Ras(V12)-Mediated Down-regulation of CCAAT/Enhancer Binding Protein beta in Immortalized Fibroblasts Requires Loss of p19(Arf) and Facilitates Bypass of Oncogene-Induced Senescence

The transcription factor CCAAT/enhancer binding protein beta (C/EBP beta) is involved in cellular responses to oncogenic and physiologic Ras signals. C/EBP beta is required for premature senescence of primary mouse fibroblasts induced by expression of H-Ras(V12), demonstrating its role in oncogene-induced senescence. Here, we have investigated the mechanisms by which Ras inhibits proliferation of normal cells but transforms immortalized cells. We show that oncogenic Ras down-regulates C/EBP beta expression in NIH 3T3 cells, which are immortalized by a deletion of the CDKN2A locus and, therefore, lack the p16(Ink4a) and p19(Arf) tumor suppressors. Ras(V12)-induced silencing of C/EBP beta occurred at the mRNA level and involved both the Raf-mitogen-activated protein/extracellular signal-regulated kinase (ERK) kinase-ERK and phosphatidylinositol 3-kinase signaling pathways. Oncogenic Ras decreased C/EBP beta expression in Ink4a/Arf(-/-) mouse embryo fibroblasts (MEF) but increased C/EBP beta levels in wild-type MEFs. C/EBP beta down-regulation in NIH 3T3 cells was reversed by expression of p19(Arf), but not of p53 or p16(Ink4a), highlighting a critical role for p19(Arf) in sustaining C/EBP 3 levels. Ectopic expression of p34 C/EBP beta (LAP) inhibited Ras(V12)-mediated transformation of NIH 3T3 cells, suppressed their tumorigenicity in nude mice, and reactivated expression of the proapoptotic Fas receptor, which is also down-regulated by Ras. Our findings indicate that Cebpb gene silencing eliminates a growth inhibitory transcription factor that would otherwise restrain oncogenesis. We propose that C/EBP beta is part of a p53-independent, p19(Arf)-mediated network that enforces Ras-induced cell cycle arrest and tumor suppression in primary fibroblasts. [Cancer lies 2009,69(6):2588-98]

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