-Hemoglobin stabilizing protein is an erythroid protein that binds and stabilizes -hemoglobin during normal erythropoiesis and in pathological states of -hemoglobin excess. -Hemoglobin stabilizing protein has been proposed as a candidate gene in some Heinz body hemolytic anemias and as a modifier gene in the -thalassemia syndromes. To gain additional insight into the molecular mechanisms controlling the erythroid-specific expression of the -hemoglobin stabilizing protein gene and provide the necessary tools for further genetic studies of these disorders, we have initiated identification and characterization of the regulatory elements controlling the human -hemoglobin stabilizing protein gene. We mapped the 5’ end of the a-hemoglobin stabilizing protein erythroid cDNA and cloned the 5’ flanking genomic DNA containing the putative AHSP gene promoter. In vitro studies using transfection of promoter/reporter plasmids in human tissue culture cell lines, DNase I foot printing analyses, and gel mobility shift assays, identified an a-hemoglobin stabilizing protein gene erythroid promoter with functionally important binding sites for GATA-1 and Oct-1-related proteins. In transgenic mice, a reporter gene directed by a minimal human -hemoglobin stabilizing protein promoter was expressed in bone marrow, spleen, and reticulocytes, but not in nonerythroid tissues. In vivo studies using chromatin immunoprecipitation assays demonstrated hyperacetylation of the promoter region and occupancy by GATA-1. The -hemoglobin stabilizing protein promoter is an excellent candidate region for mutations associated with decreased -hemoglobin stabilizing protein gene expression.