A linear signal transduction pathway involving phosphatidylinositol 3-kinase, protein kinase Cepsilon, and MAPK in mesangial cells regulates interferon-gamma-induced STAT1alpha transcriptional activation.
Interferon-gamma (IFN-gamma) exerts an pleiotropic effect in mesangial cells in inflammatory glomerular diseases. The biologic effect of IFN-gamma is mediated by STAT1alpha. The precise mechanism by which IFN-gamma stimulates the transcriptional activity of STAT1alpha is poorly understood. I investigated the role of protein kinase C (PKC) epsilon in regulating the transcriptional activation of STAT1alpha in mesangial cells. IFN-gamma increased PKCepsilon activity in a time-dependent manner with a concomitant increase in STAT1alpha transcriptional activity. Expression of constitutively active PKCepsilon mimicked the effect of IFN-gamma on STAT1alpha-dependent transcription. Expression of dominant negative PKCepsilon inhibited IFN-gamma-induced STAT1alpha-dependent transcription. Ly294002, a pharmacological inhibitor of phosphatidylinositol (PI) 3-kinase, blocked IFN-gamma-induced PKCepsilon activity and resulted in inhibition of STAT1alpha transcriptional activity but had no effect on STAT1alpha tyrosine phosphorylation and STAT1alpha-DNA complex formation. A PKC inhibitor, H7, also had no effect on STAT1alpha tyrosine phosphorylation and DNA binding. However, Ly294002 and H7 blocked IFN-gamma-induced serine phosphorylation of STAT1alpha. These data indicate that PI 3 kinase-dependent PKCepsilon regulates STAT1alpha transcriptional activity in the absence of any effect on its DNA binding capability. In addition to activating PKCepsilon, IFN-gamma increased MAPK activity, resulting in transcriptional activation of Elk-1, a nuclear target of MAPK. Ly294002 or a dominant negative PI 3-kinase significantly blocked IFN-gamma-induced MAPK activity. On the other hand, ectopic expression of constitutively active PKCepsilon significantly increased MAPK activity. IFN-gamma-stimulated MAPK phosphorylated STAT1alpha in vitro. Inhibition of MAPK activity blocked IFN-gamma-induced serine phosphorylation of STAT1alpha; but its tyrosine phosphorylation and DNA binding were partially inhibited. Finally, expression of dominant negative MAPK significantly inhibited IFN-gamma-induced STAT1alpha-dependent transcription. These data provide the first evidence that IFN-gamma stimulates PKCepsilon in a PI 3-kinase-sensitive manner to activate MAPK, which regulates STAT1alpha transcriptional activity.
Choudhury
Goutam Ghosh
GG
Department of Medicine, University of Texas Health Science Center at San Antonio, 78229-3900, USA.
D000818:Animals; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D017930:Genes, Reporter / Q000235:genetics; D005920:Glomerular Mesangium / Q000166:cytology / Q000378:metabolism*; D050832:Interferon-Stimulated Gene Factor 3; D007371:Interferon-gamma / Q000235:genetics / Q000494:pharmacology*; D020935:MAP Kinase Signaling System; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009025:Morpholines / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D051744:Protein Kinase C-epsilon; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D012694:Serine / Q000378:metabolism; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation
D002867:Chromones; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D050832:Interferon-Stimulated Gene Factor 3; D007371:Interferon-gamma; D020928:Mitogen-Activated Protein Kinases; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011493:Protein Kinase C; D051744:Protein Kinase C-epsilon; D011994:Recombinant Proteins; D012694:Serine; D014157:Transcription Factors; C079868:gamma interferon activation factor; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C497257:Prkce protein, rat