Cytoprotective peptide humanin binds and inhibits proapoptotic Bcl-2/Bax family protein BimEL.
Humanin (HN) is a recently identified endogenous peptide that protects cells against cytotoxicity induced by various stimuli. Recently, we showed that HN binds to and inhibits Bax, a proapoptotic Bcl-2 family protein, suggesting a mechanism for HN action. In this study, we identified Bim, a Bcl-2 homology 3-only member of the Bcl-2/Bax family, as an additional HN target protein. Using in vitro protein binding, immunoprecipitation, and coimmunolocalization assays, we demonstrated that HN binds directly to the extra long isoform of Bim (BimEL) but not the long (BimL) or short (BimS) isoforms. HN also protects cells against apoptosis induced by BimEL but not BimL and BimS in gene transfection studies. In contrast, mutants of HN which failed to bind BimEL failed to protect from BimEL-induced cell death. Moreover, HN inhibited BimEL-induced release of SMAC and cytochrome c from mitochondria isolated from bax-/-cells, indicating that HN can suppress BimEL independently of its effect on Bax. Finally, we demonstrate that HN prevents BimEL-induced oligomerization of Bak using isolated mitochondria. Taken together, our results indicate that the inhibition of BimEL may contribute to the antiapoptotic properties of the HN peptide.
Luciano
Frederic
F
The Burnham Institute, La Jolla, California 92037, USA.
Zhai
Dayong
D
Zhu
Xiuwen
X
Bailly-Maitre
Beatrice
B
Ricci
Jean-Ehrland
JE
Satterthwait
Arnold C
AC
Reed
John C
JC
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D051017:Apoptosis Regulatory Proteins; D000072224:Bcl-2-Like Protein 11; D019556:COS Cells; D002352:Carrier Proteins / Q000378:metabolism; D045304:Cytochromes c / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000378:metabolism; D010446:Peptide Fragments / Q000378:metabolism; D011506:Proteins / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051028:bcl-2-Associated X Protein
D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; C494772:BAK1 protein, human; C494826:BAX protein, human; C000606775:BCL2L11 protein, human; D000072224:Bcl-2-Like Protein 11; D002352:Carrier Proteins; C412403:DIABLO protein, human; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D024101:Mitochondrial Proteins; D010446:Peptide Fragments; D011506:Proteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051028:bcl-2-Associated X Protein; C427211:humanin; D045304:Cytochromes c