title: Restriction endonucleases from invasive Neisseria gonorrhoeae cause double-strand breaks and distort mitosis in epithelial cells during infection.
abstract: The host epithelium is both a barrier against, and the target for microbial infections. Maintaining regulated cell growth ensures an intact protective layer towards microbial-induced cellular damage. Neisseria gonorrhoeae infections disrupt host cell cycle regulation machinery and the infection causes DNA double strand breaks that delay progression through the G2/M phase. We show that intracellular gonococci upregulate and release restriction endonucleases that enter the nucleus and damage human chromosomal DNA. Bacterial lysates containing restriction endonucleases were able to fragment genomic DNA as detected by PFGE. Lysates were also microinjected into the cytoplasm of cells in interphase and after 20 h, DNA double strand breaks were identified by 53BP1 staining. In addition, by using live-cell microscopy and NHS-ester stained live gonococci we visualized the subcellular location of the bacteria upon mitosis. Infected cells show dysregulation of the spindle assembly checkpoint proteins MAD1 and MAD2, impaired and prolonged M-phase, nuclear swelling, micronuclei formation and chromosomal instability. These data highlight basic molecular functions of how gonococcal infections affect host cell cycle regulation, cause DNA double strand breaks and predispose cellular malignancies.
authors.affiliation: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, 106 91 Stockholm, Sweden.
authors.forename: Linda
authors.initials: L
authors.lastname: Weyler
authors.affiliation: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, 106 91 Stockholm, Sweden.
authors.forename: Mattias
authors.initials: M
authors.lastname: Engelbrecht
authors.affiliation: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, 106 91 Stockholm, Sweden.
authors.forename: Manuel
authors.initials: M
authors.lastname: Mata Forsberg
authors.affiliation: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, 106 91 Stockholm, Sweden.
authors.forename: Karl
authors.initials: K
authors.lastname: Brehwens
authors.affiliation: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, 106 91 Stockholm, Sweden.
authors.forename: Daniel
authors.initials: D
authors.lastname: Vare
authors.affiliation: Department of Chemistry and Molecular Biology, University of Gothenburg, 405 30 Gothenburg, Sweden.
authors.forename: Katarina
authors.initials: K
authors.lastname: Vielfort
authors.affiliation: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, 106 91 Stockholm, Sweden.
authors.forename: Andrzej
authors.initials: A
authors.lastname: Wojcik
authors.affiliation: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, 106 91 Stockholm, Sweden.
authors.forename: Helena
authors.initials: H
authors.lastname: Aro
mesh_terms: D004249:DNA Damage; D004262:DNA Restriction Enzymes / Q000378:metabolism*; D004847:Epithelial Cells / Q000382:microbiology; D008938:Mitosis; D009344:Neisseria gonorrhoeae / Q000201:enzymology* / Q000472:pathogenicity
chemical_list: D004262:DNA Restriction Enzymes