Crystal structure of Clostridium difficile toxin A

Chumbler, Nicole M.; Rutherford, Stacey A.; Zhang, Zhifen; Farrow, Melissa A.; Lisher, John P.; Farquhar, Erik R.; Giedroc, David P.; Spiller, Benjamin W.; Melnyk, Roman A.; Lacy, D. Borden

doi:10.1038/nmicrobiol.2015.2

Cited by 88 publications

(127 citation statements)

References 46 publications

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“…A number of C. difficile virulence factors have a Zn requirement; however, the impact of Zn on CDI has yet to be explored 14–17 . To test this, we used a mouse model of CDI that induces susceptibility to C. difficile through administration of cefoperazone (0.5 mg/ml)(Supplementary Fig.…”

mentioning

confidence: 99%

Dietary zinc alters the microbiota and decreases resistance to Clostridium difficile infection

Zackular

Moore

Jordan

et al. 2016

Nat Med

225

179

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Clostridium difficile is the most commonly reported nosocomial pathogen in the United States and is an urgent public health concern worldwide1. Over the past decade, incidence, severity, and costs associated with C. difficile infection (CDI) have increased dramatically2. CDI is most commonly initiated by antibiotic-mediated disruption of the gut microbiota; however, non-antibiotic associated CDI cases are well documented and on the rise3,4. This suggests that unexplored environmental, nutrient, and host factors likely influence CDI. Here we show that excess dietary zinc (Zn) significantly alters the gut microbiota and in turn reduces the threshold of antibiotics needed to confer susceptibility to C. difficile infection. In mice colonized with C. difficile, excess dietary Zn severely exacerbates C. difficile-associated disease by increasing toxin activity and altering the host immune response. In addition, we show that the Zn binding S100 protein calprotectin is antimicrobial against C. difficile and an essential component of the innate immune response to CDI. Together, these data suggest that nutrient Zn levels play a key role in determining susceptibility to CDI and severity of disease, and that calprotectin-mediated metal limitation is an important factor in the host immune response to C. difficile.

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mentioning

confidence: 99%

Dietary zinc alters the microbiota and decreases resistance to Clostridium difficile infection

Zackular

Moore

Jordan

et al. 2016

Nat Med

225

179

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“…3B). We presume that the residual activity of the mutant seen in the caspase-3/7 activation assay is a result of residual glucosylating activity, the effects of which are detectable only at the highest concentration tested (34,44). When the cells were challenged with higher concentrations (Ն100 pM) of TcdB, very little caspase activation was detected (Fig.…”

Section: Figmentioning

confidence: 93%

Clostridium difficile Toxins TcdA and TcdB Cause Colonic Tissue Damage by Distinct Mechanisms

