Host Acid Signal Controls Salmonella Flagella Biogenesis through CadC-YdiV Axis
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ABSTRACT:
ORGANISM(S): Salmonella enterica subsp. enterica serovar Typhimurium
PROVIDER: GSE210570 | GEO | 2022/08/06
REPOSITORIES: GEO
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Project description:Genetically engineered Salmonella Typhimurium are potent vectors for prophylactic and therapeutic measures against pathogens as well as cancer. This is based on the potent adjuvanticity that supports strong immune responses. The physiology of Salmonella is well understood. It simplifies engineering of both enhanced immune-stimulatory properties as well as safety features, thus, resulting in an appropriate balance between attenuation and efficacy for clinical applications. A major virulence factor of Salmonella is the flagellum. It is also a strong pathogen-associated molecular pattern recognized by extra- and intracellular receptors of immune cells of the host. At the same time, it represents a serious metabolic burden. Accordingly, the bacteria evolved tight regulatory mechanisms that control flagella synthesis in vivo. Here, we systematically investigated the immunogenicity and adjuvant properties of various flagella mutants of Salmonella in vitro and in a mouse cancer model in vivo. We found that mutants lacking the flagellum-specific ATPase FliHIJ or the inner membrane ring FliF displayed the greatest stimulatory capacity and strongest anti-tumor effects, while remaining safe in vivo. Scanning electron microscopy revealed the presence of outer membrane vesicles in the ΔfliF and ΔfliHIJ mutants. Finally, the combination of the ΔfliF and ΔfliHIJ mutations with our previously described attenuated and immunogenic background strain SF102 displayed strong efficacy against the highly resistant cancer cell line RenCa. We thus conclude that manipulating flagella biosynthesis has great potential for the construction of highly efficacious and versatile Salmonella vector strains.
2019-11-25 | GSE116623 | GEO
Project description:Flagella-driven motility of Salmonella enterica serovar Typhimurium facilitates host colonization. However, the large extracellular flagellum is also a prime target for the immune system. As consequence, expression of flagella is bistable within a population of Salmonella, resulting in flagellated and non-flagellated subpopulations. This allows the bacteria to maximize fitness in hostile environments. The degenerate EAL-domain protein RflP (formerly YdiV) is responsible for the bistable expression of flagella by directing the flagellar master regulatory complex FlhD4C2 to proteolytic degradation. The environmental cues controlling expression of rflP and thus the bistable flagellar biosynthesis remain ambiguous. Here, we demonstrate that RflP responds to cell envelope stress and alterations of outer membrane integrity. Lipopolysaccharide (LPS) truncation mutants of Salmonella Typhimurium exhibited increasing motility defects due to downregulation of flagellar gene expression. Transposon mutagenesis and genetic profiling revealed that σ24 (RpoE) and Rcs phosphorelay-dependent cell envelope stress response systems sense modifications of lipopolysaccharide, low pH activity of the complement system. This subsequently results in activation of RflP expression and degradation of FlhD4C2 via ClpXP. We speculate that diverse hostile environments inside the host might result in cell envelope damage and would thus trigger the repression of resource-costly and immunogenic flagella biosynthesis via activation of the cell envelope stress response.
2018-04-13 | GSE87447 | GEO
Project description:Host Acid Signal Controls Salmonella Flagella Biogenesis through CadC-YdiV Axis
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Project description:Using RNA seq, gene expression profiles were compared between wild-type Salmonella enterica Typhimurium 14028s and its isogenic srfM2 mutant and genes differentially expressed in the srfM2 mutant were identified.
2018-11-16 | GSE113988 | GEO
Project description:The flagellum provides essential motility in the pathogenic process of Salmonella host invasion. However, flagellin is strongly antigenic and can be recognized by TLR5, resulting in activation of the host immune system to clear the invading pathogen. The presence of flagella also disrupts the integrity of the bacterial cytosol, making it more vulnerable to external adversities. Salmonella must rapidly shut down the flagella synthesis system after entering the host cell to evade the host immune system and resist damage from external adverse factors. Our studies showed YdiV(STM1344) play a key role in regulating the transcription of flagella-related genes during Salmonella invasion. After the ydiV(stm1344) gene was knocked out, the pathogenicity of Salmonella was significantly reduced. Here, we performed a TMT-based comparative quantitative proteomics analysis to reveal the proteomic differences between the WT and △ydiV(stm1344) in simulated host environment medium.
