Epitranscriptomic control of stress adaptations in Escherichia coli

Abstract The impacts of various stressors on bacterial systems have been studied at the phenotypic, transcriptional, and translational levels during the early stress response. However, the contributions of RNA modifications during stress adaptation remain largely unexplored. Here, we map the epitranscriptomic changes of Escherichia coli after exposure to oxidative and acid stress using direct RNA sequencing of mRNA, rRNA, pre-tRNA, and tRNA, combined with mass spectrometry, deletion mutant phenotyping, and single-nucleotide PCR. We identified widespread, dynamic mRNA modifications that include central metabolism transcripts and increased levels of rRNA methylations (m4Cm and m5C) under both stresses, with potential consequences for translation. In uncharged pre-tRNAs, stress-specific modifications via the Mnm and Q pathways accumulated at the wobble position; these modifications proved crucial for survival. Together, these findings reveal a multifaceted layer of post-transcriptional regulation, establishing the first comprehensive view of the bacterial epitranscriptome during the early stress response.