Elastin peptides activate extracellular signal-regulated kinase 1/2 via a Ras-independent mechanism requiring both p110gamma/Raf-1 and protein kinase A/B-Raf signaling in human skin fibroblasts.
Elastin peptides (EPs) produced during cancer progression bind to the elastin binding protein (EBP) found at the surface of dermal fibroblasts, leading to the expression of collagenase-1 gene. The production of this enzyme involved in stromal reaction is caused by the sustained activation of the extracellular signal-regulated kinases 1/2 (ERK1/2) pathway via cAMP/protein kinase A (PKA) and phosphatidylinositol 3-kinase (PI3K). However, the mechanism of these signaling events remains unknown. We show that kappa-elastin (kappaE), a commonly used EP, induces maximum phosphorylation of mitogen-activated protein kinase/extracellular signal-regulated kinase (MEK)1/2 and ERK1/2 after 30 min. The simultaneous inhibition of PKA and PI3K, by N-(2-(p-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide (H89) and 2-(4-morpholynil)-8-phenyl-4H-1-bemzopyran-4-one (LY294002), respectively, blocked MEK1/2 and ERK1/2 phosphorylation, as did lactose, an EBP antagonist. kappaE induced Raf-1 phosphorylation and activation in a PI3K-dependent manner. In our system, the PI3K p110gamma is expressed and activated by betagamma-derived subunits from a pertussis toxin-sensitive G protein after fibroblast stimulation. Pertussis toxin also blocks the Raf-1/MEK1/2/ERK1/2 phosphorylation cascade. In addition, we found that B-Raf is expressed in dermal fibroblasts and activated in a PKA-dependent manner after kappaE treatment, thereby integrating PKA signals to MEK1/2. It is noteworthy that Ras involvement was excluded because ERK1/2 activation by kappaE was not blocked in RasN17-transfected fibroblasts. Together, our results identify a novel Ras-independent ERK1/2 activation system in which p110gamma/Raf-1/MEK1/2 and PKA/B-Raf/MEK1/2 cooperate to activate ERK1/2. Thus, p110gamma and B-Raf seem to be important modulators of dermal fibroblasts physiology and should now qualify as therapeutic targets in strategies aiming at limiting elastin degradation contribution to cancer progression.
Duca
Laurent
L
Laboratoire de Biochimie, UMR CNRS 6198, IFR53 Biomolécules, Faculty of Sciences, Université de Reims Champagne Ardenne, Moulin de la Housse, BP 1039, 51687 Reims Cedex 2, France.
Lambert
Elise
E
Debret
Romain
R
Rothhut
Bernard
B
Blanchevoye
Charlotte
C
Delacoux
Frédéric
F
Hornebeck
William
W
Martiny
Laurent
L
Debelle
Laurent
L
D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000502:physiology*; D004549:Elastin / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D037342:Pertussis Toxin / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology*; D010766:Phosphorylation; D048493:Proto-Oncogene Proteins B-raf / Q000502:physiology*; D019908:Proto-Oncogene Proteins c-raf / Q000502:physiology*; D015398:Signal Transduction; D012867:Skin / Q000166:cytology / Q000378:metabolism; D018631:ras Proteins / Q000502:physiology
D004549:Elastin; D037342:Pertussis Toxin; D019869:Phosphatidylinositol 3-Kinases; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf; D019908:Proto-Oncogene Proteins c-raf; D017868:Cyclic AMP-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D018631:ras Proteins