Abstract
Campylobacter jejuni is a leading cause of foodborne illnesses worldwide. As a microaerobic pathogen, C. jejuni is subjected to DNA damages caused by various stresses such as reactive oxygen species (ROS) and UV radiations. The base excision repair (BER) system plays an important role in preventing mutations associated with oxidative DNA damage, but the system remains poorly characterized in Campylobacter. In this study, a BER homolog encoded by cj0595c (named nth) in C. jejuni was analyzed for endonuclease III activity and for its role in maintaining genomic stability. It was found that inactivation of nth resulted in elevated frequencies of spontaneous fluoroquinolone-resistant (FQR) and oxidative stress resistant (OXR) mutants, compared with the wild-type strain in C. jejuni. Sequencing analysis of the FQR and OXR mutants revealed that the elevated mutation rates were associated with C → T or G → A transition in gyrA (FQR mutants) or perR (for OXR mutants). In an in vitro assay, a purified recombinant C. jejuni Nth protein demonstrated endonuclease III activity that recognized and excised the thymine glycol (Tg) base from a double stranded DNA. These findings indicate that Nth functions as a BER repair enzyme in C. jejuni and is important for the repair of DNA damage, protecting the bacteria from stresses encountered within a host and in the environment.
Highlights
Campylobacter jejuni, a major bacterial foodborne pathogen, is among the most common causes of bacterial diarrhea in humans worldwide (Nachamkin et al, 2000)
We found that the nth gene encodes Endonuclease III of the base excision repair (BER) system and inactivation of nth increased the spontaneous mutation frequencies with the C:G → T:A transition change, promoting the emergence of fluoroquinolone resistant (FQR) and oxidative stress resistant (OXR) mutants in C. jejuni
Searching for annotations of DNA glycosylases in C. jejuni revealed that cj0086c, nth, and cj1620c genes are likely to encode C. jejuni Uracil-DNA glycosylase (Ung), endonuclease III (Nth), and adenine DNA glycosylase (MutY), respectively (Table 3)
Summary
Campylobacter jejuni, a major bacterial foodborne pathogen, is among the most common causes of bacterial diarrhea in humans worldwide (Nachamkin et al, 2000). DNA repair mechanisms remain poorly defined in Campylobacter It has been revealed through whole genome sequencing, that C. jejuni has an incomplete version of a DNA repair system compared to that found in E. coli (Parkhill et al, 2000; Gundogdu et al, 2007). It lacks important genes involved in DNA repair including lexA, which allows bacteria to mount a SOS response and survive sudden increases in DNA damage (Michel, 2005). MutH and MutL, which are methyl-directed mismatch repair (MMR) enzymes and repair replication errors that arise from mis-incorporations (mismatches) and strand slippage (frameshift errors) (Modrich, 1991), and MutM for base excision repair (BER), are absent in C. jejuni
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