2022-10-12 | PXD035693 | Pride
Project description:Campylobacter jejuni produces a single flagellum on one or both poles of the cell, the polar flagella not only confer the bacterium darting motility but also involved in its virulence and metabolic processes. FlhF is a lately defined flagella biosynthesis regulator, but how it regulates flagella assembly isn’t clear at present. Previous research had indicated FlhF was associated with gene expression, to understanding its role in flagella system and its correlation with C. jejuni virulence, the global gene expression profiles of C. jejuni NCTC 11168 after inactivation of FlhF was analyzed. Transcriptome analysis revealed that ten genes were significantly differentially expressed with 6 down-regulated and 4 up-regulated, these genes are involved in diverse cellular functions including bacterial chemotaxis, flagellar assembly and two-component system, significance analysis revealed that the pathway of flagellar assembly was remarkable affected. Further studies were focused on the flagella system, FlhF was found play a global role as 63% (41/65) of the flagella genes were down-regulated, it influences flagella gene transcription from the early stage, this effect persists during the synthesis process and has a remarkable impact on the late part. The results above expand our cognition of this regulator, and also helpful for comprehensively and systematically reveal its role in flagella system.
2018-10-03 | GSE87852 | GEO
Project description:Gene expression profiles of Escherichia coli K-12 W3110 were compared as a function of steady-state external pH. Cultures were grown with aeration to an optical density at 600 nm of 0.3 in potassium-modified Luria-Bertani medium buffered at pH 5.0, 7.0, and 8.7. For each of the three pH conditions, cDNA from RNA of five independent cultures was hybridized to Affymetrix E. coli arrays. Analysis of variance with a significance level of 0.001 resulted in 98% power to detect genes showing a twofold difference in expression. Normalized expression indices were calculated for each gene and intergenic region (IG). Differential expression among the three pH classes was observed for 763 genes and 353 IGs. Hierarchical clustering yielded six well-defined clusters of pH profiles, designated Acid High (highest expression at pH 5.0), Acid Low (lowest expression at pH 5.0), Base High (highest at pH 8.7), Base Low (lowest at pH 8.7), Neutral High (highest at pH 7.0, lower in acid or base), and Neutral Low (lowest at pH 7.0, higher at both pH extremes). Flagellar and chemotaxis genes were repressed at pH 8.7 (Base Low cluster), where the cell's transmembrane proton potential is diminished by the maintenance of an inverted pH gradient. High pH also repressed the proton pumps cytochrome o (cyo) and NADH dehydrogenases I and II. By contrast, the proton-importing ATP synthase F1Fo and the microaerophilic cytochrome d (cyd), which minimizes proton export, were induced at pH 8.7. These observations are consistent with a model in which high pH represses synthesis of flagella, which expend proton motive force, while stepping up electron transport and ATPase components that keep protons inside the cell. Acid-induced genes, on the other hand, were coinduced by conditions associated with increased metabolic rate, such as oxidative stress. All six pH-dependent clusters included envelope and periplasmic proteins, which directly experience external pH. Overall, this study showed that (i) low pH accelerates acid consumption and proton export, while coinducing oxidative stress and heat shock regulons; (ii) high pH accelerates proton import, while repressing the energy-expensive flagellar and chemotaxis regulons; and (iii) pH differentially regulates a large number of periplasmic and envelope proteins. Keywords: Steady State
2006-03-23 | GSE4511 | GEO
Project description:The long external filament of bacterial flagella is composed of several thousand copies of a single protein, flagellin. Here, we explore the role played by lysine methylation of flagellin in Salmonella, which requires the methylase FliB. We show that both flagellins of Salmonella enterica serovar Typhimurium, FliC and FljB, are methylated at surface-exposed lysine residues by FliB. A Salmonella Typhimurium mutant deficient in flagellin methylation is outcompeted for gut colonization in a gastroenteritis mouse model, and methylation of flagellin promotes bacterial invasion of epithelial cells in vitro. Lysine methylation increases the surface hydrophobicity of flagellin and enhances flagella-dependent adhesion of Salmonella to phosphatidylcholine vesicles and epithelial cells.
