Bcr-Abl-Mediated Protection from Apoptosis Downstream of Mitochondrial Cytochrome c Release

Abstract

Bcr-Abl, activated in chronic myelogenous leukemias, is a potent cell death inhibitor. Previous reports have shown that Bcr-Abl prevents apoptosis through inhibition of mitochondrial cytochrome c release. We report here that Bcr-Abl also inhibits caspase activation after the release of cytochrome c. Bcr-Abl inhibited caspase activation by cytochrome c added to cell-free lysates and prevented apoptosis when cytochrome c was microinjected into intact cells. Bcr-Abl acted posttranslationally to prevent the cytochrome c-induced binding of Apaf-1 to procaspase 9. Although Bcr-Abl prevented interaction of endogenous Apaf-1 with the recombinant prodomain of caspase 9, it did not affect the association of endogenous caspase 9 with the isolated Apaf-1 caspase recruitment domain (CARD) or Apaf-1 lacking WD-40 repeats. These data suggest that Apaf-1 recruitment of caspase 9 is faulty in the presence of Bcr-Abl and that cytochrome c/dATP-induced exposure of the Apaf-1 CARD is likely defective. These data provide a novel locus of Bcr-Abl antiapoptotic action and suggest a distinct mechanism of apoptosomal inhibition.

We thank Christopher Freel for aid in manuscript preparation and Jeffrey Rathmell for critical reading of the manuscript. Additionally, we thank Kevin Wright, Seth Margolis, and Jennifer Perry for helpful comments and discussion. We also thank Ann Marie Pendergast (Duke University) for the Bcr-Abl constructs and 32D cells, Yigong Shi (Princeton University) for the catalytically inactive caspase 3 construct, and Xiaodong Wang (University of Texas Southwestern) for the full-length Apaf-1 and procaspase 9 cDNAs.

This work was supported by NIH RO1 CA102702 to S.K. and NIH RO1 NS42197 to M.D. P.B.D. is a Ruth L. Kirschstein NRSA fellow. Z.T.S. is a predoctoral fellow of the Breast Cancer Research Program of the USARMC.