Cell adhesion protects c-Raf-1 against ubiquitin-dependent degradation by the proteasome.
MAP kinase activation by growth factors depends on cell adhesion to the extracellular matrix. Disrupting the cell adhesion process in NIH 3T3 fibroblasts induced an almost complete inhibition of MAP kinase, which was impaired by proteasome inhibitors. In the absence of cell anchorage, c-Raf-1 expression was dramatically decreased after 24 h. This down-regulation was suppressed by proteasome inhibitors, suggesting that a proteasome-dependent degradation of Raf occurred in the absence of cell adhesion. Proteasome inhibitors did not affect Raf-1 levels in adherent cells, indicating that this degradation only occurred in the absence of cell adhesion. Finally, ectopic coexpression of Raf-1 and ubiquitin in HEK-293 and NIH 3T3 cells generated ubiquitylated forms of Raf-1, both in adherent and suspended cells, suggesting a possible ubiquitin-dependent degradation of the protein.
Manenti
Stéphane
S
Laboratoire de Biologie Cellulaire et Moléculaire du Contrôle de la Prolifération, CNRS UMR 5088, Université Paul Sabatier, 118 Route de Narbonne, 31062 Toulouse Cedex, France. manenti@cict.fr
Delmas
Christelle
C
Darbon
Jean-Marie
JM
D016475:3T3 Cells; D000818:Animals; D002448:Cell Adhesion; D002460:Cell Line; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D006801:Humans; D007976:Leupeptins / Q000494:pharmacology; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism*; D025801:Ubiquitin / Q000378:metabolism*
D015853:Cysteine Proteinase Inhibitors; D007976:Leupeptins; D009097:Multienzyme Complexes; D025801:Ubiquitin; D019908:Proto-Oncogene Proteins c-raf; D020928:Mitogen-Activated Protein Kinases; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde