The Evolution of the Mammalian SEC23 Paralogs and the Molecular Pathogenesis of Congenital Dyserythropoietic Anemia type II

哺乳动物 SEC23 旁系同源物的进化和先天性红细胞生成不良性贫血 II 型的分子发病机制

• 批准号: 9298412
• 负责人: Rami Khoriaty
• 金额: $ 16.96万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2020-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9298412
• 关键词: Abnormal Red Blood Cell; Accounting; Acute Erythroblastic Leukemia; Anatomy; Anemia; Animals; Biological Models; Biological Process; Bone Marrow; Bone Marrow Transplantation; Capsid Proteins; Cell Differentiation process; Cell membrane; Cell physiology; Cells; Characteristics; Complementary DNA; Complex; Congenital dyserythropoietic anemia; Data; Defect; Development; Disease; Endoplasmic Reticulum; Erythroblasts; Erythrocytes; Erythroid; Erythroid Cells; Evolution; Exhibits; Fertility; Financial compensation; Foundations; Functional disorder; Future; Genes; Genetic Engineering; Genetic Models; Genomics; Golgi Apparatus; Growth; Hematopoietic; Hematopoietic stem cells; Human; Impairment; K-562; Lead; Mass Spectrum Analysis; Molecular; Mus; Mutation; Organelles; Pancreas; Pathogenesis; Pathologic; Pathway interactions; Patients; Pattern; Perinatal; Phenotype; Production; Proteins; Proteome; Proteomics; Regulation; Regulatory Element; Role; Sampling; Secretory Cell; Techniques; Testing; Time; Tissues; Translating; Vesicle; Weaning; Work; base; improved; in vivo; insight; mouse model; new therapeutic target; novel; novel therapeutics; paralogous gene; protein complex; protein function; public health relevance; secretory protein; trafficking; type II Congenital dyserythropoietic anemia

 DESCRIPTION (provided by applicant): Congenital dyserythropoietic anemia type-II (CDAII) is an autosomal recessive disease resulting from a defect in terminal erythroid maturation, characterized by moderate anemia and increased bone marrow (BM) bi/multi- nucleated erythroblasts. CDAII results from mutations in SEC23B, one of two closely related mammalian SEC23 paralogs (SEC23A and SEC23B). SEC23 is a core component of coat protein complex-II (COPII) vesicles, which transport secretory protein cargos from the Endoplasmic Reticulum (ER) to the Golgi apparatus. BM transplantation cures CDAII, suggesting that the pathologic defect is restricted to the hematopoietic compartment. Despite the identification of the genetic defect underlying CDAII, the mechanism by which SEC23B deficiency results in CDAII remains unknown. We previously showed that, in contrast to humans, SEC23B-deficient mice die perinatally, exhibiting massive pancreatic degeneration. We also generated mice with SEC23B deficiency restricted to the hematopoietic compartment and showed that these mice lack anemia and other CDAII characteristics. In recent preliminary results, we genetically engineered the Sec23a cDNA into the genomic locus of Sec23b in mice. Mice expressing SEC23A under the normal temporal and tissue specific patterns of SEC23B appear healthy and survive past weaning. This surprising observation suggests that the two Sec23 paralogs are interchangeable at the level of protein function. Additional preliminary data demonstrate that the SEC23B/SEC23A expression ratio is higher in human BM compared to pancreas, with the reverse pattern observed in mice. Taken together, these data suggest that the tissue-specific expression patterns of the Sec23 paralogs have shifted during evolution between mouse and human and that the absence of CDAII in SEC23B-deficient mice is due to compensation from SEC23A, the latter accounting for a relatively high fraction of total SEC23 in the hematopoietic compartment of these animals (in contrast to humans). The overall objective of this proposal is to study the functional evolution of the two SEC23 paralogs and to characterize the molecular pathogenesis of CDAII. First, I will determine the extent to which the highly similar SEC23 paralogs functionally compensate for one another in-vivo. I will fully characterize SEC23B deficient mice expressing SEC23A under the endogenous Sec23b regulatory elements and assess for subtle abnormalities in pancreas function. If these mice are anatomically normal and exhibit normal survival, growth, and fertility, this will demonstrate that SEC23A and SEC23B are interchangeable at the protein level when expressed in the appropriate tissues and at the appropriate times and levels. Second, based on SEC23A and SEC23B expression data, and on our preliminary work demonstrating that the 2 SEC23 paralogs overlap in function, we hypothesize that mice with hematopoietic SEC23A (or combined SEC23A/SEC23B) deficiency will develop CDAII. We will generate and characterize these mice, and expect to generate a murine model faithfully reproducing the human CDAII phenotype. Third, based on the ER-to-Golgi transport defect in this disease, we hypothesize that CDAII results from the impaired trafficking of one or more key cargo proteins that depend on SEC23B for ER-exit, and therefore for secretion to the RBC plasma membrane. We will perform quantitative proteomic techniques to identify the critical cargo(s) that depend on SEC23B for exit from the ER in human erythroid cells. We will validate the putative cargo(s) for their roles in CDAII pathophysiology. These studies have important implications for understanding the evolutionary functions of the paralogous SEC23 genes, and for improving our fundamental understanding of the complex mechanisms by which cargos are sorted in COPII vesicles. The results from this project may lead to the identification of new therapeutic targets as well as model systems in which to test these targets. Cargos that depend on SEC23B for secretion during RBC development may represent novel targets for CDAII therapy, which are also expected to translate to other anemias due to defects of terminal erythroid maturation. We also expect to generate a mouse model faithfully reproducing human CDAII, which may be used in the future to test novel therapies. Determining the functional overlap of SEC23A and SEC23B is critical because a shared role of these paralogs would suggest that therapies that increase the expression of either paralog in erythroid cells might be effective in CDAII.

