Distinct Orai-Coupling Domains in Stim1 and Stim2 Define the Orai-Activating Site

Abstract

The ER membrane-spanning STIM1 protein is a finely-tuned sensor of ER luminal Ca2+. Small changes in ER Ca2+ induce STIM1 to undergo an intricate self-triggering process, causing it to translocate into ER-PM junctions where it couples with and activates the highly Ca2+-selective family of Orai channels in the PM. The entering Ca2+ sustains Ca2+ oscillations, maintains Ca2+ homeostasis, and provides crucial long-term Ca2+ signals in many cell types which control gene expression and cellular growth. Similar in structure and also widely expressed among cells, the little-studied STIM2 protein is reported to differ subtly from STIM1 in its N-terminal domain, affecting luminal Ca2+-sensitivity and the rate of unfolding and self-activation. The STIM1 cytoplasmic C-terminus contains the STIM-Orai activating region (SOAR) which has been structurally resolved. While the corresponding SOAR sequence in STIM2 is highly conserved, we reveal it has a profoundly diminished interaction with and ability to gate Orai1 channels. We narrowed this distinction in Orai1 activation to a small sequence in SOAR, within which substitution of a single phenylalanine in STIM1 with leucine in STIM2 confers a severe decrease in Orai1 channel-gating efficacy. This residue is strategically positioned at the structural apex of the SOAR domain. Modification of this single residue within the intact STIM1 protein reveals its pivotal role in both interaction with and gating of the Orai1 channel. The results not only pinpoint a crucial locus of STIM-Orai coupling but also reveal a physiologically profound distinction between STIM1 and STIM2.