p53-induced Regulation of Transcription in the Chromatin Context

p53 诱导的染色质转录调节

• 批准号: 10014467
• 负责人: Victor Zhurkin
• 金额: $ 10.13万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10014467
• 关键词: Affect; Affinity; Agreement; Anisotropy; Apoptosis; Apoptotic; Binding; Binding Sites; Biological Process; Cell Cycle Arrest; Cell physiology; Cells; Chromatin; Chromatin Loop; Complex; Computer Analysis; DNA; DNA Binding; DNA Methylation; Data; Deoxyribonuclease I; Dimensions; Elements; Environment; Epigenetic Process; Exhibits; Family; GADD45A gene; Gene Activation; Genes; Genetic Transcription; Genomic DNA; Genomics; Goals; Histone H2A; Histones; Human; In Vitro; Kinetics; Major Groove; Malignant Neoplasms; Measures; Modeling; Molecular Conformation; Motivation; Normal Cell; Nucleosomes; Pathway interactions; Pattern; Positioning Attribute; Publishing; Response Elements; Rotation; Scheme; Sequence Analysis; Shapes; Side; Signal Transduction; Site; Surface; TP53 gene; Tail; Time; Transactivation; Transcriptional Regulation; Tumor Suppression; Work; cancer cell; cell type; epigenetic regulation; experimental study; gel electrophoresis; gene induction; histone modification; in vivo; insight; novel; organizational structure; prevent; response; transcription factor

As a transcription factor, p53 modulates expression of thousands of genes involved in a variety of cellular functions. p53 exhibits high affinity to the response elements (REs) regulating cell cycle arrest genes (CCA-sites), but relatively low affinity to the sites associated with apoptosis (Apo-sites). Since p53 can bind nucleosomal DNA, we sought to understand if the two groups of p53 sites differ in their accessibility when embedded in nucleosomes. To this aim, we analyzed the sequence-dependent bending anisotropy of human genomic DNA containing p53 sites. We calculated rotational positioning patterns predicting that most of the CCA-sites are exposed on the nucleosomal surface. This is consistent with experimentally observed positioning of human nucleosomes. Remarkably, the sequence-dependent DNA anisotropy of both the p53 sites and flanking DNA work in concert producing strong positioning signals. By contrast, both the predicted and observed rotational settings of the Apo-sites in nucleosomes suggest that many of these sites are buried inside, thus preventing immediate p53 recognition and delaying gene induction. We also measured the p53 binding to its cognate sites embedded in the in strongly positioned '601' nucleosome. Our data suggest that the p53 affinity to DNA strongly correlates with the rotational positioning of its site in nucleosome, in agreement with the computational analysis described above. The exposed configurations of the p53 sites in nucleosome (like CCA-sites) demonstrate significantly stronger affinity to p53 compared to the buried configurations (similar to the Apo-sites). Thus, the difference in nucleosomal organization of the two sets of p53 response elements appears to be a key factor affecting the strength of p53-DNA binding and kinetics of induction of the p53 target genes. Our model differs from the earlier concept connecting the selective activation of the CCA- and Apo-genes to the binding affinities of their REs to p53. Instead, we emphasize a direct correlation between the selection of p53-induced tumor suppression pathway (apoptosis versus cell cycle arrest) and structural organization of the corresponding p53-binding sites in chromatin. We add new dimensions to the existing paradigm, namely, the relative positioning and chromatin environment of the p53 REs. Our scheme not only explains the above cases but also provides a new insight into the cellular mechanisms of activation of hundreds of genes by p53. Recently, we performed a comprehensive analysis of 25 published p53 cistromes and identified thousands binding sites in normal and cancer cells. Our analysis revealed two distinct epigenetic features underlying p53-DNA interactions in vivo. First, we found that p53 binding sites are associated with transcriptionally active histone marks (H3K4me3 and H3K36me3) in normal-cell chromatin, but with repressive histone marks (H3K27me3) in cancer-cell chromatin. Second, p53 binding sites in cancer cells are characterized by a lower level of DNA methylation than their counterparts in normal cells, probably related to global hypomethylation in cancers. In addition, regardless of the cell type, p53 sites are highly enriched in the endogenous retroviral elements of the ERV1 family, highlighting the importance of this repeat family in shaping the transcriptional network of p53. Moreover, the p53 sites exhibit an unusual combination of chromatin patterns: high nucleosome occupancy and, at the same time, high sensitivity to DNase I. Our results suggest that p53 can access its target sites in a chromatin environment that is non-permissive to most DNA-binding factors, which may allow p53 to act as a pioneer transcription factor in the context of chromatin. Furthermore, our preliminary data suggest that the p53-DNA binding affinity is modulated by the histone H2A N-tails interacting with DNA in the vicinity of the RE. Potentially, this novel observation may have far-reaching implications for epigenetic regulation of p53 (and other TFs) binding to chromatin.

