Conserved DNA Methyltransferases: A Window into Fundamental Mechanisms of Epigenetic Regulation in Bacteria

Abstract

An increasing number of studies have reported that bacterial DNA methylation has important functions beyond the roles in restriction-modification systems, including the ability of affecting clinically relevant phenotypes such as virulence, host colonization, sporulation, biofilm formation, among others. Although insightful, such studies have a largely ad hoc nature and would benefit from a systematic strategy enabling a joint functional characterization of bacterial methylomes by the microbiology community. In this opinion article, we propose that highly conserved DNA methyltransferases (MTases) represent a unique opportunity for bacterial epigenomic studies. These MTases are rather common in bacteria, span various taxonomic scales, and are present in multiple human pathogens. Apart from well-characterized core DNA MTases, like those from Vibrio cholerae, Salmonella enterica, Clostridioides difficile, or Streptococcus pyogenes, multiple highly conserved DNA MTases are also found in numerous human pathogens, including those belonging to the genera Burkholderia and Acinetobacter. We discuss why and how these MTases can be prioritized to enable a community-wide, integrative approach for functional epigenomic studies. Ultimately, we discuss how some highly conserved DNA MTases may emerge as promising targets for the development of novel epigenetic inhibitors for biomedical applications.

Keywords: antimicrobials; methylome; persistent/core genes; restriction-modification systems; virulence.

Figure 1.

Summary of MTase conservation in bacterial genomes from Genbank. (a) Phylogenetic tree of the 139 bacterial species (colored by Phylum), for which at least 10 complete genomes were available at Genbank (corresponding to a total of 5,568 genomes). Heatmap corresponds to the density (per genome per Mb) of Types I, II, III MTases and Type IIC R-M systems for each species. Bar plots indicate the percentage of the most abundant MTase(s) found in each species, assuming as inclusion criteria a minimum of 80% similarity in amino-acid sequence and less than 20% difference in protein length. Stippled lines indicate a threshold of 80% above which an MTase can be considered persistent. 100% denotes a core gene. (b) Pie-chart summarizing the percentages of species analyzed containing either persistent non-core (n.c.) MTases, core MTases, both, or none. (c) Pie-charts showing the breakdown of total, persistent, and core MTases per Type.

Figure 2.

Summary of the organization and target recognition motifs of persistent MTases based on the REBASE database. Yellow circles represent solitary MTases, whereas red ones represent complete systems. 100% means that the MTase is always found either as solitary (without a cognate endonuclease) or as part of a complete R-M system. Values below 100% indicate that both organizations are present, being the most predominant highlighted. For example, Type II GTWWAC-recognizing MTases are exclusively solitary, whereas Type III CACAG-recognizing MTases are exclusively found within complete R-M systems. GATC-recognizing MTases are found as solitary in 98.9% of the species analyzed, and the remaining 1.1% in complete systems. Target recognition motifs shown are based on the REBASE database. Circle radius is proportional to the number of species in which the MTase is present

Figure 3.

Illustrative examples of sequence similarity networks of persistent MTases conserved at different taxonomic resolutions (Supplementary Table 5). Each node represents one protein. To avoid redundancy and improve visualization, only one genome per species is shown (typically the reference/representative genome). Edges correspond to pairwise protein sequence identity >60%. Node colors correspond to different phyla.

Figure 4.

Overview of a large-scale community-wide integrative approach for bacterial methylome analyses. Core and persistent MTases can be prioritized to build a trans-omic network across multiple laboratories merging multiple functional data gathered at different experimental conditions. The latter may build upon MTase mutants generated by Transposon Insertion mutagenesis coupled with deep Sequencing (TIS), site-directed mutagenesis of methylation sites, genome-wide profiling of DNA binding proteins (ChIP-seq), transcription start site (TSS) mapping, and identification of methylation-sensitive transcription factors. Multiple layers of omics data may ultimately be commonly shared, linked to other resources (e.g., REBASE), allow for an in-depth analysis of, for example, methylation motif conservation, phase-variable DNA methyltransferases, and accelerate the research of novel epigenomic inhibitors.