Survival-factor-induced phosphorylation of bad results in its dissociation from Bcl-xL but not Bcl-2

I. Hirai, H. G. Wang

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Abstract

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 proapoptotic 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.

Original languageEnglish (US)
Pages (from-to)345-352
Number of pages8
JournalBiochemical Journal
Volume359
Issue number2
DOIs
Publication statusPublished - Oct 15 2001

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All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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