作为一种转录因子，p53调节参与多种细胞功能的数千个基因的表达。p53对调节细胞周期阻滞基因的反应元件（RE）（CCA位点）表现出高亲和力，但对与凋亡相关的位点（Apo位点）表现出相对低的亲和力。由于p53可以结合核小体DNA，我们试图了解两组p53位点嵌入核小体时的可及性是否不同。为此，我们分析了含有p53位点的人类基因组DNA的序列依赖性弯曲各向异性。我们计算了旋转定位模式预测，大部分的CCA-网站暴露在核小体表面上。这与实验观察到的人类核小体的定位一致。值得注意的是，p53位点和侧翼DNA的序列依赖性DNA各向异性协同工作，产生强烈的定位信号。相比之下，预测和观察到的旋转设置的载脂蛋白位点的核小体表明，许多这些网站被埋在里面，从而防止立即p53识别和延迟基因诱导。我们还测量了p53与其嵌入在强定位的“601”核小体中的同源位点的结合。我们的数据表明，p53与DNA的亲和力与其在核小体中的位点的旋转定位密切相关，与上述计算分析一致。核小体中p53位点的暴露构型（如CCA-位点）与埋藏构型（类似于Apo-位点）相比，对p53表现出显著更强的亲和力。因此，两组p53反应元件的核小体组织的差异似乎是影响p53-DNA结合强度和p53靶基因诱导动力学的关键因素。我们的模型不同于早期的概念连接CCA和载脂蛋白基因的选择性激活的结合亲和力的RE的p53。相反，我们强调p53诱导的肿瘤抑制途径（细胞凋亡与细胞周期阻滞）的选择和相应的p53结合位点在染色质中的结构组织之间的直接相关性。我们增加了新的维度，现有的范例，即，相对定位和染色质环境的p53 REs。我们的计划不仅解释了上述情况下，但也提供了一个新的见解，数百个基因的激活的细胞机制的p53。最近，我们对25个已发表的p53顺式序列进行了全面分析，并在正常细胞和癌细胞中鉴定了数千个结合位点。我们的分析揭示了两个不同的表观遗传特征潜在的p53-DNA相互作用在体内。首先，我们发现p53结合位点与正常细胞染色质中的转录活性组蛋白标记（H3 K4 me 3和H3 K36 me 3）相关，但与癌细胞染色质中的抑制性组蛋白标记（H3 K27 me 3）相关。第二，癌细胞中的p53结合位点的特征在于DNA甲基化水平低于正常细胞中的对应物，这可能与癌症中的整体低甲基化有关。此外，无论细胞类型如何，p53位点高度富集在ERV 1家族的内源性逆转录病毒元件中，突出了该重复序列家族在形成p53转录网络中的重要性。此外，p53位点表现出一种不寻常的染色质模式组合：高核小体占有率，同时对DNA酶I具有高敏感性。我们的研究结果表明，p53可以访问它的靶位点在染色质环境中，是不允许的大多数DNA结合因子，这可能允许p53作为一个先锋转录因子的背景下，染色质。此外，我们的初步数据表明，p53-DNA结合亲和力是由组蛋白H2 A N-尾与RE附近的DNA相互作用调节的。潜在地，这一新的观察结果可能对p53（和其他TF）与染色质结合的表观遗传调控具有深远的影响。