Novel Mechanisms of Congenital Dyserythropoietic Anemia

先天性红细胞生成不良性贫血的新机制

• 批准号: 9887377
• 负责人: PATRICK G GALLAGHER
• 金额: $ 41.84万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9887377
• 关键词: Anemia; Binding; Binding Proteins; Biological Assay; Bone Marrow; CRISPR/Cas technology; Cell Line; Cell model; Cells; Chromatin; Clustered Regularly Interspaced Short Palindromic Repeats; Congenital dyserythropoietic anemia; Data; Development; Disease; Erythrocytes; Erythroid; Erythroid Cells; Erythropoiesis; Fanconi' s Anemia; Fetal Liver; Functional disorder; Gene Expression; Gene Expression Regulation; Genes; Genetic Diseases; Genetic Models; Genetic study; Genomics; Hematology; Hematopoiesis; Hematopoietic; Hematopoietic stem cells; Hemoglobin; Hepatocyte; Homozygote; Human; Inherited; Knock-in; Knock-out; Knowledge; Light; Link; Modeling; Mus; Mutate; Mutation; Patients; Pediatrics; Phenotype; Point Mutation; Process; Production; Proteins; Rare Diseases; Regulation; Regulatory Element; Research; Research Personnel; Role; Small Interfering RNA; System; epigenomics; erythroid differentiation; experience; genetic analysis; genomic locus; in vivo Model; insight; knock-down; molecular hematology; mouse model; mutant; novel; overexpression; protein expression; therapeutic development; tool

Abstract The congenital dyserythropoietic anemias (CDAs) are a heterogeneous group of genetic diseases of red blood cell production marked by hyporegenerative anemia with the presence of multinucleated red blood cell precursors in the bone marrow. Numerous gene loci have been associated with CDA, the most common being codanin-1 (CDAI) and SEC23B (CDA II). Although a common cause of CDA worldwide, the function of codanin-1 is largely unknown. We have demonstrated that codanin-1 is a chromatin binding protein whose expression is maintained during erythroid differentiation. The objective of this application is to identify the function of CDA-associated proteins in erythropoiesis, with a particular focus on codanin-1 and a newly identified CDA protein, MACF1. This will be explored by examining three Specific Aims: (1) study of mechanisms of codanin-1 regulation of gene expression during erythropoiesis; (2) characterization of a murine CRISPR/Cas9 knockin model of a codanin-1 CDA- associated point mutations; and (3) characterization of the role of MACF1 in erythropoiesis and in the pathophysiology of CDA, including development of a knockin and knockout mutant MACF1-linked murine model of CDA. The central hypothesis is that codanin-1 and MACF1 are involved in regulation of erythroid development and differentiation. The experimental plan focuses on understanding this regulation using cell lines, human and murine primary erythroid cell systems, and mouse models. Preliminary data support the hypotheses and proposed studies. Diminished expression of codanin-1 in CDA I by siRNA and overexpression of a patient-derived point mutant codanin-1 both result in erythroid precursor multinuclearity and decreased hemoglobinization. Genetic analyses strongly support the role of MACF1 in CDA. The investigators have created numerous important tools for the proposed studies. The rationale for this proposal is that by understanding the pathophysiology of a rare disease like CDA, we will gain broad knowledge of mechanisms controlling erythropoiesis, resulting in insights applicable to development of therapeutic strategies for inherited and acquired disorders of red blood cell production. This project takes advantage of a wealth of expertise using the multiple PI format, joining 2 experienced hematologic researchers in Pediatrics at Yale. Dr. Gallagher has experience studying mechanisms of erythropoiesis and its perturbation in genetic disease as well as genomics of erythroid development and differentiation. Dr. Kupfer has studied fundamental aspects of hematopoiesis and its perturbation using Fanconi anemia as a model. Together these investigators are uniquely qualified to study diseases associated with dysfunctional erythropoiesis. The co-PIs are actively involved in molecular hematology research at Yale; both are co-PIs of the Yale Cooperative Center of Excellence in Hematology (YCCEH), providing support for some of the studies in this proposal. Together, these studies will shed important light on erythropoiesis and its perturbation in inherited and acquired disorders of erythrocyte production.

抽象的 先天性孕育性贫血（CDA）是红血的一组遗传疾病 具有多核红细胞的存在下的细胞产生，以低降解性贫血为标志 骨髓中的前体。许多基因基因座与CDA有关，最常见的是 Codanin-1（CDAI）和SEC23B（CDA II）。虽然是全球CDA的常见原因，但 codanin-1在很大程度上未知。我们已经证明，codanin-1是一种染色质结合蛋白 在红细胞分化过程中保持表达。此应用的目的是确定 CDA相关蛋白在红细胞生成中的功能，特别关注codanin-1和一个新的 鉴定出CDA蛋白，MACF1。这将通过检查三个具体目的来探讨：（1）研究 促红细胞生成期间基因表达的Codanin-1调节机制； （2）鼠的表征 CORANIN-1 CDA-相关点突变的CRISPR/CAS9敲击模型； （3）表征 MACF1在红细胞生成和CDA的病理生理学中的作用，包括开发敲击蛋白和 CDA的基因敲除突变体MACF1连接的鼠模型。中心假设是Codanin-1和MacF1 参与了红细胞发育和分化的调节。实验计划的重点 使用细胞系，人和鼠类原发性红细胞系统以及小鼠了解这种调节 型号。初步数据支持假设和拟议的研究。 codanin-1在 CDA I通过siRNA和患者来源的点突变体Codanin-1的过表达都导致红系 前体的多核性和降低血红蛋白。遗传分析强烈支持 CDA中的MACF1。研究人员为拟议的研究创建了许多重要的工具。这 该提议的理由是，通过了解CDA等罕见疾病的病理生理学，我们将获得 对控制红细胞生成的机制的广泛了解，从而产生适用于开发的见解 遗传和获得的红细胞产生疾病的治疗策略。这个项目需要 使用多个PI格式的丰富专业知识的优势，加入2位经验丰富的血液学研究人员 在耶鲁大学的儿科。 Gallagher博士具有研究促红细胞生成机制及其扰动的经验 在遗传疾病以及红细胞发育和分化的基因组学中。库普弗博士研究了 造血的基本方面及其使用Fanconi贫血作为模型的扰动。在一起 研究人员具有独特的资格研究与功能障碍性红细胞生成的疾病。 Co-Pis 在耶鲁大学积极参与分子血液学研究；两者都是耶鲁大学合作中心的共同点 血液学卓越（YCCEH），为本提案中的一些研究提供了支持。一起， 这些研究将对嗜红细胞毒素及其在遗传和获得性疾病中的扰动进行重要阐述 红细胞生产。