Abstract
Sindbis virus (SV) is an alpha virus used as a model for studying the role of apoptosis in virus infection. In this study, we examined the role of protein kinase C (PKC) in the apoptosis induced by SVNI, a virulent strain of SV. Infection of C6 cells with SVNI induced a selective translocation of PKCdelta to the endoplasmic reticulum and its tyrosine phosphorylation. The specific PKCdelta inhibitor rottlerin and a PKCdelta kinase-dead mutant increased the apoptosis induced by SVNI. To examine the role of the tyrosine phosphorylation of PKCdelta in the apoptosis induced by SVNI we used a PKCdelta mutant in which five tyrosine residues were mutated to phenylalanine (PKCdelta5). PKCdelta5-overexpressing cells exhibited increased apoptosis in response to SVNI as compared with control cells and to cells overexpressing PKCdelta. SVNI also increased the cleavage of caspase 3 in cells overexpressing PKCdelta5 but did not induce cleavage of PKCdelta or PKCdelta5. Using single tyrosine mutants, we identified tyrosines 52, 64, and 155 as the phosphorylation sites associated with the apoptosis induced by SVNI. We conclude that PKCdelta exerts an inhibitory effect on the apoptosis induced by SV and that phosphorylation of PKCdelta on specific tyrosines is required for this function.
Highlights
Sindbis virus (SV) is an alpha virus used as a model for studying the role of apoptosis in virus infection
We examined the role of protein kinase C (PKC) in the apoptosis induced by SVNI, a virulent strain of SV
To examine the role of the tyrosine phosphorylation of PKC␦ in the apoptosis induced by SVNI we used a PKC␦ mutant in which five tyrosine residues were mutated to phenylalanine (PKC␦5)
Summary
Vol 277, No 26, Issue of June 28, pp. 23693–23701, 2002 Printed in U.S.A. Infection of Glioma Cells with Sindbis Virus Induces Selective Activation and Tyrosine Phosphorylation of Protein Kinase C ␦. PKC␣, PKC⑀, and PKC have been associated with inhibition of cell apoptosis [22, 23], whereas PKC␦, , and have been described as proapoptotic kinases, and their cleavage by caspase 3 has been shown to be important for their action [24, 25] Apoptotic stimuli such as etoposide and ionizing radiation induce the cleavage of these isoforms and the accumulation of active catalytic fragments [25, 26]. We reported that etoposide induced apoptosis of C6 cells via activation of PKC␦ and its tyrosine phosphorylation on specific tyrosine residues [31]. We report that infection of C6 cells with SVNI, a neurovirulent strain of Sindbis virus, induced selective activation and tyrosine phosphorylation of PKC␦. Results emphasize that the action of PKC depends critically on its biological context
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