IFN-γ-Induced MHC Class II Expression: Transactivation of Class II Transactivator Promoter IV by IFN Regulatory Factor-1 is Regulated by Protein Kinase C-α

Abstract

Previous studies based on pharmacological evidence suggested a requirement for protein kinase C (PKC) activity in the regulation of IFN-γ-induced MHC class II (MHC-II) expression. In the present study, we investigated the molecular mechanisms by which PKC-α modulates IFN-γ-induced MHC-II expression in the mouse macrophage cell line RAW 264.7. Overexpression of a dominant-negative (DN) mutant of PKC-α inhibited the expression of IFN-γ-induced MHC-II but had no effect on IFN-γ-induced STAT1 nuclear translocation and DNA binding activity, as well as on the expression of inducible NO synthase, IFN consensus sequence binding protein, MHC class I, IFN regulatory factor (IRF)-1, and IFN-γ-inducible protein-10. Further analysis showed that IFN-γ-induced expression of the MHC class II transactivator (CIITA), a transcriptional coactivator essential for MHC-II expression, was inhibited in DN PKC-α-overexpressing cells. Studies with reporter constructs containing the promoter IV region of CIITA revealed that overexpression of a constitutively active mutant of PKC-α enhanced IRF-1, but not IRF-2, transcriptional activity. Furthermore, characterization of IRF-1 from both normal and DN PKC-α-overexpressing cells revealed differences in IRF-1 posttranslational modifications. Collectively, our data suggest a novel regulatory mechanism for IFN-γ-induced MHC-II expression, whereby PKC regulates CIITA expression by selectively modulating the transcriptional activity of IRF-1.