Previously we have reported that deregulated expression of c-myc in normal and leukemic myeloid cells blocked differentiation and, concomitantly, induced p53-independent apoptosis. Here, we show that this morbidity was due to premature recruitment of the Fas/CD95 cell death pathway which normally operates to induce apoptosis at the end of the terminal myeloid differentiation program. Analysis of the regulated components of this pathway revealed that IL6-mediated induction of differentiation resulted in rapid cell surface expression of CD95 receptor. Deregulated c-myc prevented the downregulation of CD95 ligand by maintaining its transcription, but caused premature downregulation of c-FLIP. First, the Type II (mitochondria-dependent, bcl-2-sensitive) and, then, the Type I (mitochondria-independent, bcl-2-insensitive) pathway were activated. Stable exogenous c-FLIP expression completely rescued the apoptotic phenotype. Furthermore, when the deregulated c-myc transgene was stably transduced into bone marrow cells from Fas(lpr/lpr) (CD95 receptor mutant) and FasL(gld/gld) (CD95 ligand mutant) mice, cell death was significantly suppressed relative to c-myc-transduced wild type bone marrow cells upon induction of differentiation. These data indicate that c-myc-mediated apoptosis associated with blocks in myeloid differentiation is dependent on the Fas/CD95 pathway. Our findings offer important new insights into understanding how deregulated c-myc alters normal blood cell homeostasis, and how additional mutations might promote leukemogenesis.