Constituted by STIM1 and Orai1, calcium (Ca2+) release activated Ca2+ (CRAC) channels mediate store operated Ca2+ entry (SOCE). Aberrant CRAC signaling is closely associated with immune-deficiency, many types of cancers neurodegenerative diseases. Thus it is crucial to unveil molecular mechanisms underlying its regulation, and to develop new pharmacological tools against CRAC channels. At pre-translational level, we identified a new spliced variant of STIM1 named STIM1β that could more efficiently engage and gate Orai1, resulting in enhanced SOCE responses. At post-translational level, we identified an interacting protein of STIM1, which could decrease SOCE by promoting lysosomal degradation of STIM1. Mostly based on virtual docking, we also obtained a new SOCE inhibitor that are more specific for STIM1-Orai1. Together, our findings may serve as potential targets or tools for treating CRAC related diseases.