The Tat Substrate SufI Is Critical for the Ability of Yersinia pseudotuberculosis To Cause Systemic Infection

Abstract

The twin arginine translocation (Tat) system targets folded proteins across the inner membrane and is crucial for virulence in many important humanpathogenic bacteria. Tat has been shown to be required for the virulence of Yersinia pseudotuberculosis, and we recently showed that the system is critical for different virulence-related stress responses as well as for iron uptake. In this study, we wanted to address the role of the Tat substrates in in vivo virulence. Therefore, 22 genes encoding potential Tat substrates were mutated, and each mutant was evaluated in a competitive oral infection of mice. Interestingly, a ΔsufI mutant was essentially as attenuated for virulence as the Tat-deficient strain. We also verified that SufI was Tat dependent for membrane/periplasmic localization in Y. pseudotuberculosis. In vivo bioluminescent imaging of orally infected mice revealed that both the ΔsufI and ΔtatC mutants were able to colonize the cecum and Peyer's patches (PPs) and could spread to the mesenteric lymph nodes (MLNs). Importantly, at this point, neither the ΔtatC mutant nor the ΔsufI mutant was able to spread systemically, and they were gradually cleared. Immunostaining of MLNs revealed that both the ΔtatC and ΔsufI mutants were unable to spread from the initial infection foci and appeared to be contained by neutrophils, while wild-type bacteria readily spread to establish multiple foci from day 3 postinfection. Our results show that SufI alone is required for the establishment of systemic infection and is the major cause of the attenuation of the ΔtatC mutant.KEYWORDS Yersinia pseudotuberculosis, Tat pathway, virulence, SufI, mesenteric lymph nodes, neutrophils T he genus Yersinia includes three species that are pathogenic to humans: Yersinia pestis, the causative agent of plague, and the two enteropathogens Y. enterocolitica and Y. pseudotuberculosis, which normally cause a self-limiting disease with symptoms ranging from mild diarrhea, enterocolitis, septicemia, and mesenteric lymphadenitis to reactive arthritis in humans after ingestion of contaminated food or water (1). Once it is ingested, Y. pseudotuberculosis traverses the epithelial barrier through M cells and infects the associated lymphoid tissues, such as Peyer's patches (PPs) and cecal patches, and later spreads to the mesenteric lymph nodes (MLNs). Although the infection is usually self-limited in humans, the infection caused by the two enteropathogenic Yersinia species in mice readily progresses to become systemic and disseminates to the spleen and liver (2). The main virulence arsenal of Yersinia is the type III secretion system (T3SS), which is encoded by an ϳ70-kb virulence plasmid. The T3SS enables the translocation of virulence effector proteins directly into the cytosol of the target host cell, which results in the disruption of host signaling and early immune

The Tat system and its dependent cell division proteins are critical for virulence of extra-intestinal pathogenic Escherichia coli

“…In several important bacterial pathogens, the Tat system has been reported to be crucial for pathogenesis [8,14,17,35]. Consistently, our results also revealed that the deletion of the Tat system significantly attenuated the virulence of extra-intestinal pathogenic E. coli (ExPEC).…”

“…When calculating the in vivo bacterial loads, the mice were euthanized at each indicated time points and the organs were collected, weighed, homogenized in sterile saline, and plated on to LB for cell counting. Competitive infection assay, an accurate and sensitive approach to determine relative virulence, was used to determine whether the mutant was attenuated compared with the WT strain [17,[34][35][36]. In the competitive infection assay, a similar amount of viable cells in the mid-log phase of the WT strain and each Tat-related mutant were mixed and used to inject mouse intraperitoneally.…”

Citrobacter freundii fitness during bloodstream infection

“…A second study attributed the majority of Salmonella tat mutant attenuation to the simultaneous mis-localization of three proteins, AmiA, AmiC, and SufI 50 , though in this same work, the Salmonella sufI single mutant did not exhibit any colonization defect. Yersinia pseudotuberculosis tatC mutants are also attenuated following oral inoculation of mice 37 and in this case, a sufI mutation alone recapitulates the full attenuation level observed for the tatC mutant 51 . Our results indicate that the SufI protein of C. freundii is likely Tat-secreted based on the presence of a conserved signal sequence and we have demonstrated that mutation of C. freundii sufI results in a 7- and 17-fold fitness defect in the spleen and liver, respectively (Fig.…”

“…This protein is thought to provide cell division functions based in part on co-localization with the septal ring 56 and genetic analyses demonstrating suppression of divisome mutants via overexpression of SufI 57 , 58 . However, the reduced fitness of sufI mutants during infection is intriguing, since we and others have observed that SufI is not essential for bacterial cell division in standard laboratory media (data not shown) 50 , 51 .…”

Transposon-insertion sequencing screens unveil requirements for EHEC growth and intestinal colonization

“…The EHEC Δ tatABC mutant exhibited a marked colonization defect and a modest increase in bile sensitivity. The twin-arginine translocation (Tat) protein secretion system, which transports folded protein substrates across the cytoplasmic membrane (reviewed in (89,90)), has been implicated in the pathogenicity of a variety of Gram-negative pathogens, including enteric pathogens such as Salmonella enterica serovar Typhimurium (91–93), Yersinia pseudotuberculosis (94,95), Campylobacter jejuni (96), and Vibrio cholerae (97). Attenuation of Tat mutants can reflect the combined absence of a variety of secreted factors.…”