Galbiati Laura, Mendoza-Maldonado Ramiro, Gutierrez Maria Ines, Giacca Mauro
Molecular Medicine Laboratory, International Centre for Genetic Engineering and Biotechnology, Trieste, Italy.
Cell Cycle. 2005 Jul;4(7):930-9. doi: 10.4161/cc.4.7.1784. Epub 2005 Jul 25.
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.
在此，我们报告一种将乙酰化与泛素化联系起来的新型非竞争性机制，其中转录因子E2F-1与细胞共激活因子及乙酰转移酶p300的结合决定了其乙酰化及随后的泛素化。通过使用特异性识别E2F-1乙酰化形式（AcE2F-1）的抗体，我们发现，DNA损伤后，AcE2F-1在细胞中呈时间依赖性积累，且p300的表达会增加乙酰化水平。值得注意的是，相同的DNA损伤条件也会诱导泛素化的E2F-1积累，这一事件同样会被p300的过表达显著刺激。p300对E2F-1泛素化的影响需要p300的HAT结构域以及E2F-1中三个乙酰化赖氨酸的完整性。值得注意的是，p300诱导的E2F-1泛素化不依赖于p45Skp2 E3连接酶，因为它不会扩展到其他p45Skp2靶点，并且在一个缺乏p45Skp2结合结构域但仍保留乙酰化区域的E2F-1突变体中也会发生。最后，p300诱导的E2F-泛素化不受RB的影响。
