Myeloid maturation appears to require exit from the cell cycle and leads to activation of apoptosis in the differentiated cells. The level of Bcl-2, which is known to promote cell survival, is shown here to influence both these critical steps. Bcl-2 function during myelomonocytic differentiation was investigated by introducing a deregulated bcl-2 gene into HL60 promyelocytic leukemia cells, which can be induced to exit the cell cycle and differentiate into granulocytes or monocytes. Deregulated Bcl-2 expression did not itself promote differentiation but extended the lifespan of mature cells elicited by granulocytic or monocytic inducers. Unexpectedly, in response to induction, Bcl-2 overexpression markedly potentiated and hastened cell cycle withdrawal into G(0). Enhanced survival cannot account for the elevated numbers of G(0) cells, because they arose under induction conditions that did not kill control cells. Since the cell cycle status and growth of uninduced cells was not affected by Bcl-2-overexpression, its cell cycle inhibitory activity must require an induction signal. While cell cycle withdrawal may be necessary for maturation, it was not sufficient, implicating a requirement for specific differentiative signals. These results identify, for the first time, a function for the bcl-2 proto-oncogene that is separable from its enhancement of cell survival.