Orai1 and STIM reconstitute store-operated calcium channel function

The two membrane proteins, STIM1 and Orail, have each been shown to be essential for the activation of store-operated channels ( SOC). Yet, how these proteins functionally interact is not known. Here, we reveal that STIM1 and Orail expressed together reconstitute functional SOCs. Expressed alone, Orail strongly reduces store-operated Ca2+ entry ( SOCE) in human embryonic kidney 293 cells and the Ca2+ release-activated Ca2+ current ( ICRAC) in rat basophilic leukemia cells. However, expressed along with the store-sensing STIM1 protein, Orail causes a massive increase in SOCE, enhancing the rate of Ca2+ entry by up to 103-fold. This entry is entirely store-dependent since the same coexpression causes no measurable store-independent Ca2+ entry. The entry is completely blocked by the SOC blocker, 2-aminoethoxydiphenylborate. Orail and STIM1 coexpression also caused a large gain in CRAC channel function in rat basophilic leukemia cells. The close STIM1 homologue, STIM2, inhibited SOCE when expressed alone but coexpressed with Orail caused substantial constitutive ( store-independent) Ca2+ entry. STIM proteins are known to mediate Ca2+ store-sensing and endoplasmic reticulum-plasma membrane coupling with no intrinsic channel properties. Our results revealing a powerful gain in SOC function dependent on the presence of both Orail and STIM1 strongly suggest that Orail contributes the PM channel component responsible for Ca2+ entry. The suppression of SOC function by Orail overexpression likely reflects a required stoichiometry between STIM1 and Orail.