Activation mechanism for CRAC current and store- operated Ca2+ entry: calcium influx factor and iPLA2β-dependent pathway*

Here we tested the role of calcium influx factor (CIF) and calcium-independent phospholipase A2 (iPLA2) in activation of CRAC (Ca release-activated Ca) channels and storeoperated Ca entry (SOCE) in rat basophilic leukemia (RBL-2H3) cells. We demonstrate that: 1) endogenous CIF production may be triggered by Ca release (net loss), as well as by simple buffering of free Ca within the stores; 2) specific 82 kDa variant of iPLA2β and corresponding activity are present in membrane fraction of RBL cells; 3) exogenous CIF (extracted from other species) mimics the effects of endogenous one, and activates iPLA2β when applied to cell homogenates, but not intact cells; 4) activation of ICRAC can be triggered in resting RBL cells by their dialysis with exogenous CIF; 5) molecular or functional inhibition of iPLA2β prevents activation of ICRAC, which could be rescued by cell dialysis with a human recombinant iPLA2β; 6) dependence of ICRAC on intracellular pH strictly follows pH dependence of iPLA2β activity; 7) (S)-BEL, a chiral enantiomer of suicidal substrate specific for iPLA2β could be effectively used for pharmacological inhibition of ICRAC and storeoperated Ca entry. These findings validate and significantly advance our understanding of the CIF-iPLA2–dependent mechanism of activation of ICRAC and SOCE.