The induction of cellular senescence by oncogenic signals acts as a barrier to cellular transformation and is attained, in part, by the elevation of the p16^INK4a tumor suppressor gene. p16 expression is repressed epigenetically by Polycomb, but how p16 is induced is not known. We report here that the p16 locus is H3K4-methylated in highly expressing cells. H3K4 methyltransferase MLL1 directly binds to and is required, along with its core component RbBP5, for the induction of p16 by oncogenic Ras. We further show that damaged DNA binding protein DDB1 and CUL4, which assemble distinct E3 ubiquitin ligases by recruiting various WD40 proteins, act upstream of MLL1-mediated H3K4 methylation. We showed that CUL4A directly binds to p16 and that silencing DDB1 blocks Ras-induced p16 activation. Ras expression dissociates BMI1 from the p16 locus, whereas both CUL4 and MLL1 bind to the p16 locus similarly in both normal and oncogenic stimulated cells. These results suggest that DDB1-CUL4 and MLL1 complexes constitute a novel pathway that mediates p16 activation during oncogenic checkpoint response and is repressed by the polycomb repression complexes during normal growth of young cells. [Cancer Res 2009;69(5):1809–14]