2020-04-27 | PXD017935 | Pride
Project description:Flagellum plays a crucial role in the invasion process of Salmonella, while also functioning as a significant antigen to trigger host pyroptosis. The regulation of flagella biogenesis is essential for both pathogenicity and immune escape of Salmonella. Here, we identified the conserved and unknown function protein STM0435 as a new flagellar regulator. ∆stm0435 strain exhibit higher pathogenicity in both cellular and animal infection experiments compared to wild-type Salmonella. Conjoint analysis of proteomics and transcriptomics demonstrated that almost all the flagellar genes were dramatically increased in a ∆stm0435 strain compared to wild-type Salmonella. Microscale thermophoresis assay showed that the purified STM0435 protein bound c-di-GMP with a ~194 µM KD affinity. The crystal structures of apo-STM0435 and STM0435&c-di-GMP complex were determined. Structural analysis revealed that R33, R137 and D138 of STM0435 are essential for c-di-GMP binding. Mutant that is unable to bind to c-di-GMP loses its ability to regulate motility, indicating that STM0435 affected the Salmonella flagellar biogenesis process by binding to c-di-GMP.
2023-08-05 | GSE239303 | GEO
Project description:Gene expression profiles of Escherichia coli K-12 W3110 were compared as a function of steady-state external pH. Cultures were grown with aeration to an optical density at 600 nm of 0.3 in potassium-modified Luria-Bertani medium buffered at pH 5.0, 7.0, and 8.7. For each of the three pH conditions, cDNA from RNA of five independent cultures was hybridized to Affymetrix E. coli arrays. Analysis of variance with a significance level of 0.001 resulted in 98% power to detect genes showing a twofold difference in expression. Normalized expression indices were calculated for each gene and intergenic region (IG). Differential expression among the three pH classes was observed for 763 genes and 353 IGs. Hierarchical clustering yielded six well-defined clusters of pH profiles, designated Acid High (highest expression at pH 5.0), Acid Low (lowest expression at pH 5.0), Base High (highest at pH 8.7), Base Low (lowest at pH 8.7), Neutral High (highest at pH 7.0, lower in acid or base), and Neutral Low (lowest at pH 7.0, higher at both pH extremes). Flagellar and chemotaxis genes were repressed at pH 8.7 (Base Low cluster), where the cell's transmembrane proton potential is diminished by the maintenance of an inverted pH gradient. High pH also repressed the proton pumps cytochrome o (cyo) and NADH dehydrogenases I and II. By contrast, the proton-importing ATP synthase F1Fo and the microaerophilic cytochrome d (cyd), which minimizes proton export, were induced at pH 8.7. These observations are consistent with a model in which high pH represses synthesis of flagella, which expend proton motive force, while stepping up electron transport and ATPase components that keep protons inside the cell. Acid-induced genes, on the other hand, were coinduced by conditions associated with increased metabolic rate, such as oxidative stress. All six pH-dependent clusters included envelope and periplasmic proteins, which directly experience external pH. Overall, this study showed that (i) low pH accelerates acid consumption and proton export, while coinducing oxidative stress and heat shock regulons; (ii) high pH accelerates proton import, while repressing the energy-expensive flagellar and chemotaxis regulons; and (iii) pH differentially regulates a large number of periplasmic and envelope proteins. Experiment Overall Design: Gene expression profiles of Escherichia coli K-12 W3110 were compared as a function of steady-state external pH. Overnight cultures were diluted 1:1000 in potassium-modified Luria-Bertani medium (LBK) buffered with 50 mM HOMOPIPES at pH 5.0, pH 7.0, and pH 8.7. Bacteria were cultured in baffled flasks (less than 10% volume filled) with rotation at 240 rpm, incubated at 37°C to an optical density at 600 nm of 0.3. For each of the three pH conditions, RNA was isolated from five independent cultures. Labeled cDNA was hybridized to Affymetrix antisense arrays according to standard procedures. To analyze the expression levels, Dchip software was used to generate model-based expression indices normalized to sample pH 7 replicate 1. ANOVA was used to identify genes with sitnificant expression differences among the three pH classes (p = 0.001). For genes showing significant differences, the Log2 expression ratios were determined for each pair of pH classes, and significance was determined by Tukey's test (p = 0.001).
2007-09-21 | E-GEOD-4511 | biostudies-arrayexpress