title: Overexpression of Bcl-2 or Bcl-xL inhibits Ara-C-induced CPP32/Yama protease activity and apoptosis of human acute myelogenous leukemia HL-60 cells.
abstract: 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 CPP32beta/Yama gene. Yama belongs to the interleukin 1beta 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-xL protein on Yama protease activation and PARP degradation during Ara-C-induced apoptosis. For this, we utilized HL-60/Bcl-2, HL-60/Bcl-xL, or control HL-60/neo cells, which were created by transfection of the cDNA of the bcl-2, bcl-xL, or the neomycin-resistant genes, respectively. As compared to HL-60/neo, HL-60/Bcl-2 and HL-60/Bcl-xL cells have 5-fold greater expression of Bcl-2 and Bcl-xL, respectively. However, these cell lines have similar levels of p32Yama and PARP. Treatment with 10 or 100 microM 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-xL cells, Ara-C-induced p32Yama activation by its cleavage, PARP degradation and apoptosis were significantly inhibited. High Bcl-2 and Bcl-xL 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-xL 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.
authors.affiliation: Department of Medicine, Winship Cancer Center, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
authors.forename: A M
authors.initials: AM
authors.lastname: Ibrado
authors.forename: Y
authors.initials: Y
authors.lastname: Huang
authors.forename: G
authors.initials: G
authors.lastname: Fang
authors.forename: L
authors.initials: L
authors.lastname: Liu
authors.forename: K
authors.initials: K
authors.lastname: Bhalla
mesh_terms: D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D053148:Caspase 3; D020169:Caspases; D003546:Cysteine Endopeptidases / Q000096:biosynthesis*; D003561:Cytarabine / Q000494:pharmacology*; D004790:Enzyme Induction / Q000187:drug effects; D005047:Etoposide / Q000494:pharmacology; D018922:HL-60 Cells / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D008942:Mitoxantrone / Q000494:pharmacology; D011065:Poly(ADP-ribose) Polymerases / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis; D051028:bcl-2-Associated X Protein
chemical_list: D000590:Amino Acid Chloromethyl Ketones; D000970:Antineoplastic Agents; C098738:N-acetyl-tyrosyl-valyl-alanyl-aspartyl chloromethyl ketone; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D051028:bcl-2-Associated X Protein; D003561:Cytarabine; D005047:Etoposide; D008942:Mitoxantrone; D011065:Poly(ADP-ribose) Polymerases; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D003546:Cysteine Endopeptidases