Riley Kasandra J-L, Ramirez-Alvarado Marina, Maher L James
Department of Biochemistry and Molecular Biology, Mayo Clinic College of Medicine, Rochester, Minnesota 55905, USA.
Biochemistry. 2007 Mar 6;46(9):2480-7. doi: 10.1021/bi061480v. Epub 2007 Feb 9.
The tumor suppressor protein p53 is mutated in over half of human cancers. Despite 25 years of study, the complex regulation of this protein remains unclear. After serendipitously detecting RNA binding by p53 in the yeast three-hybrid system (Y3H), we are exploring the specificity and function of this interaction. Electrophoretic mobility shift assays show that full-length p53 binds equally to RNAs that are strongly distinguished in the Y3H. RNA binding blocks sequence-specific DNA binding by p53. The C-terminus of p53 is necessary and sufficient for strong RNA interaction in vitro. Mouse and human C-terminal p53 peptides have different affinities for RNA, and an acetylated human p53 C-terminal peptide does not bind RNA. Circular dichroism spectroscopy of p53 peptides shows that RNA binding does not induce a structural change in the p53 C-terminal peptide, and C-terminal peptides do not detectably affect the structure of RNA. These results demonstrate that p53 binds RNA with little sequence specificity, RNA binding has the potential to regulate DNA binding, and RNA-p53 interactions can be regulated by acetylation of the p53 C-terminus.
肿瘤抑制蛋白p53在超过半数的人类癌症中发生突变。尽管经过了25年的研究，该蛋白的复杂调控机制仍不清楚。在通过酵母三杂交系统（Y3H）意外检测到p53与RNA结合后，我们正在探索这种相互作用的特异性和功能。电泳迁移率变动分析表明，全长p53与在Y3H中表现出显著差异的RNA具有同等程度的结合。RNA结合会阻断p53的序列特异性DNA结合。p53的C末端对于体外强RNA相互作用是必需且足够的。小鼠和人类的p53 C末端肽对RNA具有不同的亲和力，并且乙酰化的人类p53 C末端肽不与RNA结合。p53肽的圆二色光谱表明，RNA结合不会诱导p53 C末端肽的结构变化，并且C末端肽对RNA的结构没有可检测到的影响。这些结果表明，p53以几乎没有序列特异性的方式结合RNA，RNA结合具有调控DNA结合的潜力，并且RNA-p53相互作用可以通过p53 C末端的乙酰化来调控。
