title: Use of the tetracycline-controlled transcriptional silencer (tTS) to eliminate transgene leak in inducible overexpression transgenic mice.
abstract: The doxycycline-inducible reverse tetracycline transactivator (rtTA) is frequently used to overexpress transgenes in a temporally regulated fashion in vivo. These systems are, however, often limited by the levels of transgene expression in the absence of dox administration. The tetracycline-controlled transcriptional silencer (tTS), a fusion protein containing the tet repressor and the KRAB-AB domain of the kid-1 transcriptional repressor, is inhibited by doxycycline. We hypothesized that tTS would tighten control of transgene expression in rtTA-based systems. To test this hypothesis we generated mice in which the CC10 promoter targeted tTS to the lung, bred these mice with CC10-rtTA-interleukin 13 (IL-13) mice in which IL-13 was overexpressed in an inducible lung-specific fashion, and compared the IL-13 production and phenotypes of parental mice and the triple transgenic CC10-rtTA/tTS-IL-13 progeny of these crosses. In the CC10-rtTA-IL-13 mice, IL-13, mucus metaplasia, inflammation, alveolar enlargement, and enhanced lung volumes were noted at base line and increased greatly after doxycycline administration. In the triple transgenic tTS animals, IL-13 and the IL-13-induced phenotype could not be appreciated without doxycycline. In contrast, tTS did not alter the induction of IL-13 or the generation of the IL-13 phenotype by doxycycline. Thus, tTS effectively eliminated the baseline leak without altering the inducibility of rtTA-regulated transgenes in vivo. Optimal "off/on" regulation of transgene expression can be accomplished with the combined use of tTS and rtTA.
authors.affiliation: Section of Pulmonary and Critical Care Medicine, Department of Internal Medicine, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06520-8057, USA.
authors.forename: Z
authors.initials: Z
authors.lastname: Zhu
authors.forename: B
authors.initials: B
authors.lastname: Ma
authors.forename: R J
authors.initials: RJ
authors.lastname: Homer
authors.forename: T
authors.initials: T
authors.lastname: Zheng
authors.forename: J A
authors.initials: JA
authors.lastname: Elias
mesh_terms: D000818:Animals; D004268:DNA-Binding Proteins / Q000235:genetics; D004318:Doxycycline / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects*; D018793:Interleukin-13 / Q000096:biosynthesis; D008168:Lung / Q000187:drug effects; D051379:Mice; D008822:Mice, Transgenic; D010641:Phenotype; D011994:Recombinant Proteins / Q000494:pharmacology; D012097:Repressor Proteins / Q000494:pharmacology*; D013752:Tetracycline; D014157:Transcription Factors; D015533:Transcriptional Activation / Q000187:drug effects; D019076:Transgenes
chemical_list: D004268:DNA-Binding Proteins; D018793:Interleukin-13; D011994:Recombinant Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; C485822:Zfp354a protein, mouse; C034811:tetracycline resistance-encoding transposon repressor protein; D013752:Tetracycline; D004318:Doxycycline