Cells with stochastically increased methyltransferase to restriction endonuclease ratio provide an entry for bacteriophage into protected cell population

Abstract

The action of Type II restriction–modification (RM) systems depends on restriction endonuclease (REase), which cleaves foreign DNA at specific sites, and methyltransferase (MTase), which protects host genome from restriction by methylating the same sites. We here show that protection from phage infection increases as the copy number of plasmids carrying the Type II RM Esp1396I system is increased. However, since increased plasmid copy number leads to both increased absolute intracellular RM enzyme levels and to a decreased MTase/REase ratio, it is impossible to determine which factor determines resistance/susceptibility to infection. By controlled expression of individual Esp1396I MTase or REase genes in cells carrying the Esp1396I system, we show that a shift in the MTase to REase ratio caused by overproduction of MTase or REase leads, respectively, to decreased or increased protection from infection. Consistently, due to stochastic variation of MTase and REase amount in individual cells, bacterial cells that are productively infected by bacteriophage have significantly higher MTase to REase ratios than cells that ward off the infection. Our results suggest that cells with transiently increased MTase to REase ratio at the time of infection serve as entry points for unmodified phage DNA into protected bacterial populations.

Features of the DNAEscherichia coliRecN interaction revealed by fluorescence microscopy and single-molecule methods

“…In microscopy experiments of cells expressing RecN::mCherry fusion DNA of bacteria was stained with the intercalating dye DAPI (Thermo). Stained bacteria were placed on microscope slides preparation of which was described previously [13].…”

“…In microscopy experiments of cells expressing RecN::mCherry fusion DNA of bacteria was stained with the intercalating dye DAPI (Thermo). Stained bacteria were placed on microscope slides preparation of which was described previously [12]. Fluorescence microscopy was performed using a Nikon Eclipse Ti-E inverted microscope under control of Micro-Manager 2.0 software.…”

Single-base resolution quantitative genome methylation analysis in the model bacterium Helicobacter pylori by enzymatic methyl sequencing (EM-Seq) reveals influence of strain, growth phase, and methyl homeostasis

“…Although known for a long time, the precise and diverse biological functions of DNA methylation and methyl transferases (MTases) in bacteria, whether part of restriction-modification systems or as orphan genes, are still understudied. Restriction-modification [methylation] systems have been widely characterized as “immune systems” of bacteria, which prevent or minimize the entry of heterologous DNA, in particular by bacteriophage infection [ 2 ], and also contribute to DNA integrity and replication [ 3 ]. Comprehensive bacterial epigenetics is a quite recently expanded field, which has shown that numerous bacterial species express multiple MTases, which methylate DNA at specific nucleotide recognition motifs that are distributed throughout the chromosome [ 1 ].…”