Abstract
Primary fibroblasts respond to activated H-RAS(V12) by undergoing premature arrest, which resembles replicative senescence. This irreversible 'fail-safe mechanism' requires p19(ARF), p53 and the Retinoblastoma (Rb) family: upon their disruption, RAS(V12)-expressing cells fail to undergo senescence and continue to proliferate. Similarly, co-expression of oncogenes such as c-MYC or E1A rescues RAS(V12)-induced senescence. To identify novel genes that allow escape from RAS(V12)-induced senescence, we designed an unbiased, retroviral complementary DNA library screen. We report on the identification of DRIL1, the human orthologue of the mouse Bright and Drosophila dead ringer transcriptional regulators. DRIL1 renders primary murine fibroblasts unresponsive to RAS(V12)-induced anti-proliferative signalling by p19(ARF)/p53/p21(CIP1), as well as by p16(INK4a). In this way, DRIL1 not only rescues RAS(V12)-induced senescence but also causes these fibroblasts to become highly oncogenic. Furthermore, DRIL1 immortalizes mouse fibroblasts, in the presence of high levels of p16(INK4a). Immortalization by DRIL1, whose product binds the pRB-controlled transcription factor E2F1 (ref. 8), is correlated with induction of E2F1 activity. Correspondingly, DRIL1 induces the E2F1 target Cyclin E1, overexpression of which is sufficient to trigger escape from senescence. Thus, DRIL1 disrupts cellular protection against RAS(V12)-induced proliferation downstream of the p19(ARF)/p53 pathway.