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

Whole cell patch-clamp recordings and single-cell Ca2+ measurements were used to study the control of Ca2+ entry through the Ca2+ release-activated Ca2+ influx pathway (I-CRAC) in rat basophilic leukemia cells, When intracellular inositol 1,4,5-trisphosphate (InsP(3))-sensitive stores were depleted by dialyzing cells with high concentrations of InsP(3), I-CRAC inactivated only slightly in the absence of ATP, Inclusion of ATP accelerated inactivation 2-fold, The inactivation was increased further by the ATP analogue adenosine 5'-[gamma-thio]triphosphate, which is readily used by protein kinases, but not by 5'-adenylyl imidodiphosphate, another ATP analogue that is not used by kinases. Neither cyclic nucleotides nor inhibition of calmodulin or tyrosine kinase prevented the inactivation. Staurosporine and bisindolylmaleimide, protein kinase C inhibitors, reduced inactivation of I-CRAC, whereas phorbol ester accelerated inactivation of the current. These results demonstrate that a protein kinase-mediated phosphorylation, probably through protein kinase C, inactivates I-CRAC. Activation of the adenosine receptor (A(3) type) in RBL cells did not evoke much Ca2+ influx or systematic activation of I-CRAC. After protein kinase C was blocked, however, large I-CRAC was observed in all cells and this was accompanied by large Ca2+ influx, The ability of a receptor to evoke Ca2+ entry is determined, at least in part, by protein kinase C. Antigen stimulation, which triggers secretion through a process that requires Ca2+ influx, activated I-CRAC The regulation of I-CRAC by protein kinase will therefore have important consequences on cell functioning.