et al. 2016

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e As the major cause of antibiotic-associated diarrhea, Clostridium difficile is a serious problem in health care facilities worldwide. C. difficile produces two large toxins, TcdA and TcdB, which are the primary virulence factors in disease. The respective functions of these toxins have been difficult to discern, in part because the cytotoxicity profiles for these toxins differ with concentration and cell type. The goal of this study was to develop a cell culture model that would allow a side-by-side mechanistic comparison of the toxins. Conditionally immortalized, young adult mouse colonic (YAMC) epithelial cells demonstrate an exquisite sensitivity to both toxins with phenotypes that agree with observations in tissue explants. TcdA intoxication results in an apoptotic cell death that is dependent on the glucosyltransferase activity of the toxin. In contrast, TcdB has a bimodal mechanism; it induces apoptosis in a glucosyltransferase-dependent manner at lower concentrations and glucosyltransferase-independent necrotic death at higher concentrations. The direct comparison of the responses to TcdA and TcdB in cells and colonic explants provides the opportunity to unify a large body of observations made by many independent investigators. C lostridium difficile is the most common cause of antibioticassociated diarrhea in the United States, and C. difficile infection (CDI) has been steadily increasing in prevalence and severity over the last 15 years (1-3). Symptoms of CDI can range from mild diarrhea to pseudomembranous colitis, and hallmarks of the disease include neutrophil infiltration, fluid release, and necrotic lesions in the colonic epithelium (4, 5). The bacteria produce two main virulence factors, large toxins called TcdA and TcdB (6, 7).The respective function and relative importance of each toxin in pathogenesis have been active topics of investigation. Genetic knockout experiments in C. difficile have shown that both toxins are important for disease pathology, although TcdB alone is sufficient to cause death in both the hamster and mouse models (6-8). For many years, TcdA and TcdB have been thought to act synergistically, with TcdA acting as an enterotoxin and TcdB acting as a cytotoxin (9, 10). The general term enterotoxin refers to the capacity of TcdA to induce inflammation, cytokine release, and fluid secretion in animal intoxication models (11-13). While TcdB does not always induce these same phenotypes in models, such as the ileal loop model, it has been shown to disrupt the integrity of the epithelial structure in human explant and xenograft models (14, 15). TcdB is also notably more potent as a cytotoxin in cell culture models (9, 10, 16).The toxins have an N-terminal glucosyltransferase domain (GTD) that is delivered into the host cytosol by the C-terminal portion of the protein (17, 18). The GTD has been shown to target and inactivate a number of Rho-family GTPases (19,20). This inactivation has been linked to a cell rounding or cytopathic effect (CPE) (21-24) and to an apoptotic cytotoxic ef...

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“…TcdA and TcdB have four functional domains 100,106 : an amino-terminal glucosyltransferase domain (GTD), an autoprotease domain (APD), a pore-forming and delivery domain, and the combined repetitive oligopeptides (CROPS) domain, which extends from around residue 1,830 to the carboxyl terminus (FIG. 5a,b).…”

Section: Mechanisms/pathophysiologymentioning

confidence: 99%

“…5a,b). A combination of electron microscopy and X-ray crystallography studies has revealed the structural organization of these domains in TcdA and suggests that the structure of TcdB is similar 106,107 . TcdA and TcdB enter cells via receptor-mediated endocytosis 108 .…”

Section: Mechanisms/pathophysiologymentioning

confidence: 99%

Clostridium difficile infection

Smits

Lyras

Lacy

et al. 2016

Nat Rev Dis Primers

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Infection of the colon with the Gram-positive bacterium Clostridium difficile is potentially life threatening, especially in elderly people and in patients who have dysbiosis of the gut microbiota following antimicrobial drug exposure. C. difficile is the leading cause of health-care-associated infective diarrhoea. The life cycle of C. difficile is influenced by antimicrobial agents, the host immune system, and the host microbiota and its associated metabolites. The primary mediators of inflammation in C. difficile infection (CDI) are large clostridial toxins, toxin A (TcdA) and toxin B (TcdB), and, in some bacterial strains, the binary toxin CDT. The toxins trigger a complex cascade of host cellular responses to cause diarrhoea, inflammation and tissue necrosis — the major symptoms of CDI. The factors responsible for the epidemic of some C. difficile strains are poorly understood. Recurrent infections are common and can be debilitating. Toxin detection for diagnosis is important for accurate epidemiological study, and for optimal management and prevention strategies. Infections are commonly treated with specific antimicrobial agents, but faecal microbiota transplants have shown promise for recurrent infections. Future biotherapies for C. difficile infections are likely to involve defined combinations of key gut microbiota.

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Crystal structure of Clostridium difficile toxin A

Cited by 88 publications

References 46 publications

Dietary zinc alters the microbiota and decreases resistance to Clostridium difficile infection

Dietary zinc alters the microbiota and decreases resistance to Clostridium difficile infection

Clostridium difficile Toxins TcdA and TcdB Cause Colonic Tissue Damage by Distinct Mechanisms

Clostridium difficile infection

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