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ABSTRACTEnterohemorrhagic Escherichia coli (EHEC) O157:H7 is a common food-borne pathogen that can cause acute diseases. Lysine acetylation is a post-translational modification (PTM) that occurs in various prokaryotes and is regulated by CobB, the only deacetylase found in bacteria. Here, we demonstrated that CobB plays an important role in the virulence of EHEC O157:H7 and that deletion of cobB significantly decreased the intestinal colonization ability of bacteria. Using acetylation proteomic studies, we systematically identified several proteins that could be regulated by CobB in EHEC O157:H7. Among these CobB substrates, we found that acetylation at the K44 site of CesA, a chaperone for the type-III secretion system (T3SS) translocator protein EspA, weakens its binding to EspA, thereby reducing the stability of this virulence factor; this PTM ultimately attenuating the virulence of EHEC O157:H7. Furthermore, we showed that deacetylation of the K44 site, which is deacetylated by CobB, promotes the interaction between CesA and EspA, thereby increasing bacterial virulence in vitro and in animal experiments. In summary, we showed that acetylation influences the virulence of EHEC O157:H7, and uncovered the mechanism by which CobB contributes to bacterial virulence based on the regulation of CesA deacetylation.

The intracellular pathogen Salmonella enterica serovar Typhimurium (S. Typhimurium) exploits host macrophage as a crucial survival and replicative niche. To minimize host immune response stimulated by flagellin, the expression of flagellar genes is downregulated during S. Typhimurium growth within host macrophages. However, the underlying mechanisms are largely unknown. In this study, we show that STM14_1285 (named AsiR), a putative RpiR-family transcriptional regulator, which is downregulated within macrophages as previously reported and also confirmed here, positively regulates the expression of flagellar genes by directly binding to the promoter of flhDC. By generating an asiR mutant strain and a strain that persistently expresses asiR gene within macrophages, we confirmed that the downregulation of asiR contributes positively to S. Typhimurium replication in macrophages and systemic infection in mice, which could be attributed to decreased flagellar gene expression and therefore reduced flagellin-stimulated secretion of pro-inflammatory cytokines IL-1β and TNF-α. Furthermore, the acidic pH in macrophages is identified as a signal for the downregulation of asiR and therefore flagellar genes. Collectively, our results reveal a novel acidic pH signal-mediated regulatory pathway that is utilized by S. Typhimurium to promote intracellular replication and systemic pathogenesis by repressing flagellar gene expression.

Shigella is an intracellular pathogen that primarily infects the human colon and causes shigellosis. Shigella virulence relies largely on the type III secretion system (T3SS) and secreted effectors. VirF, the master Shigella virulence regulator, is essential for the expression of T3SS-related genes. In this study, we found that YhjC, a LysR-type transcriptional regulator, is required for Shigella virulence through activating the transcription of virF. Pathogenicity of the yhjC mutant, including colonization in the colons of guinea pigs as well as its ability for host cell adhesion and invasion, was significantly lowered. Expression levels of virF and nearly all VirF-dependent genes were downregulated by yhjC deletion, indicating that YhjC can activate virF transcription. Electrophoretic mobility shift assay analysis demonstrated that YhjC could bind directly to the virF promoter region. Therefore, YhjC is a novel virulence regulator that positively regulates the virF expression and promotes Shigella virulence. Additionally, genome-wide expression analysis identified the presence of other genes in the large virulence plasmid and a genome exhibiting differential expression in response to yhjC deletion, with 169 downregulated and 99 upregulated genes, indicating that YhjC also functioned as a global regulatory factor.