The tetracycline-responsive promoter contains functional interferon-inducible response elements.
Tetracycline (tet)-responsive expression vectors allow controlled inducible expression of proteins in mammalian cells. This system is widely used for experimental research both in vivo and in vitro. In our attempts to use this system to study the antiviral effect of IFNalpha on hepatitis B virus, we discovered an unexpected feature of the tet-responsive promoter (tet promoter) of the currently available expression vectors. IFNalphawas found to stimulate tet promoter activity after transient transfection in a dose- and cell type-dependent manner. By sequence inspection, an IFNalpha-stimulated response element (ISRE)-like sequence was identified in the linker regions located between the heptameric tet operator sequences. Gel shift assays revealed binding of IFN-stimulated gene factors to these sequences, indicating that they mediate the IFNalpha-mediated promoter stimulation. These data demonstrate an unexpected feature of the tet-responsive expression system which needs to be taken into account when using this system for analysis of cytokine functions in vitro and in vivo. The data also imply that the tet promoter-based expression system can be rendered non-responsive to IFNalpha by mutagenesis of the ISREs and this may be essential when considering gene therapy in vivo.
Rang
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A
Heinrich-Pette-Institut, Martinistrasse 52, D-20251 Hamburg, Germany.
Will
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H
D000900:Anti-Bacterial Agents / Q000378:metabolism* / Q000494:pharmacology; D001483:Base Sequence; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000235:genetics; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D050832:Interferon-Stimulated Gene Factor 3; D050834:Interferon-Stimulated Gene Factor 3, gamma Subunit; D016898:Interferon-alpha / Q000378:metabolism* / Q000494:pharmacology; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D013752:Tetracycline / Q000378:metabolism* / Q000494:pharmacology; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism
D000900:Anti-Bacterial Agents; D004268:DNA-Binding Proteins; C494227:IRF9 protein, human; D050832:Interferon-Stimulated Gene Factor 3; D050834:Interferon-Stimulated Gene Factor 3, gamma Subunit; D016898:Interferon-alpha; D014157:Transcription Factors; D013752:Tetracycline