DNA Methylation by Restriction Modification Systems Affects the Global Transcriptome Profile in Borrelia burgdorferi

Abstract

Prokaryote restriction modification (RM) systems serve to protect bacteria from potentially detrimental foreign DNA. Recent evidence suggests that DNA methylation by the methyltransferase (MTase) components of RM systems can also have effects on transcriptome profiles. The type strain of the causative agent of Lyme disease, <i>Borrelia burgdorferi</i> B31, possesses two RM systems with <i>N</i>6-methyladenosine (m6A) MTase activity, which are encoded by the <i>bbe02</i> gene located on linear plasmid lp25 and <i>bbq67</i> on lp56. The specific recognition and/or methylation sequences had not been identified for either of these <i>B. burgdorferi</i> MTases, and it was not previously known whether these RM systems influence transcript levels. In the current study, single-molecule real-time sequencing was utilized to map genome-wide m6A sites and to identify consensus modified motifs in wild-type <i>B. burgdorferi</i> as well as MTase mutants lacking either the <i>bbe02</i> gene alone or both <i>bbe02</i> and <i>bbq67</i> genes. Four novel conserved m6A motifs were identified and were fully attributable to the presence of specific MTases. Whole-genome transcriptome changes were observed in conjunction with the loss of MTase enzymes, indicating that DNA methylation by the RM systems has effects on gene expression. Genes with altered transcription in MTase mutants include those involved in vertebrate host colonization (e.g., <i>rpoS</i> regulon) and acquisition by/transmission from the tick vector (e.g., <i>rrp1</i> and <i>pdeB</i>). The results of this study provide a comprehensive view of the DNA methylation pattern in <i>B. burgdorferi</i>, and the accompanying gene expression profiles add to the emerging body of research on RM systems and gene regulation in bacteria.<b>IMPORTANCE</b> Lyme disease is the most prevalent vector-borne disease in North America and is classified by the Centers for Disease Control and Prevention (CDC) as an emerging infectious disease with an expanding geographical area of occurrence. Previous studies have shown that the causative bacterium, <i>Borrelia burgdorferi</i>, methylates its genome using restriction modification systems that enable the distinction from foreign DNA. Although much research has focused on the regulation of gene expression in <i>B. burgdorferi</i>, the effect of DNA methylation on gene regulation has not been evaluated. The current study characterizes the patterns of DNA methylation by restriction modification systems in <i>B. burgdorferi</i> and evaluates the resulting effects on gene regulation in this important pathogen.