Genetic Heterogeneity and Protein Function in DBA

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Diamond Blackfan anemia (DBA) is a congenital anemia that develops at birth or soon after, and is due to failure of production of erythrocytes and their precursors, with normal or near normal myeloid and platelet lineages. Patients can emit completely on corticosteroids or may become resistant to treatment, and then require regular blood transfusions, or bone marrow transplant if a histocompatible sibling donor is available. DBA patients are at increased risk of developing leukemia and other malignancies. DBA is inherited in about 10-15 percent of cases, mostly as an autosomal dominant. Recent genetic studies have led to the the surprising identification of mutations in a ribosomal protein gene, RPS19, on chromosome 19q13.2, in about 25 percent of both familial and sporadic cases (DBA1). Linkage analysis in multiplex DBA families shows strong evidence for another gene on chromosome 8p (DBA2) in about 40 percent of families, and other pedigrees do not show evidence for linkage to either chromosome 8p or 19q, indicating further genetic heterogeneity. The long term objective of this proposal is to identify and isolate the DBA2 gene. Therefore the specific aims are to (1), further define the chromosome 8p genetic map by ascertaining more families to search by linkage and haplotype analysis for recombinations in the flanking regions, and screen by cytogenetic techniques for deletions and translocations; (2) use cDNA arrays of the genes and expressed sequence tags (ESTs) in the critical region to define candidate genes by examining their pattern of expressed RNAs in erythroid cells, and by comparing the hybridization of normal and patient genomic DNA to these arrays to look for loss of heterozygosity; and (3), test candidate genes identified either as a result of progress in aim 1 and/or aim 2 for mutations in chromosome 8 linked families by SSCP, PCR heterozygosity screening and sequence analysis. Knowledge of additional genes that cause DBA may offer new insights into the molecular regulation of erythropoiesis and the process of stem cell commitment to the erythroid lineage, and it is possible that the protein encoded by the gene on chromosome 8p interacts with RPS19 in a novel pathway. Furthermore, the increased risk of malignancy in these patients suggests that the protein may act as a tumor suppressor. Thus isolating the genes that cause DBA may be important not only for devising new treatment for these patients but also for a better understanding of the regulation of erythropoiesis and the development of malignancy.