Deletion of the human β-globin LCR 5′HS4 or 5′HS1 differentially affects β-like globin gene expression in β-YAC transgenic mice

A 213 kb human β-globin locus yeast artificial chromosome (β-YAC) was modified by homologous recombination to delete 2.9 kb of cross-species conserved sequence similarity encompassing the LCR 5′ hypersensitive site (HS) 4 (Δ5′HS4 β-YAC). In three transgenic mouse lines, completion of the γ- to β-globin switch during definitive erythropoiesis was delayed relative to wild-type β-YAC mice. In addition, quantitative per-copy human β-like globin mRNA levels were similar to wild-type β-YAC transgenic lines, although β-globin gene expression was slightly decreased in the day 12 fetal liver of Δ5′HS4 β-YAC mice. A 0.8 kb 5′HS1 fragment was similarly deleted in the YAC. Three Δ5′HS1 β-YAC transgenic lines were established. ε-globin gene expression was markedly reduced, approximately 16 fold, during primitive erythropoiesis compared to wild-type β-YAC mice, but γ-globin expression levels were unaffected. However, during the fetal stage of definitive erythropoiesis, γ-globin gene expression was decreased approximately 4 fold at day 12 and approximately 5 fold at day 14. Temporal developmental expression profiles of the β-like globin genes were unaffected by deletion of 5′HS1. Decreased expression of the ε- and γ-globin genes is the first phenotype ascribed to a 5′HS1 mutation in the human β-globin locus, suggesting that this HS does indeed have a role in LCR function beyond simply a combined synergism with the other LCR HSs.