STRUCTURE AND FUNCTION OF THE MURINE BETA-GLOBIN LOCUS-CONTROL REGION 5' HS-3

被引:54
作者
HUG, BA
MOON, AM
LEY, TJ
机构
[1] WASHINGTON UNIV,JEWISH HOSP,MED CTR,DEPT MED,DIV HEMATOL ONCOL,ST LOUIS,MO 63110
[2] WASHINGTON UNIV,JEWISH HOSP,MED CTR,DEPT GENET,ST LOUIS,MO 63110
关键词
D O I
10.1093/nar/20.21.5771
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
We previously identified the murine homologue of the human beta-globin Locus Control Region (LCR) 5' HS-2. The lambda clone containing murine 5' HS-2 extends approximately 12 kb upstream from this site; here, we report the sequence of this entire upstream region. The murine homologue of 5'HS-3 is located approximately 16.0 kb upstream from the mouse (epsilon)y-globin gene, but no region homologous to human 5' HS-4 was present in our clone. Using a reporter system consisting of a human gamma-globin promoter driving the neomycin phosphotransferase gene (gamma-neo), we tested murine LCR fragments extending from -21 to -9 kb (with respect to the epsilony-globin gene cap site) for activity in classical enhancer and integration site assays in K562 and MEL cells. 5'HS-2 behaved as a powerful enhancer and increased the number of productive integration events (as measured by a colony assay) in both K562 and MEL cells. 5' HS-3 had no activity in K562 cells or in transiently transfected MEL cells, but was nearly as active as 5' HS-2 in the MEL cell colony assay. Two additional tests confirmed the identification of murine 5' HS-3: first, a DNA fragment containing 5' HS-3 confers copy number-dependent, integration-site independent inducibility on a linked beta-globin gene in the MEL cell environment. Secondly, a strong DNAsel hypersensitive site maps to the location of the 5' HS-3 functional core in chromatin derived from MEL cells. Collectively, these data suggest that we have identified the murine homologue of human 5' HS-3, and that this site is functional when integrated into the chromatin of MEL cells but not K562 cells. 5' HS-3 may therefore contain information that contributes to the development-specific expression of the beta-like globin genes.
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页码:5771 / 5778
页数:8
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