Transcriptional regulation of the rat platelet factor 4 gene: interaction between an enhancer/silencer domain and the GATA site.
We used various segments of the 5' upstream region of the rat platelet factor 4 (PF4) gene coupled to the human growth hormone gene and heterologous promoters to identify domains which are critical for tissue-specific expression. Transient expression experiments with rat bone marrow cells and other cell lines revealed a complex interplay between a core promoter domain from -97 to the transcriptional start site and an enhancer/silencer domain from -448 to -112. The core promoter contains a GATA site at -31 to -28 whose mutation to TATA or AATA decreases tissue specificity and moderately affects expression in megakaryocytes as well as a positively acting subdomain from -97 to -83 whose removal decreases overall transcription without affecting tissue specificity. The enhancer/silencer domain possesses three positively acting subdomains from -380 to -362, -270 to -257, and -137 to -120 as well as a negatively acting subdomain at -184 to -151 which is able to reduce overall transcription but has no effect on tissue specificity. The subdomain from -380 to -362 is most critical in restricting gene expression driven either by the PF4 promoter or by a heterologous promoter to the megakaryocytic lineage. The subdomains from -270 to -257 and -137 to -120 function together with the subdomain from -380 to -362 to somewhat increase tissue specificity. Simultaneous mutation of the GATA site and deletion of either the whole enhancer/silencer domain or the subdomain from -380 to -362 or -137 to -120 reduce transcription in megakaryocytes by 10- to 30-fold. On the basis of the above-described results, we propose that the megakaryocyte-specific enhancer/silencer domain and the GATA site are responsible for high-level expression of the PF4 gene in a lineage-specific manner.
Ravid
K
K
Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
Doi
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Beeler
D L
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Kuter
D J
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Rosenberg
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D000818:Animals; D001483:Base Sequence; D001853:Bone Marrow / Q000378:metabolism; D001854:Bone Marrow Cells; D002118:Calcium; D002478:Cells, Cultured; D002872:Chromosome Deletion; D004247:DNA; D004742:Enhancer Elements, Genetic; D005786:Gene Expression Regulation; D006801:Humans; D008969:Molecular Sequence Data; D009928:Organ Specificity / Q000235:genetics; D011401:Promoter Regions, Genetic; D051381:Rats; D012045:Regulatory Sequences, Nucleic Acid; D014158:Transcription, Genetic
D004247:DNA; D002118:Calcium