Regulation of the Ca2+‐sensing capacity of STIM1 by the G protein‐coupled estrogen receptor

Abstract

The stromal interaction molecule 1 (STIM1) is essential for activation of store‐operated Ca2+entry (SOCE). STIM1 contains in its intra‐endoplasmic reticular (ER) domain a canonical EF hand (cEF) that houses a Ca2+‐binding loop. This loop serves as a Ca2+sensor; Ca2+leaving its binding loop upon ER Ca2+depletion initiates conformational changes leading to STIM1 oligomerization and SOCE activation. We recently reported that activation of the G protein‐coupled estrogen receptor (GPER) clamps intracellular Ca2+signals via multifaceted inhibitory actions on both Ca2+efflux and SOCE; the latter results partly from alteration in phosphorylation at serines 575, 608 and 621 of STIM1. Here, we identified a dynamic interaction between GPER and STIM1 that directly controls the Ca2+‐sensing capacity of STIM1. GPER coimmunoprecipitates with STIM1 in resting primary endothelial cells, an association enhanced by treatment with either GPER agonist G‐1 or activation of SOCE using thapsigargin. This effect is not altered by combined S575/608/621A substitutions. Coimmunoprecipitation analysis using truncated versions of STIM1 showed that GPER associates with the cEF domain. Confocal microscopy identified a GPER's submembrane domain that localizes GPER to the ER. To assess the role of GPER‐STIM1 interaction in controlling STIM1's Ca2+sensing capacity, we generated a series of biosensors that incorporated STIM1's Ca2+‐binding loop in singlet, doublet or triplet. These biosensors faithfully report Ca2+binding to the loop, with tripled dynamic range by the triple‐loop biosensor and loss of Ca2+binding by charge substitutions targeting strategic elements for Ca2+binding. Using peptides corresponding to submembrane domains of GPER and these STIM1 biosensors, we identified a robust Ca2+‐independent interaction between a submembrane domain and the Ca2+‐binding loop. The charge substitutions that abolish Ca2+binding only reduce but do not negate the interaction, indicating that Ca2+and the GPER domain bind to different regions of the loop. Consistently, we have observed that GPER overexpression decreases SOCE and GPER knockdown substantially increases SOCE. These data reveal a new interaction between GPER and STIM1 that directly controls STIM1's Ca2+‐sensing capacity and SOCE activation.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.