A minigene containing four discrete cis elements recapitulates GATA‐1 gene expression in vivo

K Ohneda, R Shimizu, S Nishimura, Y Muraosa… - Genes to …, 2002 - Wiley Online Library
K Ohneda, R Shimizu, S Nishimura, Y Muraosa, S Takahashi, JD Engel, M Yamamoto
Genes to Cells, 2002Wiley Online Library
Abstract Background: The G ATA‐1 h aematopoietic e nhancer (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 …
Abstract
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.
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