The pathogenicity island 1-encoded small RNA InvR mediates post-transcriptional feedback control of the activator HilA in .

Pathogenicity Island 1 (SPI1) encodes a type three secretion system (T3SS) essential for invasion of intestinal epithelial cells. Many environmental and regulatory signals control SPI1 gene expression, but in most cases, the molecular mechanisms remain unclear. Many of these regulatory signals control SPI1 at a post-transcriptional level and we have identified a number of small RNAs (sRNAs) that control the SPI1 regulatory circuit. The transcriptional regulator HilA activates expression of the genes encoding the SPI1 T3SS structural and primary effector proteins. Transcription of is controlled by the AraC-like proteins HilD, HilC, and RtsA. The mRNA 5' untranslated region (UTR) is ~350-nuclotides in length and binds the RNA chaperone Hfq, suggesting it is a likely target for sRNA-mediated regulation. We used the rGRIL-seq (reverse global sRNA target identification by ligation and sequencing) method to identify sRNAs that bind to the 5' UTR. The rGRIL-seq data, along with genetic analyses, demonstrate that the SPI1-encoded sRNA InvR base pairs at a site overlapping the ribosome binding site. HilD and HilC activate both and . InvR in turn negatively regulates the translation of the mRNA. Thus, the SPI1-encoded sRNA InvR acts as a negative feedback regulator of SPI1 expression. Our results suggest that InvR acts to fine-tune SPI1 expression and prevent overactivation of expression, highlighting the complexity of sRNA regulatory inputs controlling SPI1 and virulence.