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An adaptable system for improving transposon‐based gene expression in vivo via transient transgene repression
Author(s) -
Doherty Joseph E.,
Woodard Lauren E.,
Bear Adham S.,
Foster Aaron E.,
Wilson Matthew H.
Publication year - 2013
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fj.13-232090
Subject(s) - transgene , luciferase , psychological repression , biology , repressor , microbiology and biotechnology , reporter gene , gene expression , regulation of gene expression , in vivo , transposable element , gene , transfection , genetics , mutant
Transposons permit permanent cellular genome engineering in vivo. However, transgene expression falls rapidly postdelivery due to a variety of mechanisms, including immune responses. We hypothesized that delaying initial transgene expression would improve long‐term transgene expression by using an engineered piggyBac transposon system that can regulate expression. We found that a 2‐part nonviral Tet‐KRAB inducible expression system repressed expression of a luciferase reporter in vitro. However, we also observed nonspecific promoter‐independent repression. Thus, to achieve temporary transgene repression after gene delivery in vivo , we utilized a nonintegrating version of the repressor plasmid while the gene of interest was delivered in an integrating piggyBac transposon vector. When we delivered the luciferase transposon and repressor to immunocompetent mice by hydrodynamic injection, initial luciferase expression was repressed by 2 orders of magnitude. When luciferase expression was followed long term in vivo , we found that expression was increased > 200‐fold compared to mice that received only the luciferase transposon and piggyBac transposase. We found that repression of early transgene expression could prevent the priming of luciferase‐specific T cells in vivo. Therefore, transient transgene repression postgene delivery is an effective strategy for inhibiting the antitransgene immune response and improving long‐term expression in vivo without using immunosuppression.—Doherty, J. E., Woodard, L. E., Bear, A. S., Foster, A. E., Wilson, M. H., An adaptable system for improving transposon‐based gene expression in vivo via transient transgene repression. FASEB J. 27, 3753–3762 (2013). www.fasebj.org