Search Results (15)

Background: Helicobacter hepaticus (H. hepaticus) has been demonstrated to have clinical relevance to the development of colitis in rodents. H. hepaticus produces cytolethal distending toxins (CDTs), which are identified as the most important virulence factors to the pathogenicity of CDT-producing bacteria in animals. However, the precise relationship between CDTs of H. hepaticus and intestinal barrier dysfunction remains unclear. The objective of the present study was to ascertain the impact of CdtB, the active subunit of CDTs, on the colonic mucosal barrier during H. hepaticus infection. Materials and Methods: We investigated the infection of male BALB/c mice, intestinal organoids, and IEC-6 cell monolayers by H. hepaticus or CdtB-deficient H. hepaticus (ΔCdtB). A comprehensive histopathological examination was conducted, encompassing the assessment of H. hepaticus colonization, the levels of mRNA expression for inflammatory cytokines, the expression levels of tight junction proteins, and the related signaling pathways. Results: The results demonstrate that the presence of ΔCdtB led to a mitigation of the symptoms associated with H. hepaticus-induced colitis, as evidenced by colon length shortening and the colon histological inflammation score. In addition, the levels of pro-inflammatory cytokines were reduced in the ΔCdtB group. Moreover, a downward trend was observed in the phosphorylation levels of STAT3 and nuclear factor-κB (p65). In vitro, the presence of H. hepaticus resulted in a reduction in the expression of tight junction (TJ) markers (ZO-1 and occludin) and an impairment of the F-actin structure in either the intestinal epithelium or intestinal organoids. However, these effects were reversed by CdtB deletion. Concurrently, both ROS levels and apoptosis levels were found to be significantly reduced in cells treated with the ΔCdtB strain. Mechanistically, myosin light chain kinase (MLCK) activation was observed in the H. hepaticus-infected group in vivo, whereas the MLCK inhibitor ML-7 was found to reverse the CdtB-induced alterations in TJ proteins in IEC6 cells. Conclusions: The collective findings demonstrate that CdtB plays a pivotal role in the H. hepaticus-induced colonic mucosal barrier. This is achieved through the regulation of TJs via the MLCK/pMLC2 signaling pathway, which is linked to elevations in oxidative stress and inflammation within intestinal epithelial cells. Full article

Alcohol consumption is associated with oxidative stress and an increased risk of carcinoma of the upper aero-digestive tract (UADT). Recently, it has been found that some microorganisms in the human oral cavity may locally metabolize ethanol, forming acetaldehyde, a carcinogenic metabolite of alcohol. In a cohort of patients first visited for UADT cancers, we estimated their alcohol consumption by measuring Ethyl Glucuronide/EtG (a long-lasting metabolite of ethanol) in the hair and carbohydrate-deficient transferrin/CDT (short-term index of alcohol intake) in the serum. Moreover, we analyzed, by culture-based methods, the presence of Neisseria subflava, Streptococcus mitis, Candida albicans, and glabrata (microorganisms generating acetaldehyde) in the oral cavity. According to the EtG values, we correlated drinking alcohol with endogenous oxidative stress and the investigated microorganism’s presence. We found that 55% of heavy drinkers presented microorganisms generating acetaldehyde locally. Moreover, we found that the presence of oral acetaldehyde-producing bacteria correlates with increased oxidative stress compared to patients without such bacteria. As for the study of alcohol dehydrogenase gene polymorphisms (the enzyme that transforms alcohol to acetaldehyde), we found that only the “CGTCGTCCC” haplotype was more frequent in the general population than in carcinoma patients. This pilot study suggests the importance of estimating alcohol consumption (EtG), the presence of bacteria producing acetaldehyde, and oxidative stress as risk factors for the onset of oral carcinomas. Full article

Cytolethal distending toxin (CDT) is produced by a range of Gram-negative pathogenic bacteria such as Campylobacter jejuni. CDT represents an important virulence factor that is a heterotrimeric complex composed of CdtA, CdtB, and CdtC. CdtA and CdtC constitute regulatory subunits whilst CdtB acts as the catalytic subunit exhibiting phosphatase and DNase activities, resulting in cell cycle arrest and cell death. Extracellular vesicle (EV) secretion is an evolutionarily conserved process that is present throughout all kingdoms. Mammalian EVs play important roles in regular cell-to-cell communications but can also spread pathogen- and host-derived molecules during infections to alter immune responses. Here, we demonstrate that CDT targets the endo-lysosomal compartment, partially evading lysosomal degradation and exploiting unconventional secretion (EV release), which is largely involved in bacterial infections. CDT-like effects are transferred by Caco-2 cells to uninfected heterologous U937 and homologous Caco-2 cells. The journey of EVs derived from CDT-treated Caco-2 cells is associated with both intestinal and myeloid tumour cells. EV release represents the primary route of CDT dissemination, revealing an active toxin as part of the cargo. We demonstrated that bacterial toxins could represent suitable tools in cancer therapy, highlighting both the benefits and limitations. The global cell response involves a moderate induction of apoptosