Protection from apoptosis by steel factor but not interleukin-3 is reversed through blockade of calcium influx.

Steel factor (SLF), the ligand for the c-Kit receptor, protects hemopoietic progenitors and mast cells from apoptosis. We show here that protection of 32D-Kit cells or mast cells from apoptosis by SLF is abrogated through concurrent inhibition of Ca2+ influx. In contrast, cell survival promoted by interleukin-3 is not affected by Ca2+ influx blockers. In the presence of blockers, increasing stimulation by SLF leads to greater levels of cell death in the population, indicating that it is the combination of activation by SLF with concurrent blockade of Ca2+ influx that results in apoptosis. The p815 mastocytoma, which expresses a mutated, constitutively active c-kit receptor, dies apoptotically in the presence of Ca2+ influx blockers alone. Ionomycin protects cells from SLF plus blocker-induced apoptosis, confirming specificity for Ca2+ ion blockade in cell death induction. Overexpression of bcl-2, which protects 32D-Kit cells from factor withdrawal, does not protect cells from apoptosis by SLF plus blocker. In contrast, caspase inhibitors YVAD-CHO, DEVD-FMK, and Boc-Asp-FMK protect cells from SLF plus blocker-induced death. These observations highlight the importance of SLF-stimulated Ca2+ influx in the protection of cells from apoptosis and demonstrate a new mechanism for inducing bcl-2 insensitive, caspase-dependent apoptosis through the combination of SLF stimulation with Ca2+ influx blockade.