The secRNome of Listeria monocytogenes Harbors Small Noncoding RNAs That Are Potent Inducers of Beta Interferon

Cellular sensing of bacterial RNA is increasingly recognized as a determinant of host-pathogen interactions. The intracellular pathogen Listeria monocytogenes induces high levels of type I interferons (alpha/beta interferons [IFN-α/β]) to create a growth-permissive microenvironment during infection. We previously demonstrated that RNAs secreted by L. monocytogenes (comprising the secRNome) are potent inducers of IFN-β. We determined the composition and diversity of the members of the secRNome and found that they are uniquely enriched for noncoding small RNAs (sRNAs). Testing of individual sRNAs for their ability to induce IFN revealed several sRNAs with this property. We examined ril32, an intracellularly expressed sRNA that is highly conserved for the species L. monocytogenes and that was the most potent inducer of IFN-β expression of all the sRNAs tested in this study, in more detail. The rli32-induced IFN-β response is RIG-I (retinoic acid inducible gene I) dependent, and cells primed with rli32 inhibit influenza virus replication. We determined the rli32 motif required for IFN induction. rli32 overproduction promotes intracellular bacterial growth, and a mutant lacking rli32 is restricted for intracellular growth in macrophages. rli32-overproducing bacteria are resistant to H(2)O(2) and exhibit both increased catalase activity and changes in the cell envelope. Comparative transcriptome sequencing (RNA-Seq) analysis indicated that ril32 regulates expression of the lhrC locus, previously shown to be involved in cell envelope stress. Inhibition of IFN-β signaling by ruxolitinib reduced rli32-dependent intracellular bacterial growth, indicating a link between induction of the interferon system and bacterial physiology. rli32 is, to the best of our knowledge, the first secreted individual bacterial sRNA known to trigger the induction of the type I IFN response.