The transcriptional factor Bach 1 forms a heterodimer with small Maf family, and functions as a repressor of the Maf recognition element (MARE) in vivo. To investigate the involvement of Bach 1 in the heme-dependent regulation of the expression of the Of alpha-globin gene, human erythroleukemia K562 cells were cultured with succinylacetone (SA), a heme biosynthetic inhibitor, and the level of alpha-globin mRNA was examined. A decrease of of alpha-globin mRNA was observed in SA-treated cells, which was restored by the addition of hemin. The heme-dependent expression of alpha-globin occurred at the transcriptional level since the expression of human alpha-globin gene promoter reporter gene containing hypersensitive site-40 (HS-40) was decreased when K562 cells were cultured with SA. Hemin treatment restored the decrease of the promoter activity by SA. The regulation of the HS-40 activity by heme was dependent on the NF-E2/AP-1 (NA) site, which is similar to MARE. The NA site-binding activity of Bach 1 in K562 increased upon SA-treatment, and the increase was diminished by the addition of hemin. The transient expression of Bach 1 and mutated Bach 1 lacking CP motifs suppressed the HS-40 activity, and cancellation of the repressor activity by hemin was observed when wild-type Bach 1 was expressed. The expression of NF-E2 strengthened the restoration of the Bach 1-effect by hemin. Interestingly, nuclear localization of Bach 1 increased when cells were treated with SA, while hemin induced the nuclear export of Bach 1. These results indicated that heme plays an important role in the induction of alpha-globin gene expression through disrupting the interaction of Bach 1 and the NA site in HS-40 enhancer in erythroid cells. (C) 2004 Elsevier Inc. All rights reserved.