings suggest a model in which the TRPC3 NT and CT CCs interact to shield the CT CC from binding to STIM1. Dissociation of this interaction allows STIM1 SOAR to access and bind to the CT CC, to present the STIM1 polybasic domain to the conserved negative charges on TRPCs, and, ultimately, to open the channel.