The pro-apoptotic Bcl-2-family protein Bad heterodimerizes with Bcl-2 and Bcl-xL in the outer mitochondrial membranes, nullifying their anti-apoptotic activities and promoting cell death. We report that interleukin-3 (IL-3) stimulation induces Bad phosphorylation and triggers its translocation from mitochondria to cytoplasm in cells expressing Bcl-xL but not Bcl-2. Overexpression of Bad sensitized Bcl-xL-expressing FL5.12 cells to apoptosis induced by IL-3 deprivation, but had no effect on the viability of cells expressing Bcl-2. IL-3 stimulation induced Bad phosphorylation at Ser-112, impairing its binding to Bcl-xL and resulting in its association with 14-3-3 proteins in the cytosol. However, Ser-112 phosphorylation could not trigger Bad dissociation from mitochondria in FL5.12 cells expressing Bcl-2. In 293T cells expressing Bcl-xL, Bad was phosphorylated at three serines, 112, 136 and 155, and was largely localized in the cytosolic fraction. In contrast, overexpression of Bcl-2 prevented phosphorylation of Bad at Ser-136 and Ser-155, sequestering this protein in the mitochondrial membranes. When the N-terminal regions of Bcl-2 and Bcl-xL were swapped with each other, the Bcl-xL(N)–Bcl-2 chimaeric protein (containing the N-terminal region of Bcl-xL) failed to prevent Bad phosphorylation in cells and was unable to block the cytosolic distribution of this pro-apoptotic protein. Additional experiments with the Bcl-2(N)–Bcl-xL chimaeric protein (containing the N-terminal region of Bcl-2) indicated that, although the N-terminal region of Bcl-2 is necessary, it is not sufficient for sequestering Bad in the mitochondrial membranes. These observations suggest that growth-factor-mediated phosphorylation of Bad contributes to the cytoprotective function of Bcl-xL but not Bcl-2.
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October 2001
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Research Article|
October 08 2001
Survival-factor-induced phosphorylation of Bad results in its dissociation from Bcl-xL but not Bcl-2
Itaru HIRAI;
Itaru HIRAI
Drug Discovery Program, H. Lee Moffitt Cancer Center and Research Institute, Departments of Interdisciplinary Oncology and Pharmacology & Therapeutics, University of South Florida College of Medicine, 12902 Magnolia Drive, Tampa, FL 33612, U.S.A.
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Hong-Gang WANG
Hong-Gang WANG
1
Drug Discovery Program, H. Lee Moffitt Cancer Center and Research Institute, Departments of Interdisciplinary Oncology and Pharmacology & Therapeutics, University of South Florida College of Medicine, 12902 Magnolia Drive, Tampa, FL 33612, U.S.A.
1To whom correspondence should be addressed (e-mail wanghg@moffitt.usf.edu).
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Publisher: Portland Press Ltd
Received:
April 30 2001
Revision Received:
June 13 2001
Accepted:
August 10 2001
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London ©2001
2001
Biochem J (2001) 359 (2): 345–352.
Article history
Received:
April 30 2001
Revision Received:
June 13 2001
Accepted:
August 10 2001
Citation
Itaru HIRAI, Hong-Gang WANG; Survival-factor-induced phosphorylation of Bad results in its dissociation from Bcl-xL but not Bcl-2. Biochem J 15 October 2001; 359 (2): 345–352. doi: https://doi.org/10.1042/bj3590345
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