The goals of this program project are to study the cellular and molecular events which control switching in the expression of hemoglobin genes and changes in the characteristics of erythroid cells during ontogeny and cell differentiation. We propose a multidisciplinary approach which will involve human population and family studies for detection of mutants, DNA sequencing of normal and mutant globin genomic regions, chromatin functional assays, and biochemical and molecular studies of various markers of erythroid cells and stem cells. These goals will be accomplished with the collaborative effort of investigators who have expertise in human genetics, hematology, molecular, cell, and developmental biology, and immunology and membrane biochemistry. Five projects and three core units are included. Project 1 aims to identify human mutants for molecular analysis and to isolate stem cells for analysis of their differentiation and studies of their developmental programs during ontogeny. Project 2 focuses on studies of human Lambda and Beta globin switching at the level of chromatin in normal cells and cells from mutants. Project 3 will undertake sequencing of human mutant globin DNAs and of normal globin genomic sites of regulatory importance to be revealed by the chromatin studies. Project 4 investigates the control of avian globin gene switching and the role of non-histone chromosomal proteins and specific chromosomal events occurring in erythroid stem cells and their progeny during red cell differentiation. Project 5 examines whether erythroid membrane glycoproteins and glycolipids change during ontogeny and differentiation and the phenotypic relationship and coordination of these changes in relation to the changes occurring in globin expression. Core unit B supports the projects by collection of required fresh tissues and core unit C, with production of the numbers of cultured erythroblasts required for biochemical studies. As a result of this coordinated effort we expect to obtain basic information on hemoglobin switching and its control. Understanding the control of hemoglobin switching during ontogeny has practical importance since it may lead to possibilities of in vivo modulation. This will have therapeutic implications in patients with globin disorders such as sickle cell anemia.
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