Defects in TGF beta signaling overcome senescence of mouse keratinocytes expressing v-ras(Ha)

Previous studies have shown that TGF beta 1 expression is upregulated in mo use keratinocytes infected with a v-ras(Ha) retrovirus, although the functi onal significance of this has not been clear. Here we show that v-ras(Ha) r etrovirus transduced primary mouse keratinocytes undergo hyperproliferation followed by a TGF beta 1 dependent G1 growth arrest and senescence. The gr owth arrest is accompanied by a 15-fold increase in total TGF beta 1 secret ed and a fourfold increase in secreted active TGF beta 1. When cultured in the presence of a neutralizing antibody to TGF beta 1, the senescence respo nse is suppressed. Levels of the TGF beta 1 target p15(ink4b) increase duri ng senescence as does association of this kinase inhibitor with cyclinD/cdk 4 complexes. However, p16(ink4a), p53 and p19(ARF) expression also increase during senescence. Genetic analysis shows that TGF beta 1 null and dominan t negative T beta RII expressing v-ras(Ha) keratinocytes resist the G1 grow th arrest and do not senescence. This resistance is associated with low exp ression of p15(ink4b) and p16(ink4a), constitutive Rb phosphorylation and h igh levels of cdk4 and cdk2 kinase activity. In contrast, inactivation of T GF beta 1 secretion or response does not block the induction of p53 and p19 (ARF), but the level of p21(waf1), a p53 target gene, is reduced in cyclin D/cdk4 and cyclin E/cdk2 complexes. Thus, although multiple senescence path ways are activated in response to a ras oncogene, inactivation of TGF beta 1 secretion or response is sufficient to block the senescence program. Sinc e v-ras(Ha) transduced TGF beta 1-/- keratinocytes form squamous cell carci nomas following skin grafting, these results suggest that in mouse keratino cytes, defects in TGF beta 1 signaling accelerate malignant progression by overcoming oncogene induced replicative senescence.