Hierarchical Delivery of an Essential Host Colonization Factor in Enteropathogenic Escherichia coli

Thomas, Nikhil A.; Deng, Wanyin; Baker, Noel T.; Puente, José Luis Barrio de la; Finlay, B. Brett

doi:10.1074/jbc.m706019200

Cited by 38 publications

(80 citation statements)

References 46 publications

(45 reference statements)

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“…However, LifA/Efa1/ToxB do not have a typical signal sequence for exported proteins at their N termini, and how they gain entry and act within target cells is still unresolved. Our demonstration that LifA/Efa1/ToxB can be secreted and translocated into host cells via the LEE-encoded T3SS offers an instant solution to this paradox and provides possible explanations for many seemingly contradictory functions of LifA/Efa1/ToxB, because type III effectors are known to contribute to bacterial colonization, modulate Rho GTPases and tight junctions, and even affect type III secretion (3,9,34).…”

Section: Discussionmentioning

confidence: 99%

“…As expected, the three translocators EspB, EspA, and EspD had the highest SILAC ratios in this SILAC analysis ( Table I), confirming that they are preferentially secreted under our experimental conditions. Although the translocators are the most abundant type III-secreted substrates of WT EPEC, it is known that WT EPEC can secrete small amounts of some effectors (32,34). Indeed, several effectors, including both the LEE-encoded (Map, EspG, EspH, Tir, EspF, and EspZ) and the non-LEE-encoded (NleA, NleE, NleD, and EspJ) effectors displayed high SILAC ratios, although the ratios were much lower than those of the translocators (Table I).…”

Section: Experimental Strategy For Silac Analysis Of Epec Type IIImentioning

confidence: 99%

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Quantitative Proteomic Analysis of Type III Secretome of Enteropathogenic Escherichia coli Reveals an Expanded Effector Repertoire for Attaching/Effacing Bacterial Pathogens

Deng

Hoog

et al. 2012

Molecular & Cellular Proteomics

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Type III secretion systems are central to the pathogenesis and virulence of many important Gram-negative bacterial pathogens, and elucidation of the secretion mechanism and identification of the secreted substrates are critical to our understanding of their pathogenic mechanisms and developing potential therapeutics. Stable isotope labeling with amino acids in cell culture-based mass spectrometry is a quantitative and highly sensitive proteomics tool that we have previously used to successfully analyze the type III secretomes of Citrobacter rodentium and Salmonella enterica serovar Typhimurium. In this report, stable isotope labeling with amino acids in cell culture was used to analyze the type III secretome of enteropathogenic Escherichia coli (EPEC), an important human pathogen, which, together with enterohemorrhagic E. coli and C. rodentium, represents the family of attaching and effacing bacterial pathogens. We not only confirmed all 25 known EPEC type III-secreted proteins and effectors previously identified by conventional molecular and bioinformatical techniques but also identified several new type III-secreted proteins, including two novel effectors, C_0814/NleJ and LifA, that were shown to be translocated into host cells. LifA is a known virulence factor believed to act as a toxin as well as an adhesin, but its mechanism of secretion and function is not understood. With a predicted molecular mass of 366 kDa, LifA is the largest type III effector identified thus far in any pathogen. We further demonstrated that Efa1, ToxB, and Z4332 (homologs of LifA in enterohemorrhagic E. coli) are also type III effectors. This study has comprehensively characterized the type III secretome of EPEC, expanded the repertoire of type III-secreted effectors for the attaching and effacing pathogens, and provided new insights into the mode of function for LifA/Efa1/ToxB/Z4332, an important family of virulence factors. Molecular & Cellular

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Section: Discussionmentioning

confidence: 99%

Section: Experimental Strategy For Silac Analysis Of Epec Type IIImentioning

confidence: 99%

Quantitative Proteomic Analysis of Type III Secretome of Enteropathogenic Escherichia coli Reveals an Expanded Effector Repertoire for Attaching/Effacing Bacterial Pathogens

Deng

Hoog

et al. 2012

Molecular & Cellular Proteomics

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“…CesT is a multicargo chaperone that interacts with at least nine effectors (Thomas et al, 2007). We hypothesize that initial cesT expression and corresponding CesT protein levels could serve a role in preparing EPEC for effector translocation during infection.…”

Section: Discussionmentioning

confidence: 99%

“…We speculate that the cesT promoter is a transcriptional mechanism to ready the cell with CesT protein levels that serve to stabilize effectors as soon as they are synthesized. This could be particularly important for the in vivo translocation of Tir, which has been shown to be essential for the hierarchical secretion of effector proteins (Thomas et al, 2007) and thus for their eventual injection into host cells.…”

Section: Discussionmentioning

confidence: 99%

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Dual temporal transcription activation mechanisms control cesT expression in enteropathogenic Escherichia coli

Brouwers

Thomas

2012

Microbiology

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Novel insights into the mechanism of SepL‐mediated control of effector secretion in enteropathogenic Escherichia coli

et al. 2017

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Type three secretion systems (T3SSs) are virulence determinants employed by several pathogenic bacteria as molecular syringes to inject effector proteins into host cells. Diarrhea‐producing enteropathogenic Escherichia coli (EPEC) uses a T3SS to colonize the intestinal tract. T3S is a highly coordinated process that ensures hierarchical delivery of three classes of substrates: early (inner rod and needle subunits), middle (translocators), and late (effectors). Translocation of effectors is triggered upon host‐cell contact in response to different environmental cues, such as calcium levels. The T3S substrate specificity switch from middle to late substrates in EPEC is regulated by the SepL and SepD proteins, which interact with each other and form a trimeric complex with the chaperone CesL. In this study, we investigated the link between calcium concentration and secretion regulation by the gatekeeper SepL. We found that calcium depletion promotes late substrate secretion in a translocon‐independent manner. Furthermore, the stability, formation, and subcellular localization of the SepL/SepD/CesL regulatory complex were not affected by the absence of calcium. In addition, we demonstrate that SepL interacts in a calcium‐independent manner with the major export gate component EscV, which in turn interacts with both middle and late secretion substrates, providing a docking site for T3S. These results suggest that EscV serves as a binding platform for both the SepL regulatory protein and secreted substrates during the ordered assembly of the T3SS.

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Hierarchical Delivery of an Essential Host Colonization Factor in Enteropathogenic Escherichia coli

Cited by 38 publications

References 46 publications

Quantitative Proteomic Analysis of Type III Secretome of Enteropathogenic Escherichia coli Reveals an Expanded Effector Repertoire for Attaching/Effacing Bacterial Pathogens

Quantitative Proteomic Analysis of Type III Secretome of Enteropathogenic Escherichia coli Reveals an Expanded Effector Repertoire for Attaching/Effacing Bacterial Pathogens

Dual temporal transcription activation mechanisms control cesT expression in enteropathogenic Escherichia coli

Novel insights into the mechanism of SepL‐mediated control of effector secretion in enteropathogenic Escherichia coli

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