Dataset Information

HilE is required for synergistic activation of SPI-1 gene expression in Salmonella enterica serovar Typhimurium.

ABSTRACT:

Background

Salmonella enterica serovar Typhimurium is an intestinal pathogen capable of infecting a wide range of animals. It initiates infection by invading intestinal epithelial cells using a type III secretion system encoded within Salmonella pathogenicity island 1 (SPI-1). The SPI-1 genes are regulated by multiple interacting transcription factors. The master regulator is HilD. HilE represses SPI-1 gene expression by binding HilD and preventing it from activating its target promoters. Previous work found that acetate and nutrients synergistically induce SPI-1 gene expression. In the present study, we investigated the role of HilE, nominally a repressor of SPI-1 gene expression, in mediating this response to acetate and nutrients.

Results

HilE is necessary for activation of SPI-1 gene expression by acetate and nutrients. In mutants lacking hilE, acetate and nutrients no longer increase SPI-1 gene expression but rather repress it. This puzzling response is not due to the BarA/SirA two component system, which governs the response to acetate. To identify the mechanism, we profiled gene expression using RNAseq in the wild type and a ?hilE mutant under different growth conditions. Analysis of these data suggested that the Rcs system, which regulates gene expression in response to envelope stress, is involved. Consistent with this hypothesis, acetate and nutrients were able to induce SPI-1 gene expression in mutants lacking hilE and the Rcs system.

Conclusions

While the exact mechanism is unknown, these results demonstrate the HilE, nominally a repressor of SPI-1 gene expression, can also function as an activator under the growth conditions investigated. Collectively, these results provide new insights regarding SPI-1 gene regulation and demonstrate that HilE is more complex than initially envisioned.

SUBMITTER: Hamed S

PROVIDER: S-EPMC7887791 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature

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Publications

HilE is required for synergistic activation of SPI-1 gene expression in Salmonella enterica serovar Typhimurium.

Hamed Selwan S Shawky Riham M RM Emara Mohamed M Slauch James M JM Rao Christopher V CV

BMC microbiology 20210216 1

<h4>Background</h4>Salmonella enterica serovar Typhimurium is an intestinal pathogen capable of infecting a wide range of animals. It initiates infection by invading intestinal epithelial cells using a type III secretion system encoded within Salmonella pathogenicity island 1 (SPI-1). The SPI-1 genes are regulated by multiple interacting transcription factors. The master regulator is HilD. HilE represses SPI-1 gene expression by binding HilD and preventing it from activating its target promoters ...[more]

PMID: 33593291

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Project description:The Salmonella type three secretion system (T3SS), encoded in the Salmonella pathogenicity island 1 (SPI1) locus, mediates the invasion of the host intestinal epithelium. SPI1 expression is dependent upon three AraC-like regulators: HilD, HilC, and RtsA. These regulators act in a complex feed-forward loop to activate each other and hilA, which encodes the activator of the T3SS structural genes. HilD has been shown to be the major integration point of most signals known to activate the expression of the SPI1 T3SS, acting as a switch to control induction of the system. HilE is a negative regulator that acts upon HilD. Here we provide genetic and biochemical data showing that HilE specifically binds to HilD but not to HilC or RtsA. This protein-protein interaction blocks the ability of HilD to bind DNA as shown by both an in vivo reporter system and an in vitro gel shift assay. HilE does not affect HilD dimerization, nor does it control the stability of the HilD protein. We also investigated the role of HilE during the infection of mice using competition assays. Although deletion of hilE does not confer a phenotype, the hilE mutation does suppress the invasion defect conferred by loss of FliZ, which acts as a positive signal controlling HilD protein activity. Together, these data suggest that HilE functions to restrict low-level HilD activity, preventing premature activation of SPI1 until positive inputs reach a threshold required to fully induce the system.IMPORTANCESalmonella is a leading cause of gastrointestinal and systemic disease throughout the world. The SPI1 T3SS is required for Salmonella to induce inflammatory diarrhea and to gain access to underlying tissue. A complex regulatory network controls expression of SPI1 in response to numerous physiological inputs. Most of these signals impinge primarily on HilD translation or activity. The system is triggered when HilD activity crosses a threshold that allows efficient activation of its own promoter. This threshold is set by HilE, which binds to HilD to prevent the inevitable minor fluctuations in HilD activity from inappropriately activating the system. The circuit also serves as a paradigm for systems that must integrate numerous environmental parameters to control regulatory output.

