Three intracellular signals for cytotoxic T lymphocyte-mediated killing. Independent roles for protein kinase C, Ca2+ influx, and Ca2+ release from internal stores.

Ligation of the TCR after interaction of a CTL with a relevant target cell results in a change in CTL shape with reorientation and secretion of cytoplasmic granules. Secretion can be induced by treating CTL with the combination of a phorbol ester to activate protein kinase C and ionomycin to increase the intracellular Ca2+ concentration by ion-selective permeabilization of both the plasma membrane and internal Ca2+ storage depots. Treatment of cloned murine CTL with PMA induced a change in cell shape without movement of cytoplasmic granules, whereas increased intracellular [Ca2+] after treatment with ionomycin resulted in granule movement without shape change. Granule movement induced by ionomycin did not lead to secretion in the absence of PMA. Direct activation of the TCR resulted in increased intracellular Ca2+ predominately from influx of extracellular Ca2+ with a substantially smaller contribution from release of Ca2+ from internal stores. By independently inhibiting either component of the TCR-initiated increase in intracellular Ca2+, it was determined that both sources of Ca2+ were required for granule movement and secretion. Thus, increases in intracellular Ca2+ concentration mediated by these two sources appear not to be functionally equivalent. Taken together, these results indicate that the CTL functional response that normally occurs after interaction with an antigenically relevant target cell is regulated by three independent signals.