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The role of the MBD2-NuRD complex in gamma-globin gene silencing

MBD2-NuRD 复合物在 γ-珠蛋白基因沉默中的作用

基本信息

批准号：
10208866
负责人：
GORDON D GINDER
金额：
$ 65.66万
依托单位：
VIRGINIA COMMONWEALTH UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-08-01 至 2023-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10208866
关键词：
Address Adult Animal Model Binding Bioinformatics Biological Biological Assay Bioluminescence Biophysics CHD4 gene Cancer Cell Growth Cell model ChIP-seq Chromatin Remodeling Factor Complex DNA Methylation Data Dependence Development Disease Embryo Energy Transfer Epigenetic Process Erythroid Erythroid Cells Feedback Fetal Hemoglobin Future Gene Abnormality Gene Expression Gene Expression Regulation Gene Silencing Genes Genetic Globin Goals Hematologic Neoplasms Histone Deacetylase Human Individual Knockout Mice Laboratories Link Maintenance Malignant Neoplasms Mediating Molecular Target Mus NuRD complex Pathway interactions Peptides Phenotype Play Proteins Public Health Regulation Relaxation Repressor Proteins Role Sickle Cell Anemia Structure Techniques Testing Therapeutic Time Transgenes Transgenic Organisms Tumor Suppressor Genes Ursidae Family Validation Work beta Globin beta Thalassemia chromatin immunoprecipitation chromatin remodeling effective therapy experimental study fetal gamma Globin genetic corepressor genome editing in vivo insight leukemia molecular targeted therapies novel physical separation protein protein interaction recruit small molecule therapeutic target therapy development transcription factor transcriptome sequencing

项目摘要

PROJECT SUMMARY Studies of developmental regulation of globin gene expression have provided important mechanistic insight into normal mammalian gene control and abnormal gene expression in diseases. Epigenetic mechanisms are now recognized as central to globin gene regulation as well as dysregulation of genes in leukemia and other cancers. This project is aimed at elucidating the key protein-protein interactions among components of the MBD2-NuRD chromatin remodeling complex in the context of fetal β-type globin gene silencing in adult human erythroid cells. The long term goal is to identify and validate targets for safe therapeutic activation of fetal hemoglobin expression in sickle cell anemia and β-Thalassemia. This goal will be pursued through the following collaborative aims: 1) To functionally define the roles of specific regions of MBD2 and NuRD complex components in the ability of MBD2-NuRD to silence the fetal γ-globin gene in adult human erythroid cells and 2) to biophysically and structurally characterize interfaces critical for MBD-NuRD complex formation and stability. The experimental approach employs a real-time iterative feedback between genome editing of key protein interactions in adult erythroid cells in conjunction with structural studies of key protein-protein interactions of MBD2, GATAD2A and CHD4 NuRD components using both standard NMR, and crystallographic techniques as well as state-of-the-art paramagnetic relaxation enhancement (PRE) and bioluminescence resonance energy transfer (BRET). The erythroid genes that are directly activated by disruption of MBD2-NuRD and in turn relieve γ-globin gene silencing will be characterized by both RNA-seq and ChIP-seq assays and bioinformatics analyses as well as ChIP assay validation. These experiments will identify and validate proof of principle peptide and small molecule targets for future development of therapy of sickle cell anemia and β-Thalessemia.

项目摘要珠蛋白基因表达的发育调控研究提供了重要的机制见解正常哺乳动物基因控制和疾病中的异常基因表达。表观遗传机制是现在被认为是珠蛋白基因调节以及白血病和其他疾病中基因失调的中心。癌的该项目旨在阐明蛋白质-蛋白质相互作用的关键组成部分， MBD 2-NuRD染色质重塑复合物在成人胎儿β型珠蛋白基因沉默中的作用红系细胞长期目标是确定和验证安全治疗激活胎儿的靶点，镰状细胞性贫血和β-地中海贫血血红蛋白表达这一目标将通过以下合作目标：1）功能上定义MBD 2和NuRD复合物特定区域的作用 MBD 2-NuRD沉默成人红系细胞中胎儿γ-珠蛋白基因的能力中的组分， 2)从生物药理学和结构上表征MBD-NuRD复合物形成的关键界面，稳定该实验方法采用了关键基因组编辑之间的实时迭代反馈，成人红系细胞中蛋白质相互作用及关键蛋白质-蛋白质结构研究 MBD 2、GATAD 2A和CHD 4 NuRD组分的相互作用，晶体学技术以及最先进的顺磁弛豫增强（PRE）和生物发光共振能量转移（BRET）。直接被激活的红细胞基因 MBD 2-NuRD的破坏并进而减轻γ-珠蛋白基因沉默将通过RNA-seq 和ChIP-seq测定和生物信息学分析以及ChIP测定验证。这些实验将鉴定和验证用于未来治疗发展的主要肽和小分子靶点的证据镰状细胞性贫血和β-地中海贫血。