Characterization of the 5.5 kb promoter of human thromboxane synthase (TS) gene revealed a proximal positive regulatory sequence (PPRS, -90 to -25 bp) and several distal repressive elements. The maximal promoter activity was found to reside within the first 285 bp, ∼75% of which was contributed by the PPRS. The sequence between -365 and -665 bp exerted a strong repressive effect (∼55%) on reporter gene expression independent of orientation and position, consistent with properties expected for a silencer. The sequence upstream of -665 bp to -5.5 kb contains mainly repressive elements which further reduce the promoter activity by 30%. The 65 bp PPRS worked in an orientation-independent, but position-dependent, manner and could be further divided into two independent elements, PPRS1 (-90 to -50 bp) and PPRS2 (-50 to -25 bp). While similar nuclear factor(s) from different cell types interact with PPRS2, those interacting with PPRS1 exhibit cell specificity. Internal sequence deletion and oligonucleotide competition established that a binding sequence for NF-E2 in PPRS1 (-60 tgctgattcat -50) was important for enhancing TS promoter activity in HL-60 cells. The presence of NF-E2 mRNA in HL-60 cells was demonstrated by reverse-transcription PCR amplification of the cDNA and Northern blot analysis. A 9-fold transactivation of luciferase (luc) reporter gene expression had been detected when NF-E2 cDNA was co-expressed with a TS promoter/luc construct. Despite the fact that NF-E2 and the cis-elements could alter the efficiency of TS transcription, they were not sufficient for restricting cell-specific TS expression. Analysis of the methylation status at the TS promoter in several human cell lines reveals cell-specific patterns of methylation that might correlate with TS expression. Taken together, these results suggest that the expression of human TS gene is modulated by multiple factors including cis-elements, trans-activator(s), and possibly genomic methylation.
Research Article|November 01 1996
Multiple factors regulating the expression of human thromboxane synthase gene
Seung Joon BAEK;
*Division of Human Genetics, Department of Obstetrics and Gynecology, University of Maryland School of Medicine
†Medical Biotechnology Center, University of Maryland Biotechnology Institute, Baltimore, MD 21201, U.S.A.
‡To whom correspondence should be addressed at the Division of Human Genetics, University of Maryland School of Medicine, Baltimore, MD 21201, U.S.A.
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Kuan-Der LEE, Seung Joon BAEK, Rong-Fong SHEN; Multiple factors regulating the expression of human thromboxane synthase gene. Biochem J 1 November 1996; 319 (3): 783–791. doi: https://doi.org/10.1042/bj3190783
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