The beta‐like globin genes require the upstream locus control region (LCR) for proper expression. The active elements of the LCR coincide with strong erythroid‐specific DNase I‐hypersensitive sites (HSs). We have used 5′ HS4 as a model to study the formation of these HSs. Previously, we identified a 101 bp element that is required for the formation of this HS. This element binds six proteins in vitro. We now report a mutational analysis of the HS4 HS‐forming element (HSFE). This analysis indicates that binding sites for the hematopoietic transcription factors NF‐E2 and GATA‐1 are required for the formation of the characteristic chromatin structure of the HS following stable transfection into murine erythroleukemia cells. Similarly arranged NF‐E2 and GATA binding sites are present in the other HSs of the human LCR, as well as in the homologous mouse and goat sequences and the chicken beta‐globin enhancer. A combination of DNase I and micrococcal nuclease sensitivity assays indicates that the characteristic erythroid‐specific hypersensitivity of HS4 to DNase I is the result of tissue‐specific alterations in both nucleosome positioning and tertiary DNA structure.

No abstract

Proper expression of the genes of the human beta-globin gene locus requires the associated locus control region (LCR). Structurally, the LCR is defined by the presence of four domains of erythroid-specific chromatin structure. These domains, which have been characterized as DNase I hypersensitive sites (HSs), comprise the active elements of the LCR. The major focus of this research is to define the cis -acting elements which are required for the formation of these domains of unique chromatin structure. Our previous investigations on the formation of LCR HS4 demonstrated that NF-E2 and tandem, inverted GATA binding sites are required for the formation of the native HS. Similarly arranged NF-E2 and tandem GATA sites are present within the core regions of the other human LCR HSs and are evolutionarily conserved. Using site-directed mutagenesis of human HSs 2 and 3 we have tested the hypothesis that these NF-E2 and GATA sites are common requirements for the formation of all LCR HSs. We find that mutation of these elements, and particularly the GATA elements, results in a decrease or complete loss of DNase I hypersensitivity. These data imply the presence of common structural elements within the core of each LCR HS which are required for erythroid-specific chromatin structure reorganization.

No abstract