Random Switching of the ModA11 Type III DNA Methyltransferase of Neisseria meningitidis Regulates Entner–Doudoroff Aldolase Expression by a Methylation Change in the eda Promoter Region

Phase-variable DNA methyltransferases (Mods) mediate epigenetic regulation of gene expression. These phase-variable regulons, called phasevarions, have been shown to regulate virulence and immunoevasion in multiple bacterial pathogens. How genome methylation switching mediates gene regulation is unresolved. Neisseria meningitidis remains a major cause of sepsis and meningitis worldwide. Previously, we reported that phase variation (rapid on/off switching) of the meningococcal ModA11 methyltransferase regulates 285 genes. Here we show a bioinformatic analysis that reveals only 26 of the regulated genes have a methylation site located upstream of the gene with potential for direct effect of methylation on transcription. To investigate how methylation changes are “read” to alter gene expression, we used a lacZ gene fusion approach. We showed a 182-nucleotide region upstream of the eda gene (Entner–Doudoroff aldolase) is sufficient to impart methylation-dependent regulation of eda. Site-directed mutagenesis of the 5′-ACGTm6AGG-3′ ModA11 site upstream of the eda gene showed that methylation of this site modulates eda expression. We show that eda is regulated by the PhoB homolog MisR, and that a MisR binding motif overlaps with the ModA11 methylation site. In a MisR mutant, regulation of eda is uncoupled from regulation by ModA11 phasevarion switching. The on/off switching of ModA11 leads to the presence or absence of a N6-methyladenine modification at thousands of sites in the genome. Most of these modifications have no impact on gene regulation. Moreover, the majority of the 285 gene regulon that is controlled by ModA11 phasevarion switching (259/285) are not directly controlled by methylation changes in the promoter region of the regulated genes. Our data are consistent with direct control via methylation of a subset of the regulon, like Eda, whose regulation will trigger secondary effects in expression of many genes.