DEPLETION-ACTIVATED CALCIUM CURRENT IS INHIBITED BY PROTEIN-KINASE INRBL-2H3 CELLS

Whole cell patch-clamp recordings and single-cell Ca2+ measurements we re used to study the control of Ca2+ entry through the Ca2+ release-ac tivated Ca2+ influx pathway (I-CRAC) in rat basophilic leukemia cells, When intracellular inositol 1,4,5-trisphosphate (InsP(3))-sensitive s tores were depleted by dialyzing cells with high concentrations of Ins P(3), I-CRAC inactivated only slightly in the absence of ATP, Inclusio n of ATP accelerated inactivation 2-fold, The inactivation was increas ed further by the ATP analogue adenosine 5'-[gamma-thio]triphosphate, which is readily used by protein kinases, but not by 5'-adenylyl imido diphosphate, another ATP analogue that is not used by kinases. Neither cyclic nucleotides nor inhibition of calmodulin or tyrosine kinase pr evented the inactivation. Staurosporine and bisindolylmaleimide, prote in kinase C inhibitors, reduced inactivation of I-CRAC, whereas phorbo l ester accelerated inactivation of the current. These results demonst rate that a protein kinase-mediated phosphorylation, probably through protein kinase C, inactivates I-CRAC. Activation of the adenosine rece ptor (A(3) type) in RBL cells did not evoke much Ca2+ influx or system atic activation of I-CRAC. After protein kinase C was blocked, however , large I-CRAC was observed in all cells and this was accompanied by l arge Ca2+ influx, The ability of a receptor to evoke Ca2+ entry is det ermined, at least in part, by protein kinase C. Antigen stimulation, w hich triggers secretion through a process that requires Ca2+ influx, a ctivated I-CRAC The regulation of I-CRAC by protein kinase will theref ore have important consequences on cell functioning.