A GATA box in the GATA-1 gene hematopoietic enhancer is a critical elementin the network of GATA factors and sites that regulate this gene

A region located at kbp -3.9 to -2.6 5' to the first hematopoietic exon of the GATA-1 gene is necessary to recapitulate gene expression in both the pr imitive and definitive erythroid lineages. In transfection analyses, this r egion activated reporter gene expression from an artificial promoter in a p osition- and orientation-independent manner, indicating that the region fun ctions as the GATA-1 gene hematopoietic enhancer (G1HE), However, when anal yzed in transgenic embryos in viveo, G1HE activity was orientation dependen t and also required the presence of the endogenous GATA-1 gene hematopoieti c promoter. To define the boundaries of G1HE, a series of deletion construc ts were prepared and tested in transfection and transgenic mice analyses, W e show that G1HE contains a 149-bp core region which is critical for GATA-1 gene expression in both primitive and definitive erythroid cells but that expression in megakaryocytes requires the core plus additional sequences fr om G1HE. This core region contains one GATA, one GAT, and two E boxes. Muta tional analyses revealed that only the GATA box is critical for gene-regula tory activity. Importantly, G1HE was active in SCL-/- embryos, These result s thus demonstrate the presence of a critical network of GATA factors and G ATA binding sites that controls the expression of this gene.