Erythroid expression of the human alpha-spectrin gene promoter is mediated by GATA-1- and NF-E2-binding proteins.
alpha-Spectrin is a highly expressed membrane protein critical for the flexibility and stability of the erythrocyte. Qualitative and quantitative defects of alpha-spectrin are present in the erythrocytes of many patients with abnormalities of red blood cell shape including hereditary spherocytosis and elliptocytosis. We wished to determine the regulatory elements that determine the erythroid-specific expression of the alpha-spectrin gene. We mapped the 5' end of the alpha-spectrin erythroid cDNA and cloned the 5' flanking genomic DNA containing the putative alpha-spectrin gene promoter. Using transfection of promoter/reporter plasmids in human tissue culture cell lines, in vitro DNase I footprinting analyses, and gel mobility shift assays, an alpha-spectrin gene erythroid promoter with binding sites for GATA-1- and NF-E2-related proteins was identified. Both binding sites were required for full promoter activity. In transgenic mice, a reporter gene directed by the alpha-spectrin promoter was expressed in yolk sac, fetal liver, and erythroid cells of bone marrow but not adult reticulocytes. No expression of the reporter gene was detected in nonerythroid tissues. We conclude that this alpha-spectrin gene promoter contains the sequences necessary for low level expression in erythroid progenitor cells.
INSERM U409, Association Claude Bernard, Universite Paris 7, Faculte X. Bichat, 75870 Paris Cedex 18, France.
Laurent
L
Boulanger
Denise E
DE
Sabatino
Ellice Y
EY
Wong
Amanda P
AP
Cline
Lisa J
LJ
Garrett
Michel
M
Garbarz
Didier
D
Dhermy
David M
DM
Bodine
Patrick G
PG
Gallagher
D024506:5' Flanking Region; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D018076:DNA, Complementary / Q000737:chemistry / Q000302:isolation & purification; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015672:Erythroid Precursor Cells / Q000378:metabolism*; D050981:Erythroid-Specific DNA-Binding Factors; D004920:Erythropoiesis; D050982:GATA1 Transcription Factor; D006367:HeLa Cells; D006801:Humans; D020014:K562 Cells; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D050983:NF-E2 Transcription Factor; D050984:NF-E2 Transcription Factor, p45 Subunit; D011401:Promoter Regions, Genetic; D013049:Spectrin / Q000235:genetics*; D014157:Transcription Factors / Q000378:metabolism*
D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D050981:Erythroid-Specific DNA-Binding Factors; D050982:GATA1 Transcription Factor; C494688:GATA1 protein, human; C494689:Gata1 protein, mouse; D050983:NF-E2 Transcription Factor; D050984:NF-E2 Transcription Factor, p45 Subunit; C494692:NFE2 protein, human; C494693:Nfe2 protein, mouse; D014157:Transcription Factors; D013049:Spectrin