Novel Mechanisms of Congenital Dyserythropoietic Anemia

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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.