title: Cre recombinase induces DNA damage and tetraploidy in the absence of loxP sites.
abstract: The spatiotemporal manipulations of gene expression by the Cre recombinase (Cre) of bacteriophage P1 has become an essential asset to understanding mammalian genetics. Accumulating evidence suggests that Cre activity can, in addition to excising targeted loxP sites, induce cytotoxic effects, including abnormal cell cycle progression, genomic instability, and apoptosis, which can accelerate cancer progression. It is speculated that these defects are caused by Cre-induced DNA damage at off-target sites. Here we report the formation of tetraploid keratinocytes in the epidermis of keratin 5 and/or keratin 14 promoter-driven Cre (KRT5- and KRT14-Cre) expressing mouse skin. Biochemical analyses and flow cytometry demonstrated that Cre expression also induces DNA damage, genomic instability, and tetraploidy in HCT116 cells, and live-cell imaging revealed an extension of the G 2 cell cycle phase followed by defective or skipping of mitosis as cause for the tetraploidy. Since tetraploidy eventually leads to aneuploidy, a hallmark of cancer, our findings highlight the importance of distinguishing non-specific cytopathic effects from specific Cre/loxP-driven genetic manipulations when using Cre-mediated gene deletions.
authors.affiliation: Max-Planck-Institute of Biochemistry; Department of Molecular Medicine; Martinsried, Germany.
authors.forename: Vaibhao C
authors.initials: VC
authors.lastname: Janbandhu
authors.affiliation: Max-Planck-Institute of Biochemistry; Department of Molecular Medicine; Martinsried, Germany.
authors.forename: Daniel
authors.initials: D
authors.lastname: Moik
authors.affiliation: Max-Planck-Institute of Biochemistry; Department of Molecular Medicine; Martinsried, Germany.
authors.forename: Reinhard
authors.initials: R
authors.lastname: Fässler
mesh_terms: D000818:Animals; D017101:Bacteriophage P1 / Q000201:enzymology; D045744:Cell Line, Tumor; D015179:Colorectal Neoplasms / Q000378:metabolism; D048749:Cytokinesis; D004249:DNA Damage; D000078404:Epidermal Cells; D059565:G2 Phase Cell Cycle Checkpoints; D042822:Genomic Instability; D045325:HCT116 Cells; D006801:Humans; D019426:Integrases / Q000235:genetics / Q000378:metabolism*; D053547:Keratin-14 / Q000235:genetics; D053555:Keratin-5 / Q000235:genetics; D015603:Keratinocytes / Q000166:cytology; D008822:Mice, Transgenic; D008938:Mitosis; D011401:Promoter Regions, Genetic; D057891:Tetraploidy; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D053547:Keratin-14; D053555:Keratin-5; D014764:Viral Proteins; C045073:Cre recombinase; D019426:Integrases
keywords: Cre recombinase; DNA damage; aneuploidy; apoptosis; bypass of mitosis; cancer; cytokinesis failure; homologous recombination; tetraploidy; transgenic mice