Project description:BACKGROUND:The antibiotics generally used in farm animals are rapidly losing their effectiveness all over the world as bacteria develop antibiotic resistance. Like some other pathogenic bacteria multidrug-resistant strains of Salmonella enterica serovar Typhimurium (S. Typhimurium) are also frequently found in animals and humans which poses a major public health concern. New strategies are needed to block the development of resistance and to prolong the life of traditional antibiotics. Thus, this study aimed to increase the efficacy of existing antibiotics against S. Typhimurium by combining them with opportunistic phenolic compounds gallic acid (GA), epicatechin, epicatechin gallate, epigallocatechin and hamamelitannin. Fractional inhibitory concentration indexes (FICI) of phenolic compound-antibiotic combinations against S. Typhimurium were determined. Based on the FICI and clinical importance, 1 combination (GA and ceftiofur) was selected for evaluating its effects on the virulence factors of this bacterium. Viability of Rattus norvegicus (IEC-6) cell in presence of this antibacterial combination was evaluated. RESULTS:Minimum inhibitory concentrations (MICs) of GA, epigallocatechin and hamamelitannin found against different strains of S. Typhimurium were 256, (512-1024), and (512-1024) ?g/mL, respectively. Synergistic antibacterial effect was obtained from the combination of erythromycin-epicatechin gallate (FICI: 0.50) against S. Typhimurium. Moreover, additive effects (FICI: 0.502-0.750) were obtained from 16 combinations against this bacterium. The time-kill assay and ultrastructural morphology showed that GA-ceftiofur combination more efficiently inhibited the growth of S. Typhimurium compared to individual antimicrobials. Biofilm viability, and swimming and swarming motilities of S. Typhimurium in presence of GA-ceftiofur combination were more competently inhibited than individual antimicrobials. Viabilities of IEC-6 cells were more significantly enhanced by GA-ceftiofur combinations than these antibacterials alone. CONCLUSIONS:This study suggests that GA-ceftiofur combination can be potential medication to treat S. Typhimurium-associated diarrhea and prevent S. Typhimurium-associated blood-stream infections (e.g.: fever) in farm animals, and ultimately its transmission from animal to human. Further in vivo study to confirm these effects and safety profiles in farm animal should be undertaken for establishing these combinations as medications.

Project description:Salmonella enterica is a foodborne pathogen often leading to gastroenteritis and is commonly acquired by consumption of contaminated food of animal origin. However, frequency of outbreaks linked to the consumption of fresh or minimally processed food of nonanimal origin is increasing. New infection routes of S. enterica by vegetables, fruits, nuts, and herbs have to be considered. This leads to special interest in S. enterica interactions with leafy products, e.g., salads, that are mainly consumed in a minimally processed form. The attachment of S. enterica to salad is a crucial step in contamination, but little is known about the bacterial factors required and mechanisms of adhesion. S. enterica possesses a complex set of adhesive structures whose functions are only partly understood. Potentially, S. enterica may deploy multiple adhesive strategies for adhering to various salad species and other vegetables. In this study, we systematically analyzed the contributions of the complete adhesiome, of lipopolysaccharide (LPS), and of flagellum-mediated motility of S. enterica serovar Typhimurium (STM) in adhesion to Valerianella locusta (corn salad). We deployed a reductionist, synthetic approach to identify factors involved in the surface binding of STM to leaves of corn salad, with particular regard to the expression of all known adhesive structures, using the Tet-on system. This work reveals the contribution of Saf fimbriae, type 1 secretion system-secreted BapA, an intact LPS, and flagellum-mediated motility of STM in adhesion to corn salad leaves.IMPORTANCE Transmission of gastrointestinal pathogens by contaminated fresh produce is of increasing relevance to human health. However, the mechanisms of contamination of, persistence on, and transmission by fresh produce are poorly understood. We investigated the contributions of the various adhesive structures of STM to the initial event in transmission, i.e., binding to the plant surface. A reductionist system was used that allowed experimentally controlled surface expression of individual adhesive structures and analyses of the contribution to binding to leave surfaces of corn salad under laboratory conditions. The model system allowed the determination of the relative contributions of fimbrial and nonfimbrial adhesins, the type 3 secretion systems, the O antigen of lipopolysaccharide, the flagella, and chemotaxis of STM to binding to corn salad leaves. Based on these data, future work could reveal the mechanism of binding and the relevance of interaction under agricultural conditions.

Project description:BackgroundIntestinal fibrosis is a common and serious complication of Crohn's disease characterized by the accumulation of fibroblasts, deposition of extracellular matrix, and formation of scar tissue. Although many factors including cytokines and proteases contribute to the development of intestinal fibrosis, the initiating mechanisms and the complex interplay between these factors remain unclear.MethodsChronic infection of mice with Salmonella enterica serovar Typhimurium was used to induce intestinal fibrosis. A murine protease-specific CLIP-CHIP microarray analysis was employed to assess regulation of proteases and protease inhibitors. To confirm up- or downregulation during fibrosis, we performed quantitative real-time polymerase chain reaction (PCR) and immunohistochemical stainings in mouse tissue and tissue from patients with inflammatory bowel disease. In vitro infections were used to demonstrate a direct effect of bacterial infection in the regulation of proteases.ResultsMice develop severe and persistent intestinal fibrosis upon chronic infection with Salmonella enterica serovar Typhimurium, mimicking the pathology of human disease. Microarray analyses revealed 56 up- and 40 downregulated proteases and protease inhibitors in fibrotic cecal tissue. Various matrix metalloproteases, serine proteases, cysteine proteases, and protease inhibitors were regulated in the fibrotic tissue, 22 of which were confirmed by quantitative real-time PCR. Proteases demonstrated site-specific staining patterns in intestinal fibrotic tissue from mice and in tissue from human inflammatory bowel disease patients. Finally, we show in vitro that Salmonella infection directly induces protease expression in macrophages and epithelial cells but not in fibroblasts.ConclusionsIn summary, we show that chronic Salmonella infection regulates proteases and protease inhibitors during tissue fibrosis in vivo and in vitro, and therefore this model is well suited to investigating the role of proteases in intestinal fibrosis.

Dataset Information

HilE is required for synergistic activation of SPI-1 gene expression in Salmonella enterica serovar Typhimurium.

Background

Results

Conclusions

Publications

HilE is required for synergistic activation of SPI-1 gene expression in Salmonella enterica serovar Typhimurium.

Similar Datasets

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