Molecular Biology of Human Erythrocyte Alpha Spectrin

人红细胞α血影蛋白的分子生物学

• 批准号: 7918635
• 负责人: PATRICK G GALLAGHER
• 金额: $ 41.38万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-09-15 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7918635
• 关键词: Address; Affect; Anemia; Architecture; Base Sequence; Binding; Biochemical; Biological Assay; Blood; Blood Transfusion; Cells; Chromatin; Code; DNA; DNA-Protein Interaction; Data; Defect; Deoxyribonuclease I; Disease; Enhancers; Epigenetic Process; Erythrocyte Membrane; Erythrocytes; Erythroid; Erythroid Cells; Erythropoiesis; Gene Expression; Gene Expression Regulation; Gene Structure; Genes; Genomics; Goals; Hemolytic Anemia; Hereditary Elliptocytosis; Hereditary Spherocytosis; Histones; Human; Hybridization Array; In Vitro; Inherited; Insulator Elements; Knowledge; Link; Maps; Membrane; Membrane Protein Gene; Membrane Proteins; Messenger RNA; Molecular; Molecular Biology; Mutation; Nucleic Acid Regulatory Sequences; Operative Surgical Procedures; Pathogenesis; Patients; Positioning Attribute; Principal Investigator; Protein Binding; RNA Polymerase II; Regulation; Regulatory Element; Reporter Genes; Role; Sequence Analysis; Site; Spectrin; Structure; Techniques; Technology; Tissues; Transcript; Transgenic Mice; alpha Spectrin; chromatin immunoprecipitation; genetic regulatory protein; genome sequencing; in vivo; insight; membrane biogenesis; membrane skeleton; mutant; programs; protein expression; public health relevance; research study; transcription factor

DESCRIPTION (provided by applicant): The long-term goals of this proposal are to elucidate the molecular mechanisms involved in normal and abnormal expression of the protein 1-spectrin, a critical component of the erythrocyte membrane skeleton. Abnormalities of ?-spectrin are associated with inherited hemolytic anemia, which is sometimes severe. The first aim of this proposal is to identify and characterize the cis-sequences, trans-factors, and epigenetic state, including chromatin architecture, across the ?-spectrin gene locus that regulate its expression in erythroid and nonerythroid cells. These studies address the hypothesis that common regulatory signatures in ?-spectrin and other erythrocyte membrane protein genes control their tissue-specific expression. Integration of mRNA transcript composition, genomic organization, RNA polymerase II binding, transcription factor and regulatory protein binding, and histone architecture will provide detailed knowledge of erythrocyte membrane gene structure, function, and regulation and allow us to identify a common regulatory signature that controls expression in erythroid cells. The studies in this aim combine high throughput genomic technologies with functional studies of gene expression. Techniques to be utilized include chromatin immunoprecipitation experiments followed by array hybridization (ChIP-chip) or whole genome sequencing (ChIP-seq), high throughput DNase I hypersensitive site mapping, and functional studies of gene expression. Results obtained from these studies will allow study of the role of 1-spectrin in erythropoiesis, membrane biogenesis, and inherited erythrocyte disorders. The second aim of this proposal is the identification of mutations that perturb ?- spectrin spectrin gene regulation and/or expression in patients with spectrin-linked inherited hemolytic anemias and characterization of the effect of these mutations on ?-spectrin gene structure, function, and expression. These studies address the hypothesis that defects of ?-spectrin occur in regions of functional importance and their elucidation will provide important information about the structure, function, and regulation of the ?-spectrin gene in normal and mutant erythrocytes. Nucleotide sequence analysis of amplified patient genomic DNA will be performed to identify genetic defects in cases of inherited hemolytic anemia associated with qualitative and quantitative defects of ?-spectrin. Previously known and newly identified cis-regulatory elements in the ?- spectrin gene will be interrogated and characterized in functional studies of gene regulation. Together, results from these studies will provide important information on the structure, function, and regulation of spectrin in erythroid and nonerythroid cells and shed additional insight into the pathogenesis of spectrin-linked disorders of the erythrocyte. PUBLIC HEALTH RELEVANCE: Many people suffer from anemia, or low blood count, due to abnormalities in the membranes, or lining, of their red blood cells. Some people require surgery or blood transfusions to treat the anemia. This application studies the what, why, and how of abnormalities of the lining of the red blood cell that cause the anemia.

描述（由申请人提供）：该提案的长期目标是阐明蛋白1-谱蛋白的正常和异常表达的分子机制，蛋白质1-谱蛋白是红细胞膜骨架的关键成分。 ？ - 光谱蛋白的异常与遗传性溶血性贫血有关，有时很严重。该提案的第一个目的是识别和表征横跨？ - 谱蛋白基因基因座的顺式序列，跨因素和表观遗传状态，包括染色质结构，这些基因座调节其在红细胞和非果皮细胞中的表达。这些研究解决了以下假设： - 光谱蛋白和其他红细胞膜蛋白基因控制其组织特异性表达。 MRNA转录组成，基因组组织，RNA聚合酶II结合，转录因子和调节蛋白结合以及组蛋白结构的整合将提供有关红细胞膜基因结构，功能和调节的详细知识，并使我们能够识别控制成骨细胞中表达的常见调节性签名。该目标的研究结合了高通量基因组技术与基因表达的功能研究。要使用的技术包括染色质免疫沉淀实验，然后进行阵列杂交（CHIP-CHIP）或整个基因组测序（CHIP-SEQ），高吞噬DNase I超敏位点映射以及基因表达的功能研究。从这些研究中获得的结果将允许研究1-谱蛋白在红细胞生成，膜生物发生和遗传性红细胞疾病中的作用。该提案的第二个目的是鉴定出扰动的突变？ - 光谱谱蛋白基因调节和/或表达在具有谱线连接的遗传性溶血性贫血的患者中以及这些突变对？ - 谱蛋白基因结构，功能和表达的作用的表征。这些研究解决了以下假设： - 谱蛋白的缺陷发生在功能重要性及其阐明的区域中，将提供有关正常和突变性红细胞中的 - 谱蛋白基因的结构，功能和调节的重要信息。将进行放大患者基因组DNA的核苷酸序列分析，以鉴定与遗传性溶血性贫血的遗传缺陷，与质量和定量缺陷有关？-spectrin。在基因调节的功能研究中将质疑和表征谱系基因中的先前已知和新鉴定的顺式调节元件。总之，这些研究的结果将提供有关红细胞和非刺细胞中光谱蛋白的结构，功能和调节的重要信息，并提供了对红细胞谱相关疾病的发病机理的更多见解。 公共卫生相关性：许多人因膜或衬里的红细胞异常而遭受贫血或低血分的困扰。有些人需要手术或输血来治疗贫血。该应用研究了引起贫血的红细胞内壁的异常，为什么以及如何。