Human gamma-globin gene promoter element regulates human beta-globin gene developmental specificity.
The persistence of fetal hemoglobin in many patients with deletion type beta thalassemias and the expression patterns of human globin genes in transgenic mice suggest that gamma- to beta-globin gene switching results primarily from competition of gamma- and beta-globin genes for interaction with the beta-globin locus control region (LCR). To define regulatory sequences that are essential for the competitive advantage of the gamma gene at early developmental stages, stable transgenic mouse lines were produced with LCR gamma-beta constructs containing deletions of gamma 5'-flanking DNA. All constructs contained the full 22 kb LCR, a 4.1 kb beta-globin gene and a gamma-globin gene with 1348, 383, 202, 130, 72 or 52 bp of 5'-flanking sequence. Primer extension analysis of yolk sac, fetal liver and blood RNA from these lines demonstrated that a region between -202 and -130 of the human gamma-globin gene promoter was required to suppress beta-globin gene expression at early developmental stages. Four transcription factor binding sites within this region [GATA(p), Oct1, GATA(d) and CACCC] were mutated independently in LCR gamma-beta constructs and transgenic mouse lines were produced. Only the gamma CACCC box mutation resulted in high levels of beta-globin gene expression in early embryos. These results demonstrate that the CACCC box of the human gamma-globin gene plays a critical role in human beta-globin gene developmental specificity. The data also suggest that gamma CACCC box binding factors mediate LCR-gamma interactions which normally enhance gamma-globin and suppress beta-globin gene expression in fetal erythroid cells.
Ryan
T M
TM
Department of Biochemistry and Molecular Genetics, School of Medicine and Dentistry, University of Alabama at Birmingham, Birmingham, AL 35294, USA.
Sun
C W
CW
Ren
J
J
Townes
T M
TM
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D005914:Globins / Q000235:genetics*; D051863:Host Cell Factor C1; D006801:Humans; D008099:Liver / Q000196:embryology / Q000378:metabolism; D019901:Locus Control Region / Q000235:genetics; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D009154:Mutation; D050811:Octamer Transcription Factor-1; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D014157:Transcription Factors / Q000378:metabolism*; D019076:Transgenes / Q000235:genetics; D015017:Yolk Sac / Q000378:metabolism
D004268:DNA-Binding Proteins; C497692:HCFC1 protein, human; C497693:Hcfc1 protein, mouse; D051863:Host Cell Factor C1; D050811:Octamer Transcription Factor-1; C494165:POU2F1 protein, human; C494166:Pou2f1 protein, mouse; D012333:RNA, Messenger; D014157:Transcription Factors; D005914:Globins; D004247:DNA