Genetic Heterogeneity and Protein Function in DBA

钻石黑扇贫血(DBA)是一种先天性贫血，在出生时或出生后不久就会发展起来，是由于红细胞及其前体的产生失败，具有正常或接近正常的髓系和血小板谱系。患者可能会完全排出皮质类固醇或对治疗产生抵抗力，然后需要定期输血，如果有组织相合的兄弟姐妹捐赠者，则需要骨髓移植。DBA患者患白血病和其他恶性肿瘤的风险增加。DBA在大约10%-15%的病例中是遗传的，大部分是常染色体显性遗传。最近的遗传学研究令人惊讶地在染色体19q13.2上发现了核糖体蛋白基因RPS19的突变，在大约25%的家族性和散发性病例(DBA1)中。多个DBA家系的连锁分析表明，在大约40%的家系中存在染色体8p上的另一个基因(DBA2)，而其他家系没有显示与8p或19q染色体连锁的证据，这表明进一步的遗传异质性。这项建议的长期目标是鉴定和分离DBA2基因。因此，其具体目的是(1)通过确定更多的家系，通过连锁和单倍型分析寻找侧翼区域的重组，并利用细胞遗传学技术筛选缺失和易位，进一步确定染色体8p遗传图谱；(2)利用关键区域的基因和表达序列标签(ESTs)，通过检查它们在红系细胞中表达的RNA的模式来确定候选基因，并通过比较正常和患者基因组DNA与这些阵列的杂交来寻找杂合性丢失；以及(3)通过SSCP、聚合酶链式反应杂合性筛选和序列分析，检测8号染色体连锁家系中目标1和/或目标2突变的候选基因。对导致DBA的其他基因的了解可能为红细胞生成的分子调控和干细胞对红系的承诺过程提供新的见解，并有可能由染色体8p上的基因编码的蛋白质以一种新的途径与RPS19相互作用。此外，这些患者的恶性风险增加表明，该蛋白可能是一种肿瘤抑制因子。因此，分离导致DBA的基因可能不仅对于为这些患者设计新的治疗方法，而且对于更好地了解红细胞生成的调节和恶性肿瘤的发展都是重要的。

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.