A Transgenic Mouse Model for Congenital Dyserythropoietic Anemia Type I

The Congenital Dyseyrthropoetic Anemias (CDA) are a heterogeneous group of inborn defects characterized by anemia of varying severity and morphological abnormalities of bone marrow erythroblasts. Studying their pathology has the potential to contribute to the understanding of the normal process of erythropoiesis. In CDA type I, the phenotype is largely limited to the red cell lineage with erythroblasts showing characteristic spongy nuclear chromatin on electron microscopy. Its genetic basis is due to missense mutations in the CDAN1 gene (chromosome 15q15), but the biological function of the highly conserved and non-redundant codanin-1 protein remains entirely unknown.

We have produced a mouse model for CDA type I by generating transgenic mice from an ES cell line carrying a gene-trap (splice acceptor/β-galactosidase gene/neomycin resistance gene) in intron 25 of the murine CDAN1 gene. Blood and bone marrow from CDAN1^gt/wt heterozygotes is morphologically and quantitatively indistinguishable from wild type animals. Northern blot analysis and RT-qPCR of codanin-1 RNA expression confirms a broad pattern of expression with the highest levels seen in erythroid tissue. Flow-cytometric analysis of β-galactosidase activity in erythroid cells from phenylhydrazine treated adult spleens shows the highest levels in the CD71^high/Ter119^low early erythroblasts, declining with further erythroid maturation. Furthermore, indirect immunofluorescence localizes the codanin-1/β-galactosidase fusion product in the nucleus of the CD71^high/Ter119^low early erythroblasts, particularly at the interface between euchromatin and heterochromatin. The presence of an in-frame β-gal in the gene trap insert, therefore, provides a method to analyze gene and protein expression from the CDAN1 promoter, and additional distribution data from heterozygous embryos and adults will be presented. Heterozygote crosses produced no homozygotes among liveborn progeny, suggesting embryonic lethality of this state. Analysis of embryos at different stages suggests development of homozygotes ceased at ~6.5d of gestation, prior to the onset of erythropoiesis.

These results from the first transgenic mouse model for CDA type I, highlight a non-erythroid role for codanin-1 in early embryonic development, in addition to its role in adult human erythropoiesis. The embryonic lethality of the mouse model is consistent with the absence of homozygote null mutations in the CDAN1 genes analyzed so far in human patients. Creation of mice with the CDA type I phenotype may have to await a knock-in transgenic containing a human mutation, which is currently in preparation. The model described here should be valuable for further studies of the intracellular localization of codanin-1 and the identification of any relevant protein binding partners.