A deletion of the human beta-globin locus activation region causes a major alteration in chromatin structure and replication across the entire beta-globin locus.

Abstract

Naturally occurring deletions that remove sequences located -60 kb upstream of the human adult {3-globin gene result in the failure to transcriptionally activate the cis-linked globin genes in erythroid cells. In addition, transfection, transgenic, and somatic cell hybrid studies have revealed that sequences within this region are essential for the developmentally regulated high-level expression of c/s-linked globin genes. This regulatory region located at the 5' end of the {3-globin locus has been termed the locus activation region (LAR). Using somatic cell hybrids, we have studied the chromatin structure and timing of DNA replication of the normal human I~-globin locus and a locus containing a de novo 25-kb deletion that removes elements of the LAR. As a result of this deletion, the entire 13-globin locus and sequences -100 kb 5' and 3' of the adult 13-globin gene are DNase I-resistant and do not form characteristic distant hypersensitive sites. These sequences also replicate late in S phase in an erythroid cell background. In contrast, the sequences of the normal locus are DNase I sensitive and early replicating. These results suggest that the LAR is required for both the erythroid-specific chromatin structure and timing of DNA replication over a large physical distance.

Heterogeneous nuclear ribonucleoprotein C1/C2, MeCP1, and SWI/SNF form a chromatin remodeling complex at the β-globin locus control region

“…Mutation of this GC dinucleotide disturbs the binding of the p45-p18 NF-E2 protein dimer (25) and drastically reduces the enhancer activity of the HS2 (11). In an in vitro gel shift assay, MARE sequence interacts with the NF-E2 protein complex as well as several unidentified proteins (13,26). When we fractionated the K562 nuclear extracts on a heparin-agarose column, the p45 NF-E2 eluted with the 0.42 M NaCl (Fig.…”

“…Many deletion and transgenesis studies have been performed with the ␤-globin LCR (1,13) that suggest the importance of HS2 for transactivation of ␤-like globin genes and for the formation of DNase HS sites (7,(14)(15)(16). However, deletion of the LCR DNA sequences severely reduces the transcription of the ␤-like genes, but the DNase hypersensitivity and polymerase II association with promoters of the remaining ␤-globin genes persists (17)(18)(19).…”

The role of EKLF in human beta-globin gene competition.

“…Nuclei were isolated from 13.5-day frozen fetal livers as described by Forrester et al (1990). For each genotype (EKLF +/+, EKLF -/ ) 10 livers were disrupted with 10 strokes of a type B pestle.…”

Regulation of Globin Gene Expression in Erythroid Cells

“…Nonetheless, the in vivo relevance of both the /?-LCR and HS-40 is made clear by the occurrence of thalassemia patients with specific deletions in these regions which abolish expression of the downstream globin genes [lo, 111. Moreover, deletion of the P-LCR alters chromosomal properties over a large distance (> 200 kb), including a change in replication timing of the locus from early to late in S-phase [12]. Overall, it is believed that the LCR serves to decondense or 'open' the chromatin domain to permit expression of the downstream genes.…”