title: Lineage-Specific Methyltransferases Define the Methylome of the Globally Disseminated Escherichia coli ST131 Clone.
abstract: Escherichia coli sequence type 131 (ST131) is a clone of uropathogenic E. coli that has emerged rapidly and disseminated globally in both clinical and community settings. Members of the ST131 lineage from across the globe have been comprehensively characterized in terms of antibiotic resistance, virulence potential, and pathogenicity, but to date nothing is known about the methylome of these important human pathogens. Here we used single-molecule real-time (SMRT) PacBio sequencing to determine the methylome of E. coli EC958, the most-well-characterized completely sequenced ST131 strain. Our analysis of 52,081 methylated adenines in the genome of EC958 discovered three (m6)A methylation motifs that have not been described previously. Subsequent SMRT sequencing of isogenic knockout mutants identified the two type I methyltransferases (MTases) and one type IIG MTase responsible for (m6)A methylation of novel recognition sites. Although both type I sites were rare, the type IIG sites accounted for more than 12% of all methylated adenines in EC958. Analysis of the distribution of MTase genes across 95 ST131 genomes revealed their prevalence is highly conserved within the ST131 lineage, with most variation due to the presence or absence of mobile genetic elements on which individual MTase genes are located.
  
  
  OBJECTIVE
  DNA modification plays a crucial role in bacterial regulation. Despite several examples demonstrating the role of methyltransferase (MTase) enzymes in bacterial virulence, investigation of this phenomenon on a whole-genome scale has remained elusive until now. Here we used single-molecule real-time (SMRT) sequencing to determine the first complete methylome of a strain from the multidrug-resistant E. coli sequence type 131 (ST131) lineage. By interrogating the methylome computationally and with further SMRT sequencing of isogenic mutants representing previously uncharacterized MTase genes, we defined the target sequences of three novel ST131-specific MTases and determined the genomic distribution of all MTase target sequences. Using a large collection of 95 previously sequenced ST131 genomes, we identified mobile genetic elements as a major factor driving diversity in DNA methylation patterns. Overall, our analysis highlights the potential for DNA methylation to dramatically influence gene regulation at the transcriptional level within a well-defined E. coli clone.
authors.affiliation: Australian Infectious Diseases Research Centre and School of Chemistry and Molecular Biosciences, The University of Queensland, St. Lucia, Australia.
authors.forename: Brian M
authors.initials: BM
authors.lastname: Forde
authors.affiliation: Australian Infectious Diseases Research Centre and School of Chemistry and Molecular Biosciences, The University of Queensland, St. Lucia, Australia.
authors.forename: Minh-Duy
authors.initials: MD
authors.lastname: Phan
authors.affiliation: Australian Infectious Diseases Research Centre and School of Chemistry and Molecular Biosciences, The University of Queensland, St. Lucia, Australia.
authors.forename: Jayde A
authors.initials: JA
authors.lastname: Gawthorne
authors.affiliation: Australian Infectious Diseases Research Centre and School of Chemistry and Molecular Biosciences, The University of Queensland, St. Lucia, Australia.
authors.forename: Melinda M
authors.initials: MM
authors.lastname: Ashcroft
authors.affiliation: Australian Infectious Diseases Research Centre and School of Chemistry and Molecular Biosciences, The University of Queensland, St. Lucia, Australia.
authors.forename: Mitchell
authors.initials: M
authors.lastname: Stanton-Cook
authors.affiliation: Australian Infectious Diseases Research Centre and School of Chemistry and Molecular Biosciences, The University of Queensland, St. Lucia, Australia.
authors.forename: Sohinee
authors.initials: S
authors.lastname: Sarkar
authors.affiliation: Australian Infectious Diseases Research Centre and School of Chemistry and Molecular Biosciences, The University of Queensland, St. Lucia, Australia.
authors.forename: Kate M
authors.initials: KM
authors.lastname: Peters
authors.affiliation: Faculty of Science, Division of Genetics and Molecular Biology, Institute of Biological Sciences, University of Malaya, Kuala Lumpur, Malaysia.
authors.forename: Kok-Gan
authors.identifier: http://orcid.org/0000-0002-1883-1115
authors.initials: KG
authors.lastname: Chan
authors.affiliation: Faculty of Science, Division of Genetics and Molecular Biology, Institute of Biological Sciences, University of Malaya, Kuala Lumpur, Malaysia.
authors.forename: Teik Min
authors.initials: TM
authors.lastname: Chong
authors.affiliation: Faculty of Science, Division of Genetics and Molecular Biology, Institute of Biological Sciences, University of Malaya, Kuala Lumpur, Malaysia.
authors.forename: Wai-Fong
authors.initials: WF
authors.lastname: Yin
authors.affiliation: School of Biomedical and Healthcare Science, Plymouth University Peninsula Schools of Medicine and Dentistry, Plymouth, United Kingdom.
authors.forename: Mathew
authors.initials: M
authors.lastname: Upton
authors.affiliation: Australian Infectious Diseases Research Centre and School of Chemistry and Molecular Biosciences, The University of Queensland, St. Lucia, Australia m.schembri@uq.edu.au s.beatson@uq.edu.au.
authors.forename: Mark A
authors.identifier: http://orcid.org/0000-0003-4863-9260
authors.initials: MA
authors.lastname: Schembri
authors.affiliation: Australian Infectious Diseases Research Centre and School of Chemistry and Molecular Biosciences, The University of Queensland, St. Lucia, Australia m.schembri@uq.edu.au s.beatson@uq.edu.au.
authors.forename: Scott A
authors.initials: SA
authors.lastname: Beatson
mesh_terms: D019175:DNA Methylation; D004269:DNA, Bacterial / Q000378:metabolism*; D004927:Escherichia coli Infections / Q000453:epidemiology / Q000382:microbiology; D005838:Genotype; D014943:Global Health; D006801:Humans; D008780:Methyltransferases / Q000235:genetics / Q000378:metabolism*; D014552:Urinary Tract Infections / Q000453:epidemiology / Q000382:microbiology; D056689:Uropathogenic Escherichia coli / Q000145:classification / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification
chemical_list: D004269:DNA, Bacterial; D008780:Methyltransferases