A minigene containing four discrete cis elements recapitulates GATA-1 gene expression in vivo.
BACKGROUND
The GATA-1haematopoietic enhancer (G1HE), located between 3.9 and 2.6 kb 5' to the haematopoietic first exon, is essential for GATA-1 gene transcription in erythroid cells. However, G1HE is not sufficient to confer tissue specificity on the GATA-1 gene in vivo, indicating that additional regulatory sequences are necessary.


RESULTS
We demonstrate here that two other upstream promoter elements containing a double GATA motif or two CACCC boxes are also indispensable for reporter gene expression in erythroid cells in the transgenic mouse. The combination of these three cis-acting regions was sufficient for reporter expression in primitive erythroid cells, as demonstrated by linking the elements together into a 659 bp artificial (GdC) minigene. The minigene activated the transcription of a reporter gene from either the endogenous or an exogenous thymidine kinase promoter, retaining cell type-specificity. The addition of a 320 bp fragment in the first intron to the GdC minigene sustained reporter expression in the definitive stage. Moreover, a line of transgenic mouse that expressed GATA-1 cDNA under the control of the complete 979 bp minigene rescued GATA-1 germ line mutant mice from embryonic lethality.


CONCLUSIONS
A combination of four distinct sequence motifs co-operatively serve as a fundamental functional unit for GATA-1 erythroid transcription in vivo.
Ohneda
Kinuko
K
Center for Tsukuba Advanced Research Alliance and Institute of Basic Medical Sciences, University of Tsukuba, Japan.
Shimizu
Ritsuko
R
Nishimura
Shigeko
S
Muraosa
Yasushi
Y
Takahashi
Satoru
S
Engel
James Douglas
JD
Yamamoto
Masayuki
M
D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004622:Embryo, Mammalian / Q000033:anatomy & histology / Q000502:physiology; D015672:Erythroid Precursor Cells / Q000166:cytology / Q000378:metabolism; D050981:Erythroid-Specific DNA-Binding Factors; D050982:GATA1 Transcription Factor; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D006410:Hematopoiesis / Q000502:physiology; D006801:Humans; D007438:Introns; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
D004268:DNA-Binding Proteins; D050981:Erythroid-Specific DNA-Binding Factors; D050982:GATA1 Transcription Factor; C494688:GATA1 protein, human; C494689:Gata1 protein, mouse; D009687:Nuclear Proteins; D012097:Repressor Proteins; D014157:Transcription Factors