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,Borrelia burgdorferiB31, possesses two RM systems withN6-methyladenosine (m6A) MTase activity, which are encoded by thebbe02gene located on linear plasmid lp25 andbbq67on lp56. The specific recognition and/or methylation sequences had not been identified for either of theseB. burgdorferiMTases, 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-typeB. burgdorferias well as MTase mutants lacking either thebbe02gene alone or bothbbe02andbbq67genes. 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.,rpoSregulon) and acquisition by/transmission from the tick vector (e.g.,rrp1andpdeB). The results of this study provide a comprehensive view of the DNA methylation pattern inB. burgdorferi, and the accompanying gene expression profiles add to the emerging body of research on RM systems and gene regulation in bacteria.IMPORTANCELyme 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,Borrelia burgdorferi, 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 inB. burgdorferi, 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 inB. burgdorferiand evaluates the resulting effects on gene regulation in this important pathogen.

DNA methylation from a Type I restriction modification system influences gene expression and virulence in Streptococcus pyogenes

“…Of the m6A modifications called at RM sites, 4.9% of sites were called as m6A modified in less than 50% of sequencing reads, 23.7% of sites were called as modified in between 50–75% of sequencing reads, and finally 71.4% of sites were called as modified at greater than 75% of aligned reads. Previous studies have also reported heterogeneity in the frequency of SMRT sequencing reads with base modifications; it has been hypothesized that these differences are due to timing in DNA replication and subsequent methylation [33–35]. Whether there is a temporal component accounting for the heterogeneity in m6A DNA modifications, and whether this impacts other functions of m6A modifications, such as in influencing gene transcription in S .…”

DNA Methylation Epigenetically Regulates Gene Expression in Burkholderia cenocepacia and Controls Biofilm Formation, Cell Aggregation, and Motility

“…MTase BCAL3494 was specifically linked to methylation of the CACAG motif, and MTase BCAM0992 was linked to methylation of the GTWWAC motif. This strategy of DNA methylation analysis, in which the methylome of strains lacking MTases is determined, has been used in various bacteria, as it is an effective way to find associations between predicted MTases and genome-wide methylation motifs (39,40). For example, several methylation motifs were identified in Burkholderia pseudomallei, including motifs CA-CAG and GTWWAC (41).…”

Cytosine N4-Methylation via M.Ssp6803II Is Involved in the Regulation of Transcription, Fine- Tuning of DNA Replication and DNA Repair in the Cyanobacterium Synechocystis sp. PCC 6803

“…The combined characterization of the methylome and transcriptome in wild-type and methyltransferase-defective mutant cells revealed that DNA methylation also impacts gene expression patterns in the pathogenic bacteria Vibrio cholera (Chao et al, 2015) and Helicobacter pylori (Estibariz et al, 2019). In Borrelia burgdorferi DNA methylation through the natural RM system also impacts the global transcriptome (Casselli et al, 2018).…”