Kearns Sinéad, Lurz Rudi, Orlova Elena V, Okorokov Andrei L
Institute for Structural and Molecular Biology, School of Biological Sciences, Birkbeck College, London WC1E 7HX, UK Wolfson Institute for Biomedical Research, Division of Medicine, University College London, London WC1E 6BT, UK.
Max Planck Institute for Molecular Genetics, Ihnestrasse, Berlin 14195, Germany.
Nucleic Acids Res. 2016 Jul 27;44(13):6185-99. doi: 10.1093/nar/gkw215. Epub 2016 Mar 31.
p53 tumor suppressor is a transcription factor that controls cell cycle and genetic integrity. In response to genotoxic stress p53 activates DNA repair, cell cycle arrest, apoptosis or senescence, which are initiated via p53 binding to its specific DNA response elements (RE). The consensus p53 DNA RE consists of two decameric palindromic half-site sequences. Crystallographic studies have demonstrated that two isolated p53 DNA-binding core domains interact with one half-site of the p53 DNA REs suggesting that one p53 tetramer is bound to one RE. However, our recent 3D cryo-EM studies showed that the full-length p53 tetramer is bound to only one half-site of RE.Here, we have used biochemical and electron microscopy (EM) methods to analyze DNA-binding of human and murine p53 tetramers to various p53 DNA REs. Our new results demonstrate that two p53 tetramers can interact sequence-specifically with one DNA RE at the same time. In particular, the EM structural analysis revealed that two p53 tetramers bind one DNA RE simultaneously with DNA positioned between them. These results demonstrate a mode different from that assumed previously for the p53-DNA interaction and suggest important biological implications on p53 activity as a transcriptional regulator of cellular response to stress.
p53肿瘤抑制因子是一种控制细胞周期和基因完整性的转录因子。响应基因毒性应激时，p53会激活DNA修复、细胞周期停滞、凋亡或衰老，这些过程通过p53与其特定DNA反应元件（RE）结合而启动。p53的共有DNA反应元件由两个十聚体回文半位点序列组成。晶体学研究表明，两个分离的p53 DNA结合核心结构域与p53 DNA反应元件的一个半位点相互作用，这表明一个p53四聚体与一个反应元件结合。然而，我们最近的三维冷冻电镜研究表明，全长p53四聚体仅与反应元件的一个半位点结合。在此，我们使用生化和电子显微镜（EM）方法分析人和小鼠p53四聚体与各种p53 DNA反应元件的DNA结合情况。我们的新结果表明，两个p53四聚体可以同时与一个DNA反应元件进行序列特异性相互作用。特别是，EM结构分析显示，两个p53四聚体同时结合一个DNA反应元件，且DNA位于它们之间。这些结果证明了一种与先前假设的p53-DNA相互作用模式不同的模式，并暗示了p53作为细胞应激反应转录调节因子的活性具有重要的生物学意义。
