Protein kinase C-ε confers resistance to TRAIL-induced apoptosis by altering levels of pro- and anti-apoptotic Bcl-2 family proteins

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Tumor necrosis factor-Related Apoptosis-Inducing Ligand (TRAIL) is a member of the tumor necrosis factor (TNF) family that promotes apoptosis primarily in cancer cells, without any detectable toxicity to normal cells. Protein kinase Cs (PKC) are a family of phospholipid-dependent serine/threonine kinases that play an important role in cell survival and apoptosis. Our lab has previously shown that PKCε is an anti-apoptotic protein that inhibits TNF-α-induced apoptosis in MCF-7 breast cancer cells. In the present study we investigated the mechanism by which PKCε influences the extrinsic cell death pathway. MCF-7 cells overexpressing PKCε (MCF-7/PKCε) were refractory to TRAIL-induced apoptosis compared to vector-transfected MCF-7 cells (MCF-7/Neo), as evident from the activation of caspases and the cleavage of poly(ADP-ribose) polymerase (PARP) in TRAIL-treated cells. In addition, depletion of PKCε by siRNA or inhibition of PKCε activity by overexpression of a dominant-negative mutant enhanced cellular sensitivity to TRAIL. Although TRAIL induces cell death through the extrinsic pathway, it also cooperates with the intrinsic/mitochondrial pathway to elicit cell death. Overexpression of PKCε attenuated the release of mitochondrial cytochrome c induced by TRAIL. Since Bcl-2 family proteins regulate the intrinsic cell death pathway, we compared the pattern of expression of Bcl-2 family members in MCF-7/Neo and MCF-7/PKCε cells. Overexpression of PKCε in MCF-7 cells resulted in an increase in anti-apoptotic Bcl-2 and a decrease in pro-apoptotic BID compared to MCF-7/Neo cells. Depletion of BID using siRNA conferred resistance to MCF-7/Neo cells from TRAIL-induced cell death. Furthermore, depletion of Bcl-2 from MCF-7/PKCε cells by siRNA dramatically increased their susceptibility to TRAIL-induced apoptosis. Thus, our results demonstrate that a decrease in the ratio of pro- to anti-apoptotic Bcl-2 family members was associated with the anti-apoptotic function of PKCε in TRAIL-induced apoptosis. (Supported by a grant CA71727 from the NCI)