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. 1992 Aug 11;20(15):4021–4025. doi: 10.1093/nar/20.15.4021

Intron 1 and the 5'-flanking region of the human thymidylate synthase gene as a regulatory determinant of growth-dependent expression.

A Takayanagi 1, S Kaneda 1, D Ayusawa 1, T Seno 1
PMCID: PMC334082  PMID: 1508686

Abstract

We have determined the regulatory regions responsible for the growth-dependent expression of the human thymidylate synthase (TS) gene, using a set of minigenes constructed from segments of the human TS gene and the cDNA clone. Each construct was introduced stably into a TS-negative mutant of rat fibroblast 3Y1 cells. By serum-restricted synchronization of the cloned transformant cells, we found that a minigene with the genomic 5'-flanking region and intron 1 without other introns were sufficient for the normal extent and pattern of S-phase specific expression at the levels of both mRNA and enzymatic activity. In contrast, a TS cDNA clone driven by an SV40-based expression vector showed constitutive expression. Insertion of intron 1 into the cDNA clone in the normal location, or replacement of the viral 5'-promoter region of the cDNA clone by the genomic 5'-flanking sequence converted the constitutive expression to the S-phase dependent one, but only partly, that is, coexistence of the two regions were required for the normal expression. Results obtained by nuclear run-on assay suggested that posttranscriptional controls are also involved in this regulation in consistent with our previous results with the bona fide human TS gene.

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Selected References

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