Role of C/EBP-β, p38 MAPK, and MKK6 in IL-1β-mediated C3 gene regulation in astrocytes

J Cell Biochem. 2011 Apr;112(4):1168-75. doi: 10.1002/jcb.23032.

Abstract

Complement component C3, the central player in the complement cascade and the pro-inflammatory cytokine IL-1β is expressed by activated glial cells and may contribute to neurodegeneration. This study examines the regulation of the expression of C3 by IL-1β in astroglial cells focusing on the role of the upstream kinase MKK6, p38-α MAPK, and C/EBP-β isoforms (LAP1, LAP2, or LIP) in astroglial cells. Activation of human astroglial cell line, U373 with IL-1β, led to the induction of C3 mRNA and protein expression as determined by real-time RT-PCR and Western blot analysis, respectively. This induction was suppressed by the pharmacological inhibitor of p38 MAPK (i.e., SB202190-HCl), suggesting the involvement of p38 MAPK in C3 gene expression. IL-1β also induced C3 promoter activity in U373 cells in a MAP kinase- and C/EBP-β-dependent manner. Cotransfection of C3 luciferase reporter construct with constitutively active form of the upstream kinase in the MAP kinase cascade, that is, MKK6 (the immediate upstream activator of p38 kinase) resulted in marked stimulation of the promoter activity, whereas overexpression of a dominant negative forms of MKK6 and p38α MAPK inhibited C3 promoter activity. Furthermore, a mutant form of C/EBP-β, LAP(T235A) showed reduction in IL-1β-mediated C3 promoter activation. These results suggest that the p38α, MAPK, and MKK6 play prominent roles in IL-1β and C/EBP-β-mediated C3 gene expression in astrocytes.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Astrocytes / drug effects*
  • Astrocytes / metabolism
  • Astrocytes / pathology
  • Blotting, Western
  • CCAAT-Enhancer-Binding Protein-beta / genetics
  • CCAAT-Enhancer-Binding Protein-beta / metabolism*
  • Cell Line, Tumor
  • Complement C3 / genetics
  • Complement C3 / metabolism*
  • Enzyme Inhibitors / pharmacology
  • Gene Expression Regulation / drug effects
  • Humans
  • Imidazoles / pharmacology
  • Interleukin-1beta / pharmacology*
  • Luciferases / genetics
  • Luciferases / metabolism
  • MAP Kinase Kinase 6 / genetics
  • MAP Kinase Kinase 6 / metabolism*
  • Mutation
  • Phosphorylation / drug effects
  • Promoter Regions, Genetic / genetics
  • Protein Isoforms / genetics
  • Protein Isoforms / metabolism
  • Pyridines / pharmacology
  • Reverse Transcriptase Polymerase Chain Reaction
  • p38 Mitogen-Activated Protein Kinases / antagonists & inhibitors
  • p38 Mitogen-Activated Protein Kinases / genetics
  • p38 Mitogen-Activated Protein Kinases / metabolism*

Substances

  • CCAAT-Enhancer-Binding Protein-beta
  • Complement C3
  • Enzyme Inhibitors
  • Imidazoles
  • Interleukin-1beta
  • Protein Isoforms
  • Pyridines
  • Luciferases
  • p38 Mitogen-Activated Protein Kinases
  • MAP Kinase Kinase 6
  • MAP2K6 protein, human
  • 4-(4-fluorophenyl)-2-(4-hydroxyphenyl)-5-(4-pyridyl)imidazole