Abstract
FinO domain proteins such as ProQ of the model pathogen Salmonella enterica have emerged as a new class of major RNA-binding proteins in bacteria. ProQ has been shown to target hundreds of transcripts, including mRNAs from many virulence regions, but its role, if any, in bacterial pathogenesis has not been studied. Here, using a Dual RNA-seq approach to profile ProQ-dependent gene expression changes as Salmonella infects human cells, we reveal dysregulation of bacterial motility, chemotaxis, and virulence genes which is accompanied by altered MAPK (mitogen-activated protein kinase) signaling in the host. Comparison with the other major RNA chaperone in Salmonella, Hfq, reinforces the notion that these two global RNA-binding proteins work in parallel to ensure full virulence. Of newly discovered infection-associated ProQ-bound small noncoding RNAs (sRNAs), we show that the 3'UTR-derived sRNA STnc540 is capable of repressing an infection-induced magnesium transporter mRNA in a ProQ-dependent manner. Together, this comprehensive study uncovers the relevance of ProQ for Salmonella pathogenesis and highlights the importance of RNA-binding proteins in regulating bacterial virulence programs.IMPORTANCE The protein ProQ has recently been discovered as the centerpiece of a previously overlooked "third domain" of small RNA-mediated control of gene expression in bacteria. As in vitro work continues to reveal molecular mechanisms, it is also important to understand how ProQ affects the life cycle of bacterial pathogens as these pathogens infect eukaryotic cells. Here, we have determined how ProQ shapes Salmonella virulence and how the activities of this RNA-binding protein compare with those of Hfq, another central protein in RNA-based gene regulation in this and other bacteria. To this end, we apply global transcriptomics of pathogen and host cells during infection. In doing so, we reveal ProQ-dependent transcript changes in key virulence and host immune pathways. Moreover, we differentiate the roles of ProQ from those of Hfq during infection, for both coding and noncoding transcripts, and provide an important resource for those interested in ProQ-dependent small RNAs in enteric bacteria.
Highlights
FinO domain proteins such as ProQ of the model pathogen Salmonella enterica have emerged as a new class of major RNA-binding proteins in bacteria
Salmonella virulence is predominantly mediated by effector proteins that are translocated into eukaryotic host cells via two type III secretion systems (T3SSs) encoded on the major Salmonella pathogenicity islands (SPIs): SPI-1, required for host cell invasion [2], and SPI-2, for intracellular survival [3]
To identify a cell culture-based model to test for a putative role of ProQ in Salmonella virulence, we moved the previously described proQ deletion [13] into a Salmonella strain that constitutively expresses GFP [28] and infected several established host cell types for the study of Salmonella pathogenesis
Summary
FinO domain proteins such as ProQ of the model pathogen Salmonella enterica have emerged as a new class of major RNA-binding proteins in bacteria. Of newly discovered infection-associated ProQ-bound small noncoding RNAs (sRNAs), we show that the 3=UTR-derived sRNA STnc540 is capable of repressing an infection-induced magnesium transporter mRNA in a ProQ-dependent manner Together, this comprehensive study uncovers the relevance of ProQ for Salmonella pathogenesis and highlights the importance of RNA-binding proteins in regulating bacterial virulence programs. Other SPIs as well as Salmonella motility and chemotaxis loci contribute, for example, by enabling the bacteria to sense and reach the proper host niche [4] These infection-relevant genes are controlled transcriptionally by various DNA-binding proteins, they are targeted at the RNA level by the global RBPs Hfq, CsrA, and CspC/E, each of which interacts with hundreds of different Salmonella transcripts [5,6,7,8]. Hfq, CsrA, and CspC/E are essential for Salmonella virulence in mouse models of infection [6, 9, 10]
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