title: Autoregulatory lentiviral vectors allow multiple cycles of doxycycline-inducible gene expression in human hematopoietic cells in vivo.
abstract: The efficient control of gene expression in vivo from lentiviral vectors remains technically challenging. To analyze inducible gene expression in a human setting, we generated 'human immune system' (HIS) mice by transplanting newborn BALB/c Rag2(-/-)IL-2Rgamma(c)(-/-) immunodeficient mice with human hematopoietic stem cells transduced with a doxycycline-inducible lentiviral vector. We compared several methods of doxycycline delivery to mice, and could accurately measure doxycycline in vivo using a new sensitive detection assay. Two different lentiviral vector designs with constitutive (TRECMV-V14) or autoregulatory (TREAuto-V14) expression of an optimized reverse tetracycline transactivator were used to transduce human hematopoietic stem cells. After transplantation into immunodeficient mice, we analyzed the expression of the green fluorescent protein (GFP) reporter gene in the human hematopoiesis-derived cells that develop and accumulate in the generated HIS mice. We show efficient inducible GFP expression in adult HIS mice containing TREAuto-V14-transduced human cells, whereas GFP expression is poor with the TRECMV-V14 vector. Multiple cycles of doxycycline exposure in the TREAuto-V14 group result in repeated cycles of GFP expression with no loss of intensity. These findings are of major interest for gene therapy and basic research settings that require inducible gene expression.
authors.affiliation: Laboratory of Experimental Virology, Department of Medical Microbiology, Center for Infection and Immunity Amsterdam (CINIMA), Academic Medical Center of the University of Amsterdam (AMC-UvA), Amsterdam, The Netherlands.
authors.forename: M
authors.initials: M
authors.lastname: Centlivre
authors.forename: X
authors.initials: X
authors.lastname: Zhou
authors.forename: S M
authors.initials: SM
authors.lastname: Pouw
authors.forename: K
authors.initials: K
authors.lastname: Weijer
authors.forename: W
authors.initials: W
authors.lastname: Kleibeuker
authors.forename: A T
authors.initials: AT
authors.lastname: Das
authors.forename: B
authors.initials: B
authors.lastname: Blom
authors.forename: J
authors.initials: J
authors.lastname: Seppen
authors.forename: B
authors.initials: B
authors.lastname: Berkhout
authors.forename: N
authors.initials: N
authors.lastname: Legrand
mesh_terms: D000818:Animals; D004318:Doxycycline / Q000378:metabolism / Q000494:pharmacology*; D005786:Gene Expression Regulation; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000378:metabolism; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells / Q000378:metabolism*; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D051379:Mice; D008807:Mice, Inbred BALB C; D008822:Mice, Transgenic
chemical_list: D049452:Green Fluorescent Proteins; D004318:Doxycycline