 描述（由适用提供）：先天性dyserythropoietic贫血-II（CDAII）是一种常染色体隐性疾病，是由于末端红细胞成熟的缺陷引起的，其特征是现代性贫血和增加的骨髓（BM）BI/多核红细胞。 CDAII是由Sec23b突变（两个密切相关的哺乳动物SEC23旁系同源物（SEC23A和SEC23B）之一的突变引起的。 Sec23是外套蛋白质复合物II（COPII）蔬菜的核心成分，它们将秘密蛋白肉类从内质网（ER）传递到高尔基体。 BM移植治疗CDAII，表明病理缺陷仅限于造血室。尽管鉴定出CDAII的遗传缺陷，但SEC23B缺乏导致CDAII的机制仍然未知。我们先前表明，与人类相比，Sec23b缺陷的小鼠围产期死亡，表现出巨大的胰腺变性。我们还产生了SEC23B缺乏症的小鼠，仅限于造血室，并表明这些小鼠缺乏贫血和其他CDAII特征。在最近的初步结果中，我们通常将SEC23A cDNA设计为小鼠Sec23b的基因组基因座。在正常的临时和组织特异性模式下表达Sec23a的小鼠在SEC23B的特定模式下看起来很健康，并且在断奶之前存活。这种令人惊讶的观察表明，在蛋白质功能水平上，两个SEC23旁系同源物是可以互换的。其他初步数据表明，与胰腺相比，人类BM的Sec23b/sec23a表达比更高，并且在小鼠中观察到反向模式。 Taken together, these data suggest that the tissue-specific expression patterns of the Sec23 paralogs have shifted during evolution Between mouse and human and that the absence of CDAII in SEC23B-deficiency mice is due to compensation from SEC23A, the latter accounting for a relatively high fraction of total SEC23 in the hematopoietic compartment of these animals (in contrast to humans).该提案的总体目的是研究两个SEC23旁系同源物的功能演化，并表征CDAII的分子发病机理。首先，我将确定高度相似的SEC23旁系同源物在功能上互相补偿的程度。我将充分表征在内源性SEC23B调节元素下表达SEC23A的SEC23B防御小鼠，并评估了胰腺功能微妙异常的评估。如果这些小鼠在解剖上正常且暴露的正常生存率，生长和生育能力，这将表明Sec23a和Sec23b在适当的组织以及适当的时间和适当的时间和水平时在蛋白质水平上可以互换。其次，基于SEC23A和SEC23B表达数据，以及我们的初步工作，证明了功能的2秒旁系同源物重叠，我们假设造血Sec23a（或组合的SEC23A/SEC23B）的小鼠将发展为CDAII。我们将产生和表征这些小鼠，并期望产生一个忠实地再现人类CDAII表型的鼠模型。第三，基于该疾病中的ER到高尔基运输缺陷，我们假设CDAII是由于依赖于Sec23b的一种或多种关键的货物蛋白的运输受损而导致的，这些货物属于ER-ER-EXET，因此是为了分泌RBC plasmambrane。我们将执行定量蛋白质组学技术，以识别依赖于Sec23b的关键货物，以从SEC23B出口到人类红细胞中的ER。我们将验证推定的货物在CDAII病理生理学中的作用。这些研究对理解寄生虫Sec23基因的进化功能具有重要意义，并改善了我们对在Copii蔬菜中对千粒分类的复杂机制的基本理解。该项目的结果可能导致鉴定新的治疗靶标以及用于测试这些目标的模型系统。依赖于SEC23B在RBC发育过程中分泌的嘉格斯可能代表CDAII治疗的新靶标，由于末端红细胞的成熟缺陷，这也有望转化为其他贫血。我们还期望生成一个忠实地再现人类CDAII的小鼠模型，将来可以使用该模型来测试新疗法。确定Sec23a和Sec23b的功能重叠至关重要，因为这些旁系同源物的共同作用表明，增加伴侣在红细胞细胞中任何一种旁系同生的疗法可能在CDAII中有效。