REGULATION OF T-CELL-RECEPTOR-STIMULATED BIVALENT-CATION ENTRY IN JURKAT E6 CELLS - ROLE OF PROTEIN-KINASE-C

Stimulation of Jurkat E6 cells with anti-CD3 antibody results in a cha racteristic rise in [Ca2+](i) which is due to both the release of Ca2 from intracellular stores and the entry of external Ca2+. Individual components of the [Ca2+](i) increase were investigated by measuring in tracellular Ca2+ release in the absence of external Ca2+ and determini ng influx of bivalent cations by following the entry of Mn2+. The incr ease in [Ca2+](i) induced by anti-CD3 antibody in the presence or abse nce of extracellular Ca2+ could be inhibited by the non-selective kina se inhibitor staurosporine, which also inhibits anti-CD3-stimulated ph ospholipase C activity. Stauosporine also inhibits the influx of bival ent cations induced by anti-CD3 antibody, but not that induced by depl etion of intracellular Ca2+ stores using thapsigargin. The effect of s taurosporine was compared with that of Ro 31-8425, a potent and select ive inhibitor of protein kinase C (PKC). Ro 31-8425, at concentrations up to 10 mu M, has no inhibitory effect on the anti-CD3 antibody-indu ced [Ca2+](i) increase or phospholipase C activity. These studies are consistent with the concept that augmentation of [Ca2+](i) by stimulat ed T-cell receptors requires activation of a kinase, probably a tyrosi ne kinase such as p56(lck), ZAP-70 or p59(fyn), and is independent of PKC. Phorbol esters inhibit the anti-CD3-stimulated [Ca2+](i) increase and phospholipase C activity, showing that this can be negatively reg ulated by PKC. A small potentiation of the anti-CD3 antibody-induced [ Ca2+](i) rise in the presence of extracellular Ca2+ was detected in th e presence of Ro 31-8425; this suggests that T-cell-receptor ligation can also limit the increase in [Ca2+](i) via PKC activation.