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.

Genus-Wide Comparative Genomics Analysis ofNeisseriato Identify New Genes Associated with Pathogenicity and Niche Adaptation ofNeisseriaPathogens

“…Those genes code for restriction endonucleases. Weyler et al found restriction endonucleases, which were released by intracellular Neisseria gonorrhoeae , damaged human chromosomal DNA, and distorted mitosis [103]. Besides, some other pathogenic-associated clusters were also found in Figure 7 network, such as the nitric oxide metabolic pathway (orf_1612-orf_1620) [104] and Tfp pilus assembly protein (orf_65, orf_129, orf_1665, orf_2249, orf_528, orf_1467, and orf_1757) [105].…”

Tampering of Viruses and Bacteria with Host DNA Repair: Implications for Cellular Transformation

“…Upon infection, some intestinal bacteria produce toxins that cause DNA lesions [ 170 ], potentially resulting in genome instability, and tumor initiation and progression. At the moment, four genotoxins are known to have this effect, namely colibactin (expressed by Escherichia coli ) [ 171 ], cytolethal distending toxin (CDT; expressed by certain Gram-negative bacteria [ 172 ], Shiga toxin (expressed by Shigella dysenteriae ) [ 173 , 174 , 175 ], and endonucleases (expressed by Neisseria gonorrhoeae ) [ 176 , 177 , 178 ].…”

Mechanisms of host manipulation by Neisseria gonorrhoeae

“…Overall, this review highlights the immense host manipulation capabilities the gonococcus possesses to promote pathogenesis, with additional mechanisms not covered in this review surrounding the direct manipulation of the host cell growth cycle (Jones et al, 2007;Vielfort et al, 2013;Weyler et al, 2014). Nevertheless, progress toward elucidating these infection strategies in situ will greatly support advancements in both prevention and treatment of this long-lasting human disease.…”