Regulation of E2F-1 after DNA damage by p300-mediated acetylation and ubiquitination.

Here we report a novel, noncompetitive mechanism that links acetylation and ubiquitination, in which the association of transcription factor E2F-1 with the cellular coactivator and acetyltransferase p300 determines its acetylation and subsequent ubiquitination. By using an antibody specifically recognizing the acetylated form of E2F-1 (AcE2F-1), we found that, after DNA damage, AcE2F-1 accumulates in the cells in a time-dependent manner, and that acetylation is increased by the expression of p300. Remarkably, the same DNA damaging conditions also induce the accumulation of ubiquitinated E2F-1, an event that is again markedly stimulated by p300 overexpression. The effects of p300 on E2F-1 ubiquitination require the integrity of the HAT domain of p300 and of the three acetylated lysines in E2F-1. Of note, p300-induced E2F-1 ubiquitination does not depend on the p45Skp2 E3 ligase, since it does not extend to other p45Skp2 targets and also occurs with an E2F-1 mutant devoid of the p45Skp2-binding domain but still retaining the acetylated region. Finally, p300-induced E2F-1 ubiquitination is not influenced by RB.