Influences of DNA sequence and histone features on transcription factor binding to nucleosomes

DNA 序列和组蛋白特征对转录因子与核小体结合的影响

• 批准号: 10528812
• 负责人: MARTHA L BULYK
• 金额: $ 70.74万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10528812
• 关键词: Affect; Binding; Binding Sites; Biochemical; Biological Assay; Breathing; Cells; ChIP-seq; Chromatin; Computer Analysis; DNA; DNA Binding; DNA Binding Domain; DNA Sequence; Data; Elements; Epigenetic Process; Etiology; Eukaryota; Family; Gatekeeping; Gene Expression; Gene Expression Regulation; Genetic Transcription; Genets; Genome; Genomics; Heterochromatin; Histone H2A; Histones; Human; In Vitro; Invaded; Manuscripts; Nuclear; Nucleic Acid Regulatory Sequences; Nucleosomes; Periodicity; Play; Positioning Attribute; Post-Translational Protein Processing; Preparation; Property; Reader; Regulator Genes; Regulatory Element; Role; Site; Slide; Specificity; Surveys; Technology; Variant; cell type; cofactor; genomic data; high throughput screening; high throughput technology; histone modification; in vivo; molecular modeling; novel; preference; programs; reconstitution; recruit; transcription factor

Abstract Gene expression programs are dynamically regulated by the accessibility of chromatin for transcription factor (TF) binding, but how TFs recognize specific regulatory regions occluded by nucleosomes remains unclear. Certain TFs, termed pioneer factors, can recognize their target sites within nucleosomes, leading to the opening of chromatin. By priming cis-regulatory elements for subsequent transcriptional regulatory activity, pioneers serve as gatekeepers to cellular differentiation. Although pioneers can bind nucleosomal sites, they bind only a subset of their potential recognition sites in the genome that typically varies across cell types, thus indicating their interplay with sequence, epigenetic or other cellular features. Despite the importance of pioneer factors, what restricts pioneer binding is poorly understood. Little is known about how the sequence context of their sites in nucleosomes, the presence of histone variants or post-translational modifications (PTMs) of histones, or interactions with cofactors or chromatin readers that recognize those PTMs might influence pioneer binding to nucleosomes. No high-throughput technologies have been developed to survey the impact of these many parameters on TF pioneer binding. In this project, we will develop novel, high-throughput biochemical assays to investigate how nucleosomal sequence context, histone variants or histone PTMs influence pioneer binding of human TFs to nucleosomes. We will also investigate the interplay of pioneers, cofactors, and chromatin readers in pioneer binding. Results from these biochemical assays will be validated in vitro and used in analysis of in vivo genomic data in human cells to understand how these various features contribute to TF pioneer binding in cells. As pioneer factors play crucial roles at the top of regulatory hierarchies, these results will aid in understanding how gene regulation of cell states is encoded in the genome and the mechanisms by which it is read out.

抽象的 基因表达程序受染色质的可及性动态调节 转录因子（TF）结合，但是TF如何识别特定的调节区域被遮挡 核小体尚不清楚。某些TF，称为先驱因素，可以识别其目标 核小体内的位点，导致染色质开放。通过启动顺式调节 随后的转录调节活动的元素，先驱者是守门人 细胞分化。尽管开拓者可以结合核小体部位，但它们仅结合一个子集 它们在基因组中的潜在识别位点通常会随细胞类型而变化，因此 指示它们与序列，表观遗传或其他细胞特征的相互作用。 尽管先锋因素很重要，但限制了先锋绑定的原因知之甚少。 关于其位点在核小体中的序列上下文的了解知之甚少 组蛋白变体或组蛋白的翻译后修饰（PTM）或与 识别这些PTM的辅助因子或染色质读取器可能会影响先锋的结合 核小组。尚未开发高通量技术来调查 这些在TF开拓者结合上的许多参数。 在这个项目中，我们将开发出新颖的高通量生化测定法，以研究如何 核小体序列上下文，组蛋白变异或组蛋白PTM会影响先锋的结合 人类TF到核小体。我们还将研究开拓者，辅助因子和 先锋结合中的染色质读取器。这些生化测定的结果将在 体外并用于分析人类细胞中体内基因组数据，以了解这些数据 各种特征有助于细胞中的TF先锋结合。 随着先锋因素在监管层次结构的顶部起着至关重要的作用，这些结果将有助于 了解如何在基因组中编码细胞态的基因调节 读出的机制。