Regulation of JNK by MKK-7 in fibroblast-like synoviocytes

Arthritis Rheum. 2006 Jul;54(7):2127-35. doi: 10.1002/art.21919.


Objective: JNK regulates matrix metalloproteinase (MMP) gene expression and joint destruction in rheumatoid arthritis (RA). Previous studies demonstrated that the 2 upstream MAPK kinases (MKK-4 and MKK-7) are phosphorylated in RA synovium and form a complex with JNK in fibroblast-like synoviocytes (FLS). However, the functional hierarchy of MKK-4 and MKK-7 in FLS has not been determined. We determined the relative contributions of these MKKs by evaluating the effect of MKK-4 and MKK-7 gene knockdown in cultured FLS.

Methods: FLS were transfected with MKK-4 and/or MKK-7 small interfering RNA, and protein levels were determined by immunoblotting. After stimulation with interleukin-1/beta (IL-1beta), tumor necrosis factor alpha(TNFalpha, or anisomycin, kinase function was determined by in vitro kinase assay. Activator protein 1 (AP-1) binding and transcriptional activity were determined by electrophoretic mobility shift assay and AP-1-luciferase promoter assay, respectively. MMP-3 expression was determined by enzyme-linked immunosorbent assay and quantitative polymerase chain reaction.

Results: IL-1beta-induced JNK phosphorylation was dependent on MKK-7 but not on MKK-4; however, anisomycin-activated JNK required both kinases. In vitro kinase assay demonstrated that IL-1beta-or TNFalpha induced JNK activity was only MKK-7 dependent, while anisomycin-activated JNK was both MKK-4 and MKK-7 dependent. IL-1beta-induced AP-1 binding activity and AP-1-driven gene expression were strictly MKK-7 dependent. Finally, MMP-3 production only required MKK-7, and there was no effect of MKK-4 deficiency.

Conclusion: These data indicate that only MKK-7 is required for JNK activation in FLS after cytokine stimulation; however, other forms of cellular stress utilize MKK-4. Thus, JNK function might be modulated by targeting MKK-7 to suppress cytokine-mediated FLS activation while leaving other stress responses intact.

Publication types

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

MeSH terms

  • Anisomycin / metabolism
  • Arthritis, Rheumatoid / genetics
  • Arthritis, Rheumatoid / metabolism*
  • Arthritis, Rheumatoid / physiopathology
  • Cells, Cultured
  • Fibroblasts / drug effects
  • Fibroblasts / metabolism*
  • Fibroblasts / physiology
  • Gene Expression Regulation / drug effects
  • Gene Expression Regulation / physiology
  • Humans
  • Interleukin-1 / metabolism
  • JNK Mitogen-Activated Protein Kinases / genetics
  • JNK Mitogen-Activated Protein Kinases / metabolism*
  • MAP Kinase Kinase 4 / metabolism*
  • MAP Kinase Kinase 7 / genetics
  • MAP Kinase Kinase 7 / metabolism*
  • Matrix Metalloproteinase 3 / genetics
  • Matrix Metalloproteinase 3 / metabolism
  • Phosphorylation / drug effects
  • RNA, Small Interfering / genetics
  • Synovial Membrane / drug effects
  • Synovial Membrane / metabolism*
  • Synovial Membrane / physiopathology
  • Transcription Factor AP-1 / genetics
  • Transcription Factor AP-1 / metabolism
  • Transfection
  • Tumor Necrosis Factor-alpha / metabolism


  • Interleukin-1
  • RNA, Small Interfering
  • Transcription Factor AP-1
  • Tumor Necrosis Factor-alpha
  • Anisomycin
  • JNK Mitogen-Activated Protein Kinases
  • MAP Kinase Kinase 4
  • MAP Kinase Kinase 7
  • MAP2K4 protein, human
  • MAP2K7 protein, human
  • Matrix Metalloproteinase 3