The molecular pathophysiology of the congenital dyserythropoietic anemias

项目摘要 由于红细胞生成（红细胞[RBC]生成）缺陷导致的贫血是死亡的主要原因， 世界范围内的发病率。先天性红细胞生成不良性贫血（CDAs）是一组终末期疾病， 红系成熟缺陷的特点是无效的红细胞生成和独特的骨髓（BM）发现- ings。CDAII是最常见的CDA亚型，其次是CDAI。CDAII是一种常染色体隐性遗传病 由编码外壳蛋白复合物-2核心组分的SEC 23 B的功能缺失突变引起 （COPII）囊泡，其将分泌蛋白从内质网（ER）运输到高尔基体的附件- ratus。CDAI也是一种常染色体隐性遗传病，由CDAN 1的功能缺失突变引起，编码CDAN 1基因。 CODANIN 1，被认为在染色质组装中起作用。尽管鉴定出了 由于CDAI和CDAI II的潜在缺陷，这些疾病的病理生理学仍然未知。CODANIN 1具有 已显示与SEC 23 B共定位于细胞内，表明CDAI和CDAI II可能共享一个com-located位点。 单或相关的分子发病机制。我们先前产生了造血SEC 23 B缺陷的小鼠; 这些小鼠没有表现出RBC缺陷。我们接下来通过对酵母、斑马鱼、小鼠和人类的研究表明， 在人细胞中，SEC 23 B与其密切相关的蛋白质SEC 23 A在功能上重叠，并且其表达与人细胞中的表达水平相关。 SEC 23旁系同源物的模式在进化中发生了变化（SEC 23 B是主要表达的旁系同源物， 人BM，在小鼠BM中具有相当的SEC 23 A和SEC 23 B水平）。本提案的总体目标是 是为了描述CDA的分子发病机制。在早期的初步结果中，i）我们产生了小鼠， 具有红系特异性组合Sec 23 a/Sec 23 b缺失;这些小鼠表现出显著的红系表型; ii）我们还产生了造血细胞Cdan 1缺失的小鼠，这表明了严重的血液学退化。 完美。在这个建议中，我们的目标是表征CODANIN 1和SEC 23在红细胞生成中的关键作用， 定义由Cdan 1或Sec 23 a/Sec 23 b组合缺失引起的造血缺陷的程度。我们 本研究还旨在确定Sec 23 b和Cdan 1在体内的遗传相互作用，并确定细胞内 SEC 23 B缺陷型红系细胞与野生型红系细胞中CODANIN 1的运输。此外，根据ER- 在某些酸化的人血清中，CDAII中的至高尔基体转运缺陷和CDAII RBC的溶解，这表明 红细胞膜异常，我们假设CDAII是由一种或多种 关键货物蛋白质的RBC质膜。在初步结果中，我们证明了显着的消耗 与对照RBC质膜相比，CDAII中只有5种蛋白质。我们将通过定义来扩展我们的研究- 在额外的CDAII患者样本中检测这些货物的分泌，我们将确定这些货物的作用， GOS在CDAII的病理生理学中的作用我们提出的研究对于理解 CDAs的病理生理学，并有望为开发新的治疗方法奠定基础， 这些疾病可能与由缺陷性终末红细胞成熟引起的其他贫血有关。

Project Summary Anemia due to defects in erythropoiesis (red blood cell [RBC] production) is a major source of mortality and morbidity worldwide. The Congenital Dyserythropoietic Anemias (CDAs) are a group of disorders of terminal erythroid maturation defects characterized by ineffective erythropoiesis and distinctive bone marrow (BM) find- ings. CDAII is the most common CDA subtype, followed by CDAI. CDAII is an autosomal recessive disease resulting from loss-of-function mutations in SEC23B, encoding a core component of coat protein complex-2 (COPII) vesicles, which transport secretory proteins from the Endoplasmic reticulum (ER) to the Golgi appa- ratus. CDAI, also an autosomal recessive disease, results from loss-of-function mutations in CDAN1, encoding CODANIN1, which is proposed to play a role in chromatin assembly. Despite the identification of the genetic defects underlying CDAI and CDAII, the pathophysiology of these disorders remains unknown. CODANIN1 has been shown to co-localize intracellularly with SEC23B, suggesting that CDAI and CDAII might share a com- mon or related molecular pathogenesis. We previously generated mice with hematopoietic SEC23B deficiency; these mice did not exhibit a RBC defect. We next showed through studies in yeast, zebrafish, mice, and hu- man cells that SEC23B overlaps in function with its closely related paralog, SEC23A, and that the expression pattern of the SEC23 paralogs has shifted in evolution (SEC23B is the predominantly expressed paralog in human BM, with comparable SEC23A and SEC23B levels in mouse BM). The overall objective of this proposal is to characterize the molecular pathogenesis of the CDAs. In early preliminary results, i) we generated mice with erythroid-specific combined Sec23a/Sec23b deletion; these mice exhibit a profound erythroid phenotype; ii) we also generated mice with hematopoietic Cdan1 deletion, which demonstrate a profound hematologic de- fect. In this proposal, we aim to characterize the critical roles of CODANIN1 and SEC23 in erythropoiesis and define the extent of the hematopoietic defect resulting from Cdan1 or combined Sec23a/Sec23b deletion. We also aim to define the genetic interaction between Sec23b and Cdan1 in vivo and determine the intracellular trafficking of CODANIN1 in SEC23B-deficient versus wild-type erythroid cells. Furthermore, based on the ER- to-Golgi transport defect in CDAII and on the lysis of CDAII RBC in some acidified human sera, which suggests an RBC membrane abnormality, we hypothesize that CDAII results from impaired trafficking of one or more key cargo proteins to the RBC plasma membrane. In preliminary results we demonstrate significant depletion of only 5 proteins in CDAII compared to control RBC plasma membranes. We will expand our studies by defin- ing the secretion of these cargos in additional CDAII patient samples and we will identify the roles of these car- gos in the pathophysiology of CDAII. Our proposed studies have important implications for understanding the pathophysiology of the CDAs and are expected to lay the foundation for the development of novel therapies for these disorders, which may be relevant to other anemias resulting from defective terminal erythroid maturation.