The major aim of this proposal is to experimentally define the cis-acting DNA sequences responsible for the tissue-specific and stage-specific expression of the beta-globin gene family, using the transgenic mouse as an assay system. The globin genes represent an interesting system for the study of developmental gene regulation, and the control of their expression is an issue of medical significance because of the many hereditary defects that affect hemoglobin synthesis in man. While the globin genes are one of the best characterized mammalian gene families, very little is known about their developmental regulation. We have recently demonstrated that a cloned adult beta-globin gene introduced into the mouse germ line is subject to tissue-specific and stage-specific regulation. The transgenic mouse therefore appears to be a powerful experimental system for studying the molecular control of hemoglobin switching. Our approach will be to microinject cloned human and mouse globin genes into the mouse zygote, and to analyze the expression of the introduced genes, at the RNA and protein levels, in the resulting transgenic mice and their progeny. We will first introduce individual embryonic, fetal and adult globin genes, with varying amounts of flanking DNA, to analyze the DNA sequences required for efficient and tissue-specific expression. We will also construct and test the expression of hybrid genes, incorporating portions of two different globin genes, to identify the DNA sequences that confer distinct patterns of developmental regulation (i.e., the sequences that control hemoglobin switching). We will produce transgenic mice carrying gamma globin genes from patients with Hereditary Persistence of Fetal Hemoglobin, to test how specific mutations in these genes affect their developmental regulation. Finally, we will attempt to """"""""cure"""""""" a mutant mouse strain that is a model of human beta thalassemia, by introducing the missing beta-globin gene.
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