Ontogenic changes in erythroid gene expression

描述（由申请人提供）：胎儿期造血以红细胞生成为主，以适应胎儿血容量的快速扩张。一些发育上的差异被认为是区分胎儿和成人红细胞生成的重要因素，如红细胞祖细胞对红细胞生成素的反应和球蛋白表达的变化。我们假设胎儿和成人红细胞前体之间存在一些未知基因的差异调节，以适应胎儿和成人红细胞的不同功能。微阵列技术是一种强大的工具，用于比较细胞群之间的基因表达。为了最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，我们已经开始努力改进用于分离不同成熟阶段红细胞前体的工具。利用噬菌体展示技术，开发了一组单链可变片段抗体（scFv Abs），该抗体可识别未成熟的有核红细胞。我们的目的是确定这些新试剂识别的细胞表面标记物的分子特性和表达模式。在红细胞生成过程中，至少有三种细胞表面标记物可以被单克隆抗体识别，它们具有独特但重叠的表达。此外，为了鉴定scFv抗体识别的基因产物，我们建议表征这些细胞表面分子在胎儿和成年红细胞发育过程中的表达。本研究的第二个目的是分析胎儿和成年红细胞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细胞前体，然后使用微阵列技术分析基因表达来实现。这些实验将描述红细胞发育过程中基因的表达，并比较成人红细胞和胎儿红细胞之间的表达。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.