Regulation of Globin Gene Switching by O-GlcNAc Post-Translational Modification

O-GlcNAc 翻译后修饰调控球蛋白基因转换

• 批准号: 8610686
• 负责人: KENNETH R PETERSON
• 金额: $ 26.27万
• 依托单位: UNIVERSITY OF KANSAS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-01-01 至 2016-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8610686
• 关键词: Address; Adult; Adverse effects; Affect; African American; Binding Sites; Biochemical; Biological; Cell Culture Techniques; Cell Cycle; Cell physiology; Cells; Chromatin Remodeling Factor; Complex; Cooley' s anemia; Cytoplasmic Protein; DNA Binding; DNA-Binding Proteins; Data; Development; Disease; Drosophila genus; Enzymes; Erythrocytes; Erythroid; Erythroid Cells; Erythropoiesis; Fetal Hemoglobin; Fetal Liver; Functional disorder; Gene Expression; Gene Silencing; Genes; Genetic Transcription; Globin; Glucosamine; Goals; Hemoglobinopathies; Hereditary Disease; Homeobox Genes; Human; Knock-out; Knockout Mice; Knowledge; Lead; Mediating; Modeling; Modification; Molecular; Molecular Target; Mus; NuRD complex; Nuclear Proteins; O-GlcNAc transferase; Outcome; Outcome Study; Pathway interactions; Patients; Physiological; Play; Polycomb; Post-Translational Protein Processing; Promoter Regions; Proteins; Regulation; Relative (related person); Repression; Repressor Proteins; Research; Role; Serine; Sickle Cell Anemia; Site; Switch Genes; System; Testing; Thalassemia; Therapeutic; Threonine; Transcription Coactivator; Transcription Repressor/Corepressor; Transgenic Mice; Transgenic Organisms; Translations; Up-Regulation; base; biological adaptation to stress; chromatin immunoprecipitation; drug development; effective therapy; fetal; gain of function; gene repression; human GATA1 protein; in vivo; loss of function; mouse model; new therapeutic target; novel; null mutation; peptide O-linked N-acetylglucosamine-beta-N-acetylglucosaminidase; promoter; public health relevance; transcription factor

DESCRIPTION (provided by applicant): Understanding the molecular mechanisms underlying the human ?- to ¿-globin gene switch has long been recognized as important in the treatment of hemoglobinopathies such as sickle cell disease (SCD), Cooley's anemia and ¿-thalassemias, since a wealth of evidence demonstrates that increased fetal hemoglobin (HbF) significantly decreases the pathophysiology associated with these diseases in patients. Our goal in this study is to understand the role of post-translation modifications (PTMs) of transcription factors involved in ?-globin gene silencing during the adult definitive erythropoiesis. Our clinica goal is to identify new molecular targets based on the outcome of this study that can be modulated therapeutically for up-regulation of ?-globin synthesis to treat these diseases. We recently demonstrated that one mode of ?-globin silencing occurs at the GATA binding sites located at -566 or -567 relative to the A?-globin or G?-globin gene CAP sites, respectively, and is mediated through the DNA-binding moiety GATA-1 and its recruitment of co-repressor partners, FOG-1 and Mi2, a component of the NuRD chromatin remodeling complex. Post-translational modifications of transcription factors may play a critical role in regulating the formation of transcriptional repressor complexes or activator complexes, particularly when both types of complexes might utilize the same DNA-binding protein, as is the case for GATA-1 in erythroid cells. We propose that the O-GlcNAcylation (O-GlcNAc) PTM is involved in regulating ?-globin transcription by controlling assembly of the GATA-1-FOG-1-Mi2 (NuRD) transcriptional repressor complex upstream of the ?-globin genes. A single Specific Aim will test the hypothesis that O-GlcNAcylation of GATA-1-FOG-1-Mi2 (NuRD) transcription/chromatin remodeling factors regulates their participation in a multi-protein repressor complex. In Specific Aim 1a, we will investigate the regulation of the ?-globin genes by O-GlcNAcylation. Chromatin immunoprecipitation (ChIP) analyses will be utilized to examine the spatial and temporal arrangement of the O-GlcNAc cycling enzymes, O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA), the GATA-1-FOG-1-Mi2 axis of proteins at the A?- globin promoter and the O-GlcNAc status of the promoter itself in cell cultures following terminal differentiation or induction of globin gene expression, with or without OGT/OGA knockdown or OGA inhibition, or in ¿-YAC transgenic mice during fetal liver definitive erythropoiesis. In Specific Aim 1b, we will determine whether O-GlcNAc regulates ¿-like globin gene expression during erythropoietic development in vivo in erythroid-specific floxed OGT conditional knockout ¿-YAC mice, or in OGT or OGA enforced-expression ¿-YAC bi-transgenic (bigenic) mice. Our proposed studies represent a new avenue of research in the globin gene switching field. The knowledge we gain from these studies will reveal novel therapeutic targets for which highly-specific treatments may be developed to increase HbF for the treatment of these red blood cells diseases without the side-effects associated with broad-spectrum therapies.

描述（由申请人提供）：了解人类的分子机制？长期以来，人们一直认为-到-珠蛋白基因开关在治疗血红蛋白病(如镰状细胞病（SCD）、库利贫血（Cooley’s anemia）和-地中海贫血（-thalassemia）中发挥着重要作用，因为大量证据表明，胎儿血红蛋白（HbF）的增加显著降低了患者与这些疾病相关的病理生理学。我们在这项研究中的目标是了解转录因子的翻译后修饰（PTMs）的作用。-珠蛋白基因在成人终性红细胞生成过程中的沉默。我们的临床目标是根据这项研究的结果确定新的分子靶点，这些靶点可以通过治疗性调节来上调？-珠蛋白合成来治疗这些疾病我们最近展示了？-珠蛋白沉默发生在GATA结合位点-566或-567处。-球蛋白还是G？-珠蛋白基因CAP位点分别为，和