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：
9976500
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/9976500
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.

项目摘要球蛋白基因表达的发育调控研究提供了重要的机械洞察力进入疾病中正常的哺乳动物基因控制和异常基因表达。表观遗传机制是现在被认为是环球蛋白基因调节的核心以及白血病和其他基因的失调癌症。该项目旨在阐明该项目之间的关键蛋白质 - 蛋白质相互作用在成年人的胎儿β型球蛋白基因沉默的背景下，MBD2-nurd染色质重塑复合物红细胞细胞。长期目标是识别和验证胎儿安全治疗激活的靶标镰状细胞性贫血和β-丘脑贫血的血红蛋白表达。这个目标将通过以下协作目的：1）在功能上定义MBD2和Nurd Complex的特定区域的作用 MBD2-nurd在成年人类红细胞细胞中胎儿γ-珠蛋白基因沉默的能力的成分 2）生物物理和结构表征对MBD-nurd复合物形成至关重要的接口和稳定。实验方法采用了钥匙基因组编辑之间的实时迭代反馈成年红细胞细胞中的蛋白质相互作用与关键蛋白质蛋白的结构研究结合使用标准NMR和晶体学技术以及最先进的顺磁性松弛增强（PRE）和生物发光共振能量转移（BRET）。直接激活的红细胞基因 MBD2-nurd的破坏和拯救γ-珠蛋白基因沉默将以RNA-Seq的表征以及CHIP-SEQ分析和生物信息学分析以及CHIP测定验证。这些实验会识别和验证原理胡椒和小分子靶标的证明镰状细胞性贫血和β-硫代贫血。