Alternative splicing switches STIM1 targeting to specialized membrane contact sites and modifies SOCE

Alternative splicing is a potent modifier of protein function. Stromal interaction molecule 1 (Stim1) is the essential activator molecule of store-operated Ca2+ entry (SOCE) and a sorting regulator of certain ER proteins such as Stimulator of interferon genes (STING). Here, we characterize a conserved new variant, Stim1A, where splice-insertion translates into an additional C-terminal domain. We find prominent expression of exonA mRNA in testes, astrocytes, kidney and heart and confirm Stim1A protein in Western blot of testes. In situ, endogenous Stim1 with domain A, but not Stim1 without domain A localizes to unique adhesion junctions and to specialized membrane retrieval sites (tubulobulbar complexes) in testes. Functionally, using Ca2+ imaging and patch-clamp analysis, Stim1A shows a dominant-negative effect on SOCE and ICRAC, despite normal clustering and interaction with Orai1 investigated by combined TIRF and FRET analyses and as confirmed by an increased SOCE upon knock-down of endogenous Stim1A in astrocytes. Mutational analyses in conjunction with imaging and patch-clamp analyses of residues either in domain A or within the N-terminal region of Orai1 demonstrate a specific defect in stabilized channel gating. Our findings demonstrate that cell-type specific splicing of STIM1 adds both an intracellular targeting switch and adapts SOCE to meet the Ca2+ requirements of specific subcellular contact sites.