Tight control of gene expression in mammalian cells by tetracycline-responsive promoters.
- 15 June 1992
- journal article
- Published by Proceedings of the National Academy of Sciences in Proceedings of the National Academy of Sciences of the United States of America
- Vol. 89 (12), 5547-5551
- https://doi.org/10.1073/pnas.89.12.5547
Abstract
Control elements of the tetracycline-resistance operon encoded in Tn10 of Escherichia coli have been utilized to establish a highly efficient regulatory system in mammalian cells. By fusing the tet repressor with the activating domain of virion protein 16 of herpes simplex virus, a tetracycline-controlled transactivator (tTA) was generated that is constitutively expressed in HeLa cells. This transactivator stimulates transcription from a minimal promoter sequence derived from the human cytomegalovirus promoter IE combined with tet operator sequences. Upon integration of a luciferase gene controlled by a tTA-dependent promoter into a tTA-producing HeLa cell line, high levels of luciferase expression were monitored. These activities are sensitive to tetracycline. Depending on the concentration of the antibiotic in the culture medium (0-1 microgram/ml), the luciferase activity can be regulated over up to five orders of magnitude. Thus, the system not only allows differential control of the activity of an individual gene in mammalian cells but also is suitable for creation of "on/off" situations for such genes in a reversible way.Keywords
This publication has 30 references indexed in Scilit:
- Modulation of firefly luciferase stability and impact on studies of gene regulationGene, 1991
- RNA Polymerase II Transcription Blocked by Escherichia coli Lac RepressorScience, 1990
- Negative effect of the transcriptional activator GAL4Nature, 1988
- Oestrogen and glucocorticoid responsive elements are closely related but distinctNature, 1987
- Lac repressor can regulate expression from a hybrid SV40 early promoter containing a lac operator in animal cellsCell, 1987
- Kinetic and equilibrium characterization of the Tet repressor-tetracycline complex by fluorescence measurementsJournal of Molecular Biology, 1986
- A very strong enhancer is located upstream of an immediate early gene of human cytomegalovirusCell, 1985
- Construction of a fusion gene that confers resistance against hygromycin B to mammalian cells in cultureExperimental Cell Research, 1985
- Regulation of metallothionein–thymidine kinase fusion plasmids injected into mouse eggsNature, 1982
- A Rapid and Sensitive Method for the Quantitation of Microgram Quantities of Protein Utilizing the Principle of Protein-Dye BindingAnalytical Biochemistry, 1976