Calcium inhibition and calcium potentiation of Orai1, Orai2, and Orai3 calcium release-activated calcium channels

Abstract

The recent discoveries of Stim1 and Orai proteins have shed light on the molecular makeup of both the endoplasmic reticulum Ca2+ sensor and the calcium release-activated calcium (CRAC) channel, respectively. In this study, we investigated the regulation of CRAC channel function by extracellular Ca2+ for channels composed primarily of Orai1, Orai2, and Orai3, by coexpressing these proteins together with Stim1, as well as the endogenous channels in HEK293 cells. As reported previously, Orai1 or Orai2 resulted in a substantial increase in CRAC current (Icrac), but Orai3 failed to produce any detectable Ca2+-selective currents. However, sodium currents measured in the Orai3-expressing HEK293 cells were significantly larger in current density than Stim1-expressing cells. Moreover, upon switching to divalent free external solutions, Orai3 currents were considerably more stable than Orai1 or Orai2, indicating that Orai3 channels undergo a lesser degree of depotentiation. Additionally, the difference between depotentiation from Ca 2+ and Ba2+ or Mg2+ solutions was significantly less for Orai3 than for Orai1 or -2. Nonetheless, the Na+ currents through Orai1, Orai2, and Orai3, as well as the endogenous store-operated Na+ currents in HEK293 cells, were all inhibited by extracellular Ca2+ with a half-maximal concentration of ∼20 μM. We conclude that Orai1, -2, and -3 channels are similarly inhibited by extracellular Ca 2+, indicating similar affinities for Ca2+ within the selectivity filter. Orai3 channels appeared to differ from Orai1 and -2 in being somewhat resistant to the process of Ca2+ depotentiation.