Integrative DNA methylome and transcriptome analysis reveals DNA adenine methylation is involved in Salmonella enterica Typhimurium response to oxidative stress

ABSTRACT Salmonella could survive and replicate in macrophages, where it encounters multiple stresses. Deficiency of a DNA adenine methyltransferase impairs the survival of Salmonella enterica serovar Typhimurium (S. Typhimurium) in hydrogen peroxide. To investigate whether DNA methylation is involved in the expression of oxidative stress-responsive genes, we combined RNA-seq and single-molecule real-time sequencing to integrate transcriptome and methylome analysis. Here, we show that (i) the entire amount of m6A GATC remains stable during oxidative stress; (ii) no significant association is observed between DNA methylation and transcription level in most genes; (iii) coincidence of level change between transcription and m6A GATC in the regulatory regions is identified in 49 genes under oxidative stress. Some of them are known to contribute to bacterial defenses against oxidative stress through reducing H2O2 levels, directing aberrant protein product degradation and inducing outer membrane protein expression. Specifically, the transcription level of ahpCF is negatively correlated to the m6A GATC level in its regulatory region. The transcription of smpB is elevated along with the decrease in m6A level at position 2,879,954 on the minus strand. In contrast, mRNA levels of STM14_2773 and ycfR are positively correlated with m6A GATC content in their regulatory regions. Highly stable DNA methylome and coupled change of m6A GATC with gene expression in specific positions suggest that DNA methylation homeostasis at genome-wide and plasticity in specific regions are crucial for bacterial response to oxidative stress. These findings provide new insights into the epigenetic regulatory mechanisms of gene expression in S. Typhimurium in the host microenvironment. IMPORTANCE The intracellular pathogen Salmonella enterica serovar Typhimurium (S. Typhimurium) comes across a wide variety of stresses from entry to dissemination, such as reactive oxygen species. To adapt itself to oxidative stress, Salmonella must adopt various and complex strategies. In this study, we revealed that DNA adenine methyltransferase was essential for S. Typhimurium to survive in hydrogen peroxide. We then screened out oxidative stress-responsive genes that were potentially regulated by DNA methylation in S. Typhimurium. Our results show that the DNA methylome is highly stable throughout the genome, and the coupled change of m6A GATC with gene expression is identified in only a few positions, which suggests the complexity of the DNA methylation and gene expression regulation networks. The results may shed light on our understanding of m6A-mediated gene expression regulation in bacteria.