In mammalian nuclei, a select number of tissue-specific gene loci exhibit broadly distributed patterns of histone modifications, such as histone hyperacetylation, that are normally associated with active gene promoters. Previously, we characterized such hyperacetylated domains within mammalian ␤-globin gene loci, and determined that within the murine locus, neither the ␤-globin locus control region nor the gene promoters were required for domain formation. Here, we identify a developmentally specific erythroid enhancer, hypersensitive site-embryonic 1 (HS-E1), located within the embryonic ␤-globin domain in mouse, which is homologous to a region located downstream of the human embryonic ⑀-globin gene. This sequence exhibits nuclease hypersensitivity in primitive erythroid cells and acts as an enhancer in gain-offunction assays. Deletion of HS-E1 from the endogenous murine ␤-globin locus results in significant decrease in the expression of the embryonic ␤-globin genes and loss of the domain-wide pattern of histone hyperacetylation. The data suggest that HS-E1 is an enhancer that is uniquely required for ␤-like globin expression in primitive erythroid cells, and that it defines a novel class of enhancer that works in part by domainwide modulation of chromatin structure. (Blood. 2011;117(19):5207-5214)

In mammals, the complex tissue- and developmental-specific expression of genes within the β-globin cluster is known to be subject to control by the gene promoters, by a locus control region (LCR) located upstream of the cluster, and by sequence elements located across the intergenic regions. Despite extensive investigation, however, the complement of sequences that is required for normal regulation of chromatin structure and gene expression within the cluster is not fully defined. To further elucidate regulation of the adult β-globin genes, we investigate the effects of two deletions engineered within the endogenous murine β-globin locus. First, we find that deletion of the β2-globin gene promoter, while eliminating β2-globin gene expression, results in no additional effects on chromatin structure or gene expression within the cluster. Notably, our observations are not consistent with competition among the β-globin genes for LCR activity. Second, we characterize a novel enhancer located 3′ of the β2-globin gene, but find that deletion of this sequence has no effect whatsoever on gene expression or chromatin structure. This observation highlights the difficulty in assigning function to enhancer sequences identified by the chromatin “landscape” or even by functional assays.