Abstract
Previous studies based on pharmacological evidence suggested a requirement for protein kinase C (PKC) activity in the regulation of IFN-gamma-induced MHC class II (MHC-II) expression. In the present study, we investigated the molecular mechanisms by which PKC-alpha modulates IFN-gamma-induced MHC-II expression in the mouse macrophage cell line RAW 264.7. Overexpression of a dominant-negative (DN) mutant of PKC-alpha inhibited the expression of IFN-gamma-induced MHC-II but had no effect on IFN-gamma-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-gamma-inducible protein-10. Further analysis showed that IFN-gamma-induced expression of the MHC class II transactivator (CIITA), a transcriptional coactivator essential for MHC-II expression, was inhibited in DN PKC-alpha-overexpressing cells. Studies with reporter constructs containing the promoter IV region of CIITA revealed that overexpression of a constitutively active mutant of PKC-alpha enhanced IRF-1, but not IRF-2, transcriptional activity. Furthermore, characterization of IRF-1 from both normal and DN PKC-alpha-overexpressing cells revealed differences in IRF-1 posttranslational modifications. Collectively, our data suggest a novel regulatory mechanism for IFN-gamma-induced MHC-II expression, whereby PKC regulates CIITA expression by selectively modulating the transcriptional activity of IRF-1.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Cell Line
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DNA-Binding Proteins / biosynthesis
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / metabolism
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DNA-Binding Proteins / physiology*
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Histocompatibility Antigens Class II / biosynthesis*
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Histocompatibility Antigens Class II / genetics
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Humans
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Interferon Regulatory Factor-1
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Interferon Regulatory Factor-2
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Interferon-gamma / physiology*
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Isoenzymes / physiology
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Mice
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Nuclear Proteins*
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Phosphoproteins / biosynthesis
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Phosphoproteins / genetics
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Phosphoproteins / metabolism
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Phosphoproteins / physiology*
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Promoter Regions, Genetic / immunology*
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Protein Kinase C / biosynthesis
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Protein Kinase C / genetics
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Protein Kinase C / physiology*
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Protein Kinase C-alpha
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Protein Processing, Post-Translational / genetics
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Protein Processing, Post-Translational / immunology
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RNA, Messenger / antagonists & inhibitors
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RNA, Messenger / metabolism
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Repressor Proteins*
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Trans-Activators / genetics*
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Trans-Activators / metabolism*
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Transcription Factors*
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Transcriptional Activation / immunology*
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Transfection
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Up-Regulation / genetics
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Up-Regulation / immunology
Substances
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DNA-Binding Proteins
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Histocompatibility Antigens Class II
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IRF1 protein, human
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IRF2 protein, human
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Interferon Regulatory Factor-1
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Interferon Regulatory Factor-2
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Irf1 protein, mouse
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Irf2 protein, mouse
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Isoenzymes
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MHC class II transactivator protein
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Nuclear Proteins
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Phosphoproteins
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RNA, Messenger
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Repressor Proteins
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Trans-Activators
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Transcription Factors
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Interferon-gamma
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PRKCA protein, human
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Prkca protein, mouse
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Protein Kinase C
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Protein Kinase C-alpha