Ara-C has been shown to induce apoptosis of human acute myelogenous leukemia HL-60 cells. The DNA repair enzyme poly(ADP-ribose) polymerase (PARP) is known to be degraded during apoptosis. PARP as a substrate is cleaved by the Yama protease, encoded by the CPP32β/Yama gene. Yama belongs to the interleukin 1β converting enzyme/ced-3 family of cysteine proteases that are activated as a cascade, producing proteolytic cleavage of specific substrates that results in the morphological and biochemical features of apoptosis. In the present studies, we determined the effect of high intracellular levels of the antiapoptosis Bcl-2 or Bcl-x(L) protein on Yama protease activation and PARP degradation during Ara-C-induced apoptosis. For this, we utilized HL-60/Bcl-2, HL-60/Bcl-x(L), or control HL-60/neo cells, which were created by transfection of the cDNA of the bcl-2, bcl-x(L), or the neomycin-resistant genes, respectively. As compared to HL-60/neo, HL-60/Bcl- 2 and HL-60/Bcl-x(L) cells have 5-fold greater expression of Bcl-2 and Bcl- x(L), respectively. However, these cell lines have similar levels of p32Yama and PARP. Treatment with 10 or 100 μM Ara-C for 4 h produced DNA fragmentation and morphological features of apoptosis in HL-60/neo cells. This was associated with the cleavage and activation of p32Yama and PARP degradation but not with the induction of Yama mRNA. In contrast, in HL- 60/Bcl-2 and HL-60/Bcl-x(L) cells, Ara-C-induced p32Yama activation by its cleavage, PARP degradation and apoptosis were significantly inhibited. High Bcl-2 and Bcl-x(L) levels in these cells also inhibited Yama protease activity, PARP degradation, and apoptosis due to clinically relevant concentrations of etoposide and mitoxantrone. These results suggest that the activation of the Yama protease and PARP degradation are involved in Ara-C-, etoposide-, or mitoxantrone-induced apoptosis. In addition, they suggest that Bcl-2 and Bcl-x(L) antagonize drug-induced apoptosis by a mechanism that interferes in the activity of a key cysteine protease that is involved in the execution of apoptosis.
|Original language||English (US)|
|Number of pages||6|
|State||Published - Oct 15 1996|
ASJC Scopus subject areas
- Cancer Research