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• W3157519273 abstract "Abstract The tetracycline (Tet)-controlled inducible system is the most widely used reversible system for transgenic expression in mice. Previously, we generated a GnRH-CreTeR mouse model, using a first-generation Tet-inducible system to temporally induce expression of Cre recombinase in GnRH neurons. Recently, the Tet-inducible system has undergone several modifications to significantly reduce previous limitations that include leaky background expression and lower sensitivity to tetracycline induction. Therefore, we have developed a novel mouse model bearing a Tet-inducible kisspeptin-Cre allele (iKiss-Cre mouse) that will enable temporal control over the selective deletion of genes from Kiss1 neurons. This temporally controlled gene deletion will eliminate a longstanding technical limitation of conventional steroid receptor knockout models in which steroid regulation of the axis is confounded by steroid developmental and organizational effects in the reproductive axis. Two mouse lines were generated. The first line targets kisspeptin neurons with a third generation Tet-inducible reverse tetracycline transactivator (rtTA, Tet-On 3G) expressed under the control of the Kiss1 allele. Using CRISPR-Cas9 technology, we inserted a cassette containing an internal ribosome entry site (IRES) sequence followed by the rtTA downstream of the Kiss1 coding region as was previously done using Cre recombinase. Transcription of the recombinant Kiss1 allele yields a bicistronic messenger RNA, from which both kisspeptin and rtTA are independently translated. The second mouse line, TRE-Cre mice, was constructed to express Cre recombinase under control of the PTRE3G promoter. The PTRE3G promoter is bidirectional to allow simultaneous monitoring of Cre expression and a second florescent protein reporter (ZsGreen1). The Kiss1-rtTA only binds to and activates the PTRE3G promoter in the presence of doxycycline (an analog of tetracycline). Offspring from breeding of these mouse lines, iKiss-Cre, result in a system capable of generating Tet-induced expression of Cre recombinase in kisspeptin neurons. In order to document specificity and sensitivity of this system, we performed immunofluorescent staining and observed colocalization of kisspeptin with Cre recombinase in brain sections of iKiss-Cre mice only after doxycycline treatment. We will use this model to investigate negative feedback actions of E2 on the adultpulse generator after E2 exerts its organizational actions on maturing kisspeptin neurons. We expect that this mouse model will become a major tool used by the neuroendocrine community and serve as proof of principle for development of similar inducible knockout models employing the current generation of inducible methodologies." @default.
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• W3157519273 date "2021-05-01" @default.
• W3157519273 modified "2023-10-08" @default.
• W3157519273 title "Development of a Novel Tetracycline-Inducible Kiss1-Cre Mouse Line for Temporally Controlled Gene Deletion in Kisspeptin Neurons" @default.
• W3157519273 doi "https://doi.org/10.1210/jendso/bvab048.1090" @default.
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