Cited by 104Virus-based gene therapy researchEpigenetics and DNA MethylationAnimal Genetics and Reproduction

Extensive Methylation of Promoter Sequences Silences Lentiviral Transgene Expression During Stem Cell Differentiation In Vivo

Open access
Herbst, Friederike, Ball, Claudia R, Tuorto, Francesca, Nowrouzi, Ali, Wang, Wei, Zavidij, Oksana, Dieter, Sebastian M, Fessler, Sylvia +6 more·2012·Molecular Therapy

Abstract

Lentiviral vectors (LV) are widely used to stably transfer genes into target cells investigating or treating gene functions. In addition, gene transfer into early murine embryos may be improved to efficiently generate transgenic mice. We applied lentiviral gene transfer to generate a mouse model transgenic for SET binding protein-1 (Setbp1) and enhanced green fluorescent protein (eGFP). Neither transgenic founders nor their vector-positive offspring transcribed or expressed the transgenes. Bisulfite sequencing of the internal spleen focus-forming virus (SFFV) promoter demonstrated extensive methylation of all analyzed CpGs in the transgenic mice. To analyze the impact of Setbp1 on epigenetic silencing, embryonic stem cells (ESC) were differentiated into cardiomyocytes (CM) in vitro. In contrast to human promoters in LV, virally derived promoter sequences were strongly methylated during differentiation, independent of the transgene. Moreover, the commonly used SFFV promoter (SFFVp) was highly methylated with remarkable strength and frequency during hematopoietic differentiation in vivo in LV but less in γ-retroviral (γ-RV) backbones. In summary, we conclude that LV using an internal SFFVp are not suitable to generate transgenic mice or perform constitutive expression studies in differentiating cells. Choosing the appropriate promoter is also crucial to allow stable transgene expression in clinical gene therapy.

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Integrity signals

Registry

Crossref · 10.1038/mt.2012.46

Cites

45 of 45 references verified0 citation errors · 0 not checkable

Statistics

Claims the paper makes

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • Several methods have been described to limit continuous expression of lentiviral transgenes, such as natural promoter silencing and self-limiting CRISPR-Cas genetic circuits.

    “Several methods have been described to limit continuous expression of lentiviral transgenes as natural promoter silencing and self-limiting CRISPR-Cas genetic circuits41,59,60.”

    From Allele specific repair of splicing mutations in cystic fibrosis through AsCas12a genome editing (2019) · cites this paper as [60]

    The Discussion (page not provided) reports that methylation of the lentiviral SFFV promoter silenced transgene expression, supporting natural promoter silencing as a way expression can cease. It does not describe that silencing as a deliberate method to limit expression or discuss self-limiting CRISPR-Cas circuits. (LLM, conf=0.86)

    Experimental — accuracy not yet measured

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

The self-check needs the paper’s full text.

Cites

Flagged first, then by citations

1 of the 46 references couldn’t be matched to a known paper.

Cited by

Sorted by citations

Authors
Herbst, F., Ball, C. R., Tuorto, F., Nowrouzi, A., Wang, W., Zavidij, O., Dieter, S. M., Fessler, S., van der Hoeven, F., Kloz, U., Lyko, F., Schmidt, M., von Kalle, C., & Glimm, H.
Affiliations
German Cancer Research Center (DE)
Corresponding author
Glimm, Hanno
Venue
Molecular Therapy
Publisher
Elsevier BV
Published
2012-05
Volume
20
Issue
5
Pages
1014-1021
ISSN
1525-0016 (ISSN-L)
OpenAlex
W2031887473
Cited by
104
References
46
Citations by year
2026: 6 · 2025: 4 · 2024: 12 · 2023: 8 · 2022: 9 · 2021: 13 · 2020: 9 · 2019: 5 · 2018: 4 · 2017: 7 · 2016: 8 · 2015: 6 · 2014: 4 · 2013: 6 · 2012: 2
FWCI
3.47
Type
journal article
Language
en
Open access
Locations
  • Molecular Therapy (publishedVersion, cc-by-nc-nd)
  • Europe PMC (PubMed Central) (publishedVersion)
  • PubMed Central (publishedVersion)
  • PubMed (publishedVersion)
Funders
  • Wellcome Trust
  • Bundesministerium für Bildung und Forschung
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Virus-based gene therapy research
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Epigenetics and DNA Methylation
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Animal Genetics and Reproduction
Keywords
lentiviral vectors, DNA methylation, promoter methylation, epigenetic silencing, transgene expression, SETBP1, embryonic stem cell differentiation, cardiomyocyte differentiation, hematopoietic differentiation, transgenic mice, gammaretroviral vectors, CpG methylation, gene therapy
MeSH terms
Animals, Carrier Proteins, Cell Differentiation, Genetic Vectors, Humans, Mice, Transgenic, Promoter Regions, Genetic, Spleen Focus-Forming Viruses, Stem Cells, Lentivirus, Sequence Analysis, DNA, Founder Effect, CpG Islands, Transgenes, DNA Methylation, Genes, Essential, Gene Silencing, Myocytes, Cardiac, Epigenesis, Genetic, Green Fluorescent Proteins, Mice

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

Are you an author, or do you see a problem with this page? Request a review