The Sigma1 Receptor Competes with STIM1 to Bind Orai1 to Regulate Store Operated Calcium Entry (SOCE)

Abstract

The Sigma1 receptor is an ER chaperone protein targeted to mitochondrial associated ER membrane regions (MAMs). An increase in its expression has been observed in a variety of cancer cells and it exerts an anti-apoptotic action, promoting cell proliferation and inhibiting the apoptotic response to tumor suppressor genes such as p53 and Bax. Although its mechanism of action remains unclear, there is evidence to suggest that Sigma1R regulates inter-organelle Ca2+ signaling thereby controlling cellular bioenergetics, and is also a regulator of plasma membrane ion channels. We have investigated its role in the regulation of store operated Ca2+ entry (SOCE) triggered by ER Ca2+ store depletion. Increased expression of Sigma1R in HEK293 cells profoundly inhibited SOCE, whereas knockdown of endogenous Sigma1R in CHO cells increased SOCE. An agonist at Sigma1R, SKF10047, further inhibited SOCE whereas the antagonists, BD1047, relieved this inhibition.Co-immunoprecipitation experiments showed a novel interaction between Sigma1R and STIM1 which is ligand dependent, SKF10047 promoted the interaction and BD1047 reduced it. Total internal reflection fluorescence (TIRF) microscopy showed the recruitment of Sigma1R to ER-plasma membrane junctions was STIM1 dependent, whereas elevated Sigma1R expression slowed STIM1 recruitment. Plasma membrane biotinylation experiments also showed that STIM1 promoted Sigma1R recruitment to a surface complex while Sigma1R reduced the levels of STIM1 in the complex. Increased expression of STIM1 but not Orai1 partially rescued the inhibition of SOCE in cells over-expressing Sigma1R. Imaging of purified plasma membrane Orai1 channels complexes by atomic force microscopy (AFM) showed hexameric Orai1 decorated by both Sigma1R and STIM1. Overall our results suggest that Sigma1R inhibits SOCE in two ways; first, by binding STIM1 and reducing its recruitment to surface Orai1 channels and second, by directly modulating the STIM1-Orai1 complex.