Journal of Biological Chemistry
Volume 284, Issue 44, 30 October 2009, Pages 30039-30048
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Mechanisms of Signal Transduction
Mcl-1 Degradation during Hepatocyte Lipoapoptosis*

https://doi.org/10.1074/jbc.M109.039545Get rights and content
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The mechanisms of free fatty acid-induced lipoapoptosis are incompletely understood. Here we demonstrate that Mcl-1, an anti-apoptotic member of the Bcl-2 family, was rapidly degraded in hepatocytes in response to palmitate and stearate by a proteasome-dependent pathway. Overexpression of a ubiquitin-resistant Mcl-1 mutant in Huh-7 cells attenuated palmitate-mediated Mcl-1 loss and lipoapoptosis; conversely, short hairpin RNA-targeted knockdown of Mcl-1 sensitized these cells to lipoapoptosis. Palmitate-induced Mcl-1 degradation was attenuated by the novel protein kinase C (PKC) inhibitor rottlerin. Of the two human novel PKC isozymes, PKCδ and PKCθ, only activation of PKCθ was observed by phospho-immunoblot analysis. As compared with Jurkat cells, a smaller PKCθ polypeptide and mRNA were expressed in hepatocytes consistent with an alternative splice variant. Short hairpin RNA-mediated knockdown of PKCθ reduced Mcl-1 degradation and lipoapoptosis. Likewise, genetic deletion of Pkcθ also attenuated Mcl-1 degradation and cytotoxicity by palmitate in primary hepatocytes. During treatment with palmitate, rottlerin inhibited phosphorylation of Mcl-1 at Ser159, a phosphorylation site previously implicated in Mcl-1 turnover. Consistent with these results, an Mcl-1 S159A mutant was resistant to degradation and improved cell survival during palmitate treatment. Collectively, these results implicate PKCθ-dependent destabilization of Mcl-1 as a mechanism contributing to hepatocyte lipoapoptosis.

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*

This work was supported, in whole or in part, by National Institutes of Health Grants R01 DK41876 (to G. J. G.), T32 07198 (to H. C. M.), K01 DK79875 (to J. L. M.), R01 CA69008 (to S. H. K.), and the optical microscopy core of P309K 84567. This work was also supported by the Mayo Foundation.

The on-line version of this article (available at http://www.jbc.org) contains a supplemental figure.