Different contribution of BH3-only proteins and caspases to doxorubicin-induced apoptosis in p53-deficient leukemia cells.
Bcl-2 family proteins are key regulators of the intrinsic apoptotic pathway, either facilitating (Bax, Bak, BH3-only) or inhibiting (Bcl-2, Bcl-x(L), Mcl-1, A1) mitochondrial release of apoptogenic factors. The role of caspases in this process is a matter of controversy. We have analyzed the relative contribution of caspases and Bcl-2 family of proteins in the induction phase of apoptosis triggered by doxorubicin in two p53-deficient leukemia cell lines, Jurkat and U937. First, we have found that caspases are dispensable for the induction phase of doxorubicin-induced apoptosis in both cell lines but they are needed to speed up the execution phase in Jurkat cells, not expressing Bax. Thus, down-regulation of Bak expression by siRNA significantly prevented doxorubicin-induced apoptosis in Jurkat but not in U937 cells. Reduction of Mcl-1 protein levels with siRNA increased sensitivity to apoptosis in both cell lines. Moreover, our results indicate that the contribution of BH3-only proteins to apoptosis is cell line specific. In Jurkat cells simultaneous silencing of Bim and PUMA was necessary to reduce doxorubicin-induced apoptosis. In U937 cells silencing of Bim or Noxa reduced sensitivity to doxorubicin. Immunoprecipitation experiments discarded an interaction between Mcl-1 and Bak in both cell lines and underscored the role of Bim and PUMA as mediators of Bax/Bak activation.
López-Royuela
Nuria
N
Departamento de Bioquimica, Biologia Molecular y Celular, Facultad de Ciencias, Universidad de Zaragoza, Pedro Cerbuna 12, 50009 Zaragoza, Spain.
Pérez-Galán
Patricia
P
Galán-Malo
Patricia
P
Yuste
Victor J
VJ
Anel
Alberto
A
Susín
Santos A
SA
Naval
Javier
J
Marzo
Isabel
I
D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D020169:Caspases / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D004249:DNA Damage; D004317:Doxorubicin / Q000494:pharmacology*; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D006801:Humans; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism*; D016159:Tumor Suppressor Protein p53 / Q000235:genetics*
D019253:Proto-Oncogene Proteins c-bcl-2; D016159:Tumor Suppressor Protein p53; D004317:Doxorubicin; D020169:Caspases