Small RNA interactome of pathogenic E. coli revealed through crosslinking of RNase E

Shafagh A Waters, Sean P McAteer, Grzegorz Kudla, Ignatius Pang, Nandan P Deshpande, Timothy G Amos, Kai Wen Leong, Marc R Wilkins, Richard Strugnell, David L Gally, David Tollervey, Jai J Tree

Research output: Contribution to journalArticlepeer-review

Abstract

RNA sequencing studies have identified hundreds of non-coding RNAs in bacteria, including regulatory small RNA (sRNA). However, our understanding of sRNA function has lagged behind their identification due to a lack of tools for the high-throughput analysis of RNA-RNA interactions in bacteria. Here we demonstrate that in vivo sRNA-mRNA duplexes can be recovered using UV-crosslinking, ligation and sequencing of hybrids (CLASH). Many sRNAs recruit the endoribonuclease, RNase E, to facilitate processing of mRNAs. We were able to recover base-paired sRNA-mRNA duplexes in association with RNase E, allowing proximity-dependent ligation and sequencing of cognate sRNA-mRNA pairs as chimeric reads. We verified that this approach captures bona fide sRNA-mRNA interactions. Clustering analyses identified novel sRNA seed regions and sets of potentially co-regulated target mRNAs. We identified multiple mRNA targets for the pathotype-specific sRNA Esr41, which was shown to regulate colicin sensitivity and iron transport in E. coli Numerous sRNA interactions were also identified with non-coding RNAs, including sRNAs and tRNAs, demonstrating the high complexity of the sRNA interactome.

Original languageEnglish
Pages (from-to)374-387
JournalEMBO Journal
Volume36
Issue number3
Early online date11 Nov 2016
DOIs
Publication statusPublished - 1 Feb 2017

Keywords

  • CLIP-Seq
  • CRAC
  • EHEC
  • enterohaemorrhagic E. coli
  • non-coding RNA

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