title: Conditional and inducible transgene expression in endothelial and hematopoietic cells using Cre/loxPand tetracycline-off systems.
abstract: In the present study, the tetracycline-off and Cre/loxPsystems were combined to gain temporal and spatial control of transgene expression. Mice were generated that carried three transgenes: Tie2-tTA, tet-O-Cre and either the ZEG or ZAP reporter. Tie2-tTA directs expression of tetracycline-controlled transactivator (tTA) in endothelial and hematopoietic cells under the control of the Tie2 promoter. Tet-O-Cre produces Cre recombinase from a minimal promoter containing the tet-operator (tetO). ZEG or ZAP contains a strong promoter and aloxP-flanked stop sequence, followed by an enhanced green fluorescence protein (EGFP) or human placental alkaline phosphatase (hPLAP) reporter. In the presence of tetracycline, the tTA transactivator produced by Tie-2-tTA is disabled and Cre is not expressed. In the absence of tetracycline, the tTA binds tet-O-Cre to drive the expression of Cre, which recombines theloxPsites of the ZEG or ZAP transgene and results in reporter gene expression. In the present study, the expression of the ZEG or ZAP reporter genes in embryos and adult animals with and without tetracycline treatment was examined. In the presence of tetracycline, no reporter gene expression was observed. When tetracycline was withdrawn, Cre excision was activated and the reporter genes were detected in endothelial and hematopoietic cells. These results demonstrate that this system may be used to bypass embryonic lethality and access adult phenotypes.
authors.affiliation: Laboratory of Microvascular Medicine, Medical Research Center, Shandong Provincial Qianfoshan Hospital, Shandong University, Jinan, Shandong 250014, P.R. China ; Molecular and Cellular Biology Division, Sunnybrook Health Science Centre, Toronto, Ontario M4N 3M5, Canada ; Department of Medical Biophysics, University of Toronto, Toronto, Ontario M5G 1L7, Canada.
authors.forename: Ju
authors.initials: J
authors.lastname: Liu
authors.affiliation: Theodor-Kocher-Institute, University of Berne, Berne CH-3012, Switzerland.
authors.forename: Urban
authors.initials: U
authors.lastname: Deutsch
authors.affiliation: Molecular and Cellular Biology Division, Sunnybrook Health Science Centre, Toronto, Ontario M4N 3M5, Canada ; Department of Medical Biophysics, University of Toronto, Toronto, Ontario M5G 1L7, Canada.
authors.forename: Iris
authors.initials: I
authors.lastname: Fung
authors.affiliation: Molecular and Cellular Biology Division, Sunnybrook Health Science Centre, Toronto, Ontario M4N 3M5, Canada ; Department of Medical Biophysics, University of Toronto, Toronto, Ontario M5G 1L7, Canada.
authors.forename: Corrinne G
authors.initials: CG
authors.lastname: Lobe
keywords: Cre/loxP system; endothelial cells; hematopoietic cell; tetracycline; transgenic mice