Ontogenic changes in erythroid gene expression

描述(申请人提供)：在胎儿时期，红细胞生成是主要的造血作用，以适应胎儿血量的快速扩张。已知一些发育差异可以区分胎儿和成人的红细胞生成，例如红系祖细胞的珠蛋白表达和对促红细胞生成素的反应性的变化。我们假设一些未知基因在胎儿和成人红系前体之间有不同的调控，以适应胎儿和成人红细胞的不同功能。微阵列技术是比较细胞群体间基因表达的有力工具。为了更好地应用这项技术来破译胎儿和成人红细胞生成过程中的基因表达，我们已经着手改进用于分离不同成熟阶段的红系前体的工具。利用噬菌体展示技术研制了一组识别未成熟有核红细胞的单链可变区抗体(ScFv Abs)。我们的目标是确定这些新试剂识别的细胞表面标记的分子同一性和表达模式。在红细胞生成过程中，至少有三个细胞表面标志物似乎被具有独特但重叠表达的单链抗体识别。此外，为了鉴定单链抗体识别的基因产物，我们建议对这些细胞表面分子在胎儿和成人红系发育过程中的表达进行表征。这项建议的第二个目的是描述胎儿和成人红系发育不同阶段的基因表达。这将通过从胎儿肝脏和成人骨髓中分离不同分化阶段的红系前体细胞，然后使用微阵列技术分析基因表达来完成。这些实验将描述红系发育过程中基因的表达，并比较成人和胎儿红细胞的表达。对红系祖细胞生长和成熟过程中基因表达的详细了解可能有助于改进血红蛋白疾病的基因治疗方法。

DESCRIPTION (provided by applicant): Erythropoiesis dominates hematopoiesis during fetal life to accommodate the rapid expansion in fetal blood volume. Some developmental differences are known to distinguish fetal from adult erythropoiesis such as changes in globin expression and responsiveness to erythropoietin by erythroid progenitors. We hypothesize that a number of unknown genes are differentially regulated between fetal and adult erythroid precursors to accommodate the distinct functions of fetal and adult erythrocytes. Microarray technology is a powerful tool used to compare gene expression between cell populations. To best apply this technology to decipher gene expression during fetal and adult erythropoiesis we have embarked on an effort to improve the tools used to isolate erythroid precursors at different stages of maturation. A panel of single-chain variable fragment antibodies (scFv Abs) was developed, using phage display technology, which recognizes immature nucleated erythrocytes. We aim to determine the molecular identity and expression pattern of the cell-surface markers recognized by these novel reagents. At least three cell surface markers appear to be recognized by the scFv Abs with unique but overlapping expression during erythropoiesis. In addition, to identify the gene products recognized by the scFv Abs we propose to characterize the expression of these cell surface molecules during fetal and adult erythroid development. The second aim of this proposal is to profile gene expression at different stages of fetal and adult erythroid development. This will be accomplished by isolating erythroid precursors at different stages of differentiation from fetal liver and adult bone marrow and then analyzing gene expression using microarray technology. These experiments will both profile the expression of genes during erythroid development and compare expression between adult and fetal erythrocytes. A detailed understanding of gene expression that occurs during the growth and maturation of erythroid progenitors is likely to contribute to improved methods of genetic therapy for hemoglobinopathies.