Protein kinase Calpha is involved in interferon regulatory factor 3 activation and type I interferon-beta synthesis.

Protein kinase C (PKC) isoforms are critically involved in the regulation of innate immune responses. Herein, we investigated the role of conventional PKCalpha in the regulation of IFN-beta gene expression mediated by the Toll-like receptor 3 (TLR3) signaling pathway. Inhibition of conventional PKC (cPKC) activity in monocyte-derived dendritic cells or ...
TLR3-expressing cells by an isoform-specific inhibitor, Goe6976, selectively inhibited IFN-beta synthesis induced by double-stranded RNA polyinosine-polycytidylic acid. Furthermore, reporter gene assays confirmed that PKCalpha regulates IFN-beta promoter activity, since overexpression of dominant negative PKCalpha but not PKCbeta(I) repressed interferon regulatory factor 3 (IRF-3)-dependent but not NF-kappaB-mediated promoter activity upon TLR3 engagement in HEK 293 cells. Dominant negative PKCalpha inhibited IRF-3 transcriptional activity mediated by overexpression of TIR domain-containing adapter inducing IFN-beta and Tank-binding kinase-1. Additional biochemical analysis demonstrated that Goe6976-treated dendritic cells exhibited IRF-3 phosphorylation, dimerization, nuclear translocation, and DNA binding activity analogous to their control counterparts in response to polyinosine-polycytidylic acid. In contrast, co-immunoprecipitation experiments revealed that TLR3-induced cPKC activity is essential for mediating the interaction of IRF-3 but not p65/RelA with the co-activator CREB-binding protein. Furthermore, PKCalpha knock-down with specific small interfering RNA inhibited IFN-beta expression and down-regulated IRF-3-dependent promoter activity, establishing PKCalpha as a component of TLR3 signaling that regulates IFN-beta gene expression by targeting IRF-3-CREB-binding protein interaction. Finally, we analyzed the involvement of cPKCs in other signaling pathways leading to IFN-beta synthesis. These experiments revealed that cPKCs play a role in the synthesis of IFN-beta induced via both TLR-dependent and -independent pathways.
Mesh Terms:
Active Transport, Cell Nucleus, Carbazoles, Cell Line, Cell Nucleus, Down-Regulation, Enzyme Inhibitors, Humans, Indoles, Interferon Regulatory Factor-3, Interferon-beta, NF-kappa B, Phosphorylation, Poly I-C, Promoter Regions, Genetic, Protein Kinase C, Protein Kinase C-alpha, Protein-Serine-Threonine Kinases, RNA, Double-Stranded, RNA, Small Interfering, Signal Transduction, Toll-Like Receptor 3, Transcription Factor RelA, Transcription, Genetic
J. Biol. Chem.
Date: May. 18, 2007
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