Analysis of the Campylobacter jejuni Genome by SMRT DNA Sequencing Identifies Restriction-Modification Motifs

Abstract

Campylobacter jejuni is a leading bacterial cause of human gastroenteritis. The goal of this study was to analyze the C. jejuni F38011 strain, recovered from an individual with severe enteritis, at a genomic and proteomic level to gain insight into microbial processes. The C. jejuni F38011 genome is comprised of 1,691,939 bp, with a mol.% (G+C) content of 30.5%. PacBio sequencing coupled with REBASE analysis was used to predict C. jejuni F38011 genomic sites and enzymes that may be involved in DNA restriction-modification. A total of five putative methylation motifs were identified as well as the C. jejuni enzymes that could be responsible for the modifications. Peptides corresponding to the deduced amino acid sequence of the C. jejuni enzymes were identified using proteomics. This work sets the stage for studies to dissect the precise functions of the C. jejuni putative restriction-modification enzymes. Taken together, the data generated in this study contributes to our knowledge of the genomic content, methylation profile, and encoding capacity of C. jejuni.

Methylation-dependent DNA discrimination in natural transformation of Campylobacter jejuni

“…These methylation motifs can be paired with their corresponding MTase genes using bioinformatics and genetic analysis. To date, 10 Campylobacter strains have been sequenced using this method, including four C. jejuni isolates (18)(19)(20)(21)(22). These strains include two laboratory isolates, one ovine isolate, and one human isolate.…”

“…Numerous DNA MTases have been identified in different C. jejuni strains. Active DNA MTases of C. jejuni, and the sequences that they methylate, were identified using single-molecule real-time (SMRT) sequencing (18)(19)(20)(21)(22)(23). With this approach, one methylation motif-RA m6 ATTY-was present in all four strains tested to date.…”

Mechanisms and roles of the LuxS system, methyl recycling, and DNA methylation on the physiology of Campylobacter jejuni

“…DNA is methylated at specific bases, with the three most common base modifications in prokaryotes being N4-methylcytosine (m4C), C5-methylcytosine (m5C) and N6methyladenine (m6A) (Korlach and Turner, 2012). C. jejuni is, as of now, known to possess both m6A and m5C base modifications (Murray et al, 2012;Mou et al, 2015;O'Loughlin et al, 2015). Enzymes that carry out base modifications, known as methyltransferases (MTases), recognize specific sequences motifs and transfer methyl groups from SAM to the specific base located at the motif (Vasu and Nagaraja, 2013).…”

“…There are also (Roberts et al, 2010). However, the only known characterized R-M systems in C. jejuni are Types I and II, which are described in more detail below (Murray et al, 2012;Mou et al, 2015;O'Loughlin et al, 2015).…”

“…In recent years, SMRT sequencing has become increasingly popular for its use in profiling methylomes of important pathogens, including E. coli (Fang et al, 2012), Helicobacter pylori (Krebes et al, 2014), Salmonella enterica (Pirone-Davies et al, 2015), Klebsiella pneumoniae (Doi et al, 2014), Enterococcus faecalis (Huo et al, 2015), and Campylobacter jejuni (Murray et al, 2012;Mou et al, 2015;O'Loughlin et al, 2015). In C. jejuni, SMRT sequencing has revealed several new findings on the methylome of this pathogen, including sequence motifs, base modification types, associated methyltransferases, and methylome characterization (including hypo-and hypermethylated regions of the genome, the genes present in these areas) and comparison of methylomes between different strains of C. jejuni (Murray et al, 2012;Mou et al, 2015;O'Loughlin et al, 2015). In particular, our group found that the genome-wide methylation patterns and the types of genes that were hypo-and hypermethylated were very similar between 11168 and 81-176.…”