The interplay between the chromatin remodeler INO80 and histone variant H2A.Z.

染色质重塑因子 INO80 和组蛋白变体 H2A.Z 之间的相互作用。

基本信息

批准号：
8887625
负责人：
Blaine Bartholomew
金额：
$ 43.42万
依托单位：
UNIVERSITY OF TX MD ANDERSON CAN CTR
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-05-01 至 2019-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8887625
关键词：
Acetylation Address B-Lymphocytes Binding Cell Cycle Cell Proliferation Cells Centromere Chromatin Chromatin Fiber Chromatin Remodeling Factor Chromatin Structure Chromosomes Complex DNA DNA Crosslinking DNA Damage DNA Repair DNA biosynthesis Development Eukaryota Event Gene Activation Gene Expression Genes Genetic Transcription Genome Genome Stability Goals Histone H3 Histones ISWI In Vitro Link Lymphoma Lysine Maintenance Malignant Neoplasms Maps Mission Modeling Molecular Mutagenesis Neoplasm Metastasis Nucleosomes Oncogene Activation Pathway interactions Patients Process Promoter Regions Public Health RNA Polymerase II Reaction Recruitment Activity Refractory Regulation Research Resolution Role Site Specificity Suggestion Techniques Transcriptional Activation Variant War Work base cell transformation chromatin remodeling crosslink dimer embryonic stem cell genome integrity human disease in vivo mutant prevent promoter public health relevance repaired single molecule telomere telomere loss

项目摘要

DESCRIPTION (provided by applicant): The objective of this proposal is to examine the interplay between the ATP-dependent remodeler INO80 and the histone variant H2A.Z in order to better understand their roles in activating transcription and preventing bidirectional transcription from promoter regions. It has been confusing as to the role of H2A.Z in transcription given the contradictory information. Incorporation of H2A.Z in vitro into nucleosomes and nucleosomal arrays stabilizes the nucleosome more than canonical H2A and promotes higher-order folding into a chromatin fiber, all of which suggests H2A.Z should be refractory to transcription. However, H2A.Z is a consistent marker of active genes in higher eukaryotes and has been thought to "poise" genes for transcription. Early in transcription H2A.Z is rapidly replaced by H2A from promoter regions and H2A.Z nucleosomes are more rapidly turned over and lost than canonical nucleosomes leading to the suggestion that they can "open" the chromatin structure. Although we know INO80 exchanges H2A.Z for H2A during remodeling, we know little about how this happens and if the reaction intermediates involved predispose nucleosomes to become more accessible and prone to disassemble. In addressing whether INO80 remodeling of H2A.Z nucleosomes can explain the discrepancy between in vitro and in vivo observations, the resolution of this dilemma will not only answer basic question in transcription but also potentially in DNA repair, genomic stability linked to the formation of centromeres and telomeres, and replication fork stability; all of which involve INO80 and H2A.Z. The binding interface between INO80 and nucleosomes will be mapped by determining the INO80 domains associated with specific sites in core histones and nucleosomal and extranucleosomal DNA when INO80 is bound. The functional relevance of the crosslinked regions in INO80 for binding and remodeling will be examined by deletion or mutagenesis of conserved residues within these domains. The structural dynamics of histone exchange will be examined using ensemble and single molecule techniques to determine the rate and order of events such as disruption of the contacts of H2A.Z-H2B with DNA and the H3-H4 tetramer, and its eventual loss from nucleosomes. We will determine the parts of INO80 facilitating dimer ejection and the entry of the incoming H2A-H2B dimer using the described mutants. The effects of acetylation of lysine 56 of histone H3 (H3K56ac) on the contacts of INO80 with nucleosomes will be examined by histone and DNA crosslinking. Alterations in the reaction pathways of H2A.Z and H2A exchange when nucleosomes contain H3K56ac versus unacetylated H3 will be examined using the same ensemble and single molecule techniques to understand how acetylation enhances the exchange reaction.

描述（通过应用程序证明）：研究ATP依赖性重塑和Hist One变体H2A.Z之间的解释的目的，以更好地在激活转录和从启动子区域进行预先转录方面的作用。在转录中，H2A.Z矛盾的阵列稳定了核小体，而不是规范的H2A，并促进了较高的染色质纤维，所有这些都表明H2A.Z应该是转录的转录早期的“平衡”基因。用于重塑过程中的H2a的.z发生，中间体会使volded核体变得更加易于拆卸。所有这些都涉及INO80和H2A.Z。INO80和核聚体之间的结合界面与核心组蛋白和核小体和核核体外DNA中的特定位点相关的INO80域，当时INO80通过INO80结合了INO80的区域。将使用集合和单个分子检查组蛋白交换的y域的删除或杂种。核小体的损失我们将使用分解的H3（H3K56AC）在Ino80的接触中促进输入的H2A-H2B二聚体的二聚体当核糖体含有H3K56AC与非乙酰化H3时，H2A.Z和H2A交换的反应途径将使用相同的Semble和单分子技术检查，以了解乙酰化如何增强交换反应。