Modulation of lipopolysaccharide-induced NF-IL6 activation by protein kinase C-alpha in a mouse macrophage cell line.
We have previously shown that overexpression of a dominant-negative (DN) mutant of protein kinase C-alpha (PKC-alpha) in RAW 264.7 macrophages inhibited lipopolysaccharide (LPS)-induced IL-1alpha, inducible nitric oxide synthase and cyclooxygenase-2 expression. This inhibition was not related to defective NF-kappaB nuclear translocation, suggesting that PKC-alpha might be involved in the modulation of other LPS-inducible transcription factors. In the present study, we have investigated the impact of PKC-alpha on the activation of AP-1 and NF-IL6 in LPS-treated RAW 264.7 macrophages. Electrophoretic mobility shift assays and luciferase reporter constructs revealed that LPS-induced AP-1 transcriptional activity was normal in DN PKC-alpha-overexpressing RAW 264.7 cells. In contrast, LPS-induced DNA-binding and transcriptional activities of NF-IL6 were inhibited in DN PKC-alpha-overexpressing RAW 264.7 cells and correlated with an impairment of NF-IL6 nuclear translocation. Conversely, overexpression of either wild-type PKC-alpha or a constitutively active PKC-alpha mutant significantly enhanced LPS-stimulated NF-IL6-dependent promoter activity. Finally, LPS-induced expression of two genes regulated by NF-IL6, namely IL-1beta and granulocyte colony-stimulating factor, was impaired in DN PKC-alpha-overexpressing RAW 264.7 cells. Taken together, these results suggest that regulation of NF-IL6 activity constitutes one of the mechanisms by which PKC-alpha modulates LPS-induced gene expression in the mouse macrophage cell line RAW 264.7.
D021581:Active Transport, Cell Nucleus; D000818:Animals; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000378:metabolism*; D051743:CCAAT-Enhancer-Binding Protein-delta; D022762:CCAAT-Enhancer-Binding Proteins / Q000378:metabolism; D002460:Cell Line; D005786:Gene Expression Regulation; D016179:Granulocyte Colony-Stimulating Factor / Q000235:genetics; D007375:Interleukin-1 / Q000235:genetics; D007527:Isoenzymes / Q000502:physiology*; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008264:Macrophages / Q000378:metabolism*; D051379:Mice; D011493:Protein Kinase C / Q000502:physiology*; D051571:Protein Kinase C-alpha; D012333:RNA, Messenger / Q000032:analysis; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014157:Transcription Factors
D022782:CCAAT-Enhancer-Binding Protein-beta; D022762:CCAAT-Enhancer-Binding Proteins; C497253:Cebpd protein, mouse; D007375:Interleukin-1; D007527:Isoenzymes; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D018808:Transcription Factor AP-1; D014157:Transcription Factors; D051743:CCAAT-Enhancer-Binding Protein-delta; D016179:Granulocyte Colony-Stimulating Factor; C496630:Prkca protein, mouse; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha