Controlled re-activation of epigenetically silenced Tet promoter-driven transgene expression by targeted demethylation.
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Authors
Gödecke, NataschaZha, Lisha
Spencer, Shawal
Behme, Sara
Riemer, Pamela
Rehli, Michael
Hauser, Hansjörg
Wirth, Dagmar
Issue Date
2017-09-19
Metadata
Show full item recordAbstract
Faithful expression of transgenes in cell cultures and mice is often challenged by locus dependent epigenetic silencing. We investigated silencing of Tet-controlled expression cassettes within the mouse ROSA26 locus. We observed pronounced DNA methylation of the Tet promoter concomitant with loss of expression in mES cells as well as in differentiated cells and transgenic animals. Strikingly, the ROSA26 promoter remains active and methylation free indicating that this silencing mechanism specifically affects the transgene, but does not spread to the host's chromosomal neighborhood. To reactivate Tet cassettes a synthetic fusion protein was constructed and expressed in silenced cells. This protein includes the enzymatic domains of ten eleven translocation methylcytosine dioxygenase 1 (TET-1) as well as the Tet repressor DNA binding domain. Expression of the synthetic fusion protein and Doxycycline treatment allowed targeted demethylation of the Tet promoter in the ROSA26 locus and in another genomic site, rescuing transgene expression in cells and transgenic mice. Thus, inducible, reversible and site-specific epigenetic modulation is a promising strategy for reactivation of silenced transgene expression, independent of the integration site.Citation
Controlled re-activation of epigenetically silenced Tet promoter-driven transgene expression by targeted demethylation. 2017, 45 (16):e147 Nucleic Acids Res.Affiliation
Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr.7, D38124 Braunschweig, Germany.Journal
Nucleic acids researchPubMed ID
28934472Type
ArticleLanguage
enISSN
1362-4962ae974a485f413a2113503eed53cd6c53
10.1093/nar/gkx601
Scopus Count
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- Creative Commons
Except where otherwise noted, this item's license is described as http://creativecommons.org/licenses/by-nc-sa/4.0/