NH2-terminal BH4 domain of Bcl-2 is functional for heterodimerization with Bax and inhibition of apoptosis.
The Bcl-2 family proteins comprise pro-apoptotic as well as anti-apoptotic members. Heterodimerization between members of the Bcl-2 family proteins is a key event in the regulation of apoptosis. We report here that Bcl-2 protein was selectively cleaved by active caspase-3-like proteases in CTLL-2 cell apoptosis in response to interleukin-2 deprivation. Structural and functional analyses of the cleaved fragment revealed that the NH2-terminal region of Bcl-2 (1-34 amid acids) was required for its anti-apoptotic activity and heterodimerization with pro-apoptotic Bax protein. Site-directed mutagenesis of the NH2-terminal region showed that substitutions of hydrophobic residues of BH4 domain resulted in the loss of ability to form a heterodimer with Bax. Particularly instructive was that the V15E mutant of Bcl-2, which completely lost the ability to form a heterodimer with Bax, failed to inhibit Bax- and staurosporine-induced apoptosis. Our results suggest that the BH4 domain of Bcl-2 is critical for its heterodimerization with Bax and for exhibiting anti-apoptotic activity. Therefore, agents interferring with the critical residues of the BH4 domain may provide a new strategy in cancer therapy by impairing Bcl-2 function.
Hirotani
M
M
Institute of Molecular and Cellular Biosciences, University of Tokyo, 1-1-1, Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan.
Zhang
Y
Y
Fujita
N
N
Naito
M
M
Tsuruo
T
T
D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D019281:Dimerization; D004789:Enzyme Activation; D006801:Humans; D006868:Hydrolysis; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D051028:bcl-2-Associated X Protein
C494826:BAX protein, human; C494827:Bax protein, mouse; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D020169:Caspases