One-Dimensional Search Dynamics of Tumor Suppressor p53 Regulated by a Disordered C-Terminal Domain
One-Dimensional Search Dynamics of Tumor Suppressor p53 Regulated by a Disordered C-Terminal Domain
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DOI：
10.1016/j.bpj.2017.04.038
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发表时间：
2017-06-06
影响因子：
3.4
通讯作者：
Kamagata, Kiyoto
中科院分区：
文献类型：
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作者：
Murata, Agato;Itoh, Yuji;Kamagata, Kiyoto
肿瘤抑制基因p53在细胞条件的巨大变化中，沿着沿着DNA滑动并找到其靶序列。为了阐明p53如何在不同浓度的二价阳离子如Ca 2+和Mg 2+下保持有效的靶搜索，我们制备了两种p53突变体，每种突变体具有其两个DNA结合结构域之一，Core泰特突变体具有结构化的核心结构域加上四聚化（泰特）结构域，TetCT突变体具有泰特加上无序的C末端结构域。我们研究了它们在不同[Mg 2 +]下从DNA上的平衡和动力学解离以及沿DNA的沿着搜索动力学。尽管在较高的[Mg 2 +]下，CoreTet与DNA的结合明显减弱，但TetCT的结合轻微依赖于[Mg 2 +]。单分子荧光测量结果表明，CoreTet沿着DNA的一维扩散由快搜索模式和慢搜索模式组成，其比例强烈依赖于[Mg 2 +]。相反，TetCT的扩散仅由快速模式组成。无序的C-末端结构域可以与DNA结合而不受[Mg 2 +]的影响，并且可以维持两种搜索模式和p53搜索距离的平衡。这些结果表明，p53在不同的[Mg 2 +]下调节p53与DNA之间的复合物的四级结构，并且它维持沿着DNA的靶搜索。
Tumor suppressor p53 slides along DNA and finds its target sequence in drastically different and changing cellular conditions. To elucidate how p53 maintains efficient target search at different concentrations of divalent cations such as Ca2+ and Mg2+, we prepared two mutants of p53, each possessing one of its two DNA-binding domains, the CoreTet mutant having the structured core domain plus the tetramerization (Tet) domain, and the TetCT mutant having Tet plus the disordered C-terminal domain. We investigated their equilibrium and kinetic dissociation from DNA and search dynamics along DNA at various [Mg2+]. Although binding of CoreTet to DNA becomes markedly weaker at higher [Mg2+], binding of TetCT depends slightly on [Mg2+]. Single-molecule fluorescence measurements revealed that the one-dimensional diffusion of CoreTet along DNA consists of fast and slow search modes, the ratio of which depends strongly on [Mg2+]. In contrast, diffusion of TetCT consisted of only the fast mode. The disordered C-terminal domain can associate with DNA irrespective of [Mg2+], and can maintain an equilibrium balance of the two search modes and the p53 search distance. These results suggest that p53 modulates the quaternary structure of the complex between p53 and DNA under different [Mg2+] and that it maintains the target search along DNA.