Diverse effects of RacV12 on cell transformation by Raf: partial inhibition of morphological transformation versus deregulation of cell cycle control.
Activated Raf kinases and Rac GTPases were shown to cooperate in the oncogenic transformation of fibroblasts, which is characterised by the disassembly of the cellular actin cytoskeleton, a nearly complete loss of focal adhesion complexes and deregulated cell proliferation. This is surprising since the Rac GTPase induces actin structures and the adhesion of suspended cells to extracellular matrix proteins. NIH 3T3 cells expressing a hydroxytamoxifen-inducible oncogenic c-Raf-1-oestrogen receptor fusion protein (c-Raf-1-BxB-ER, N-BxB-ER cells) undergo morphological transformation upon stimulation of the Raf kinase. We show that treatment with the Rac, Rho and Cdc42 activating Escherichia coli toxin CNF1 or coexpression of an activated RacV12 mutant partially inhibits and reverses the disassembly of cellular actin structures and focal adhesion complexes by oncogenic Raf. Activation of the Rac GTPase restores actin structures and focal adhesion complexes at the cellular boundary, leading to spreading of the otherwise spindle-shaped Raf-transformed cells. Actin stress fibres, however, which are regulated by the function of the Rho GTPase, are disassembled by oncogenic Raf even in the presence of activated Rac and Rho. With respect to the RacV12-mediated spreading of Raf-transformed cells, we postulate an anti-oncogenic function of the activated Rac. Another feature of cell transformation is the deregulation of cell cycle control. NIH 3T3 cells expressing high levels of the c-Raf-1-BxB-ER protein undergo a cell cycle arrest upon stimulation of the oncogenic Raf kinase. Our results show that in N-BxB-ER-RacV12 cells the expression of the activated RacV12 mediates cell proliferation in the presence of high-intensity Raf signals and high levels of the Cdk inhibitor p21(Cip1). These results indicate a pro-oncogenic function of the Rac GTPase with respect to the deregulation of cell cycle control.
Kerkhoff
Eugen
E
Institut für Medizinische Strahlenkunde und Zellforschung (MSZ), Universität Würzburg, Versbacher Strasse 5, D-97078 Würzburg, Germany.
Leberfinger
Cornelia B
CB
Schmidt
Gudula
G
Aktories
Klaus
K
Rapp
Ulf R
UR
D016475:3T3 Cells; D000818:Animals; D001427:Bacterial Toxins / Q000494:pharmacology; D002411:Cation Exchange Resins; D002453:Cell Cycle / Q000187:drug effects; D002461:Cell Line, Transformed / Q000187:drug effects*; D003603:Cytotoxins / Q000494:pharmacology; D029968:Escherichia coli Proteins; D008055:Lipids; D051379:Mice; D019908:Proto-Oncogene Proteins c-raf; D013629:Tamoxifen / Q000031:analogs & derivatives / Q000494:pharmacology; D014162:Transfection; D020744:rac GTP-Binding Proteins / Q000494:pharmacology*
D001427:Bacterial Toxins; D002411:Cation Exchange Resins; D003603:Cytotoxins; D029968:Escherichia coli Proteins; D008055:Lipids; C086724:Lipofectamine; D013629:Tamoxifen; C060296:cytotoxic necrotizing factor type 1; D019908:Proto-Oncogene Proteins c-raf; D020744:rac GTP-Binding Proteins