Fanconi anemia and apoptosis: The fancc gene product functions downstream of caspase 3

Hematopoietic progenitor cells (HPC) from mice and children with type C Fanconi anemia (FA-C) are hypersensitive to mitotic inhibitory factors, including interferon-γ (IFN-γ) and tumor necrosis factor-α (TNFα)(Blood, 90:974,1997). We tested the hypothesis that IFN/fasL induced programmed cell death in FA-C cells involves the ordered activation of specific caspase molecules. In immunoblot experiments, the FA-C lymphoblastoid cell line HSC536N treated with an agonistic fas antibody (100 ng/ml) and IFN-γ(1 ng/ml) contained activated caspase 3, 6, and 7 (but not 1, 2, 4, or 10) by 60 minutes and PARP cleavage products by 180 minutes. Caspase 3 activation by immunoblot and flow cytometric analysis was identical in FA-C cells and FA cells complemented by FANCC cDNA transfer even though the latter cells were less apoptotic. The apoptotic effects of fas agonists in IFN-γ-treated human FA-C CD34+ cells, FA-C murine progenitor cells, and EBV-transformed lymphoblasts from a child with FA of the C type, were blocked when the cells were pretreated any one of three inhibitors of caspase 3 protease. Caspase 1 inhibitors did not block the effect of IFN and fas ligand and caspase 1 inhibition did not prevent caspase 3 cleavage but caspase 8 inhibition did block caspase 3 activation. We conclude that in IFN-primed FA-C cells, fas-induced apoptosis involves the activation of caspase 8, which controls activation of caspase 3 family members but that the function of FANCC in suppressing normal apoptotic responses is downstream of caspase 3.