RIPseq of VP882 phage activation in V. cholerae

Many, if not all, bacteria use quorum sensing (QS) to control gene expression and collective behaviours, and more recently QS has also been discovered in bacteriophages (phages). Phages can produce communication molecules of their own, or “listen in” on the host's communication processes, in order to switch between lytic and lysogenic modes of infection. In this project, we studied the interaction of Vibrio cholerae, the causative agent of cholera disease, with the lysogenic vibriophage VP882. The lytic cycle of VP882 is induced by the QS molecule DPO (3,5-dimethylpyrazin-2-ol), however, the global regulatory consequences of DPO-mediated VP882 activation have remained unclear. Using a combination of transcriptomic, genetic, and biochemical approaches, we discovered that induction of VP882 results in binding of phage transcripts to the major RNA chaperone Hfq, which in turn outcompete and down-regulate host-derived Hfq-dependent small RNAs (sRNAs). VP882 itself also encodes Hfq-binding sRNAs and we demonstrate that one of these sRNAs, named VpdS, modulates the expression of multiple host and phage mRNAs through a base-pairing mechanism and thereby promotes phage replication. We further show that host-derived sRNAs can affect phage replication by interfering with the translation of phage mRNAs and thus might be part of the phage defence arsenal of the host. Taken together, our data draw a complex picture of post-transcriptional interactions occurring between host- and phage-derived transcripts that together determine the phage-mediated lysis program. Overall design: To idenity Hfq-binding RNAs in Vibrio cholerae during VP882 activation, we used RNA co-immunoprecipitation followed by RNA-sequencing (RIP-seq). V. cholerae ?tdh + JSP-1269 (VP882::kanR) and ?tdh hfq::3XFLAG + JSP-1269 (VP882::kanR) were cultivated in M9 medium to OD600 of 0.2 and VP882 phage was activated by 25 µM DPO and 0.035% arabinose to induce the lytic cycle. Cells were subjected to RNA-seq analysis. Three biological replicates for each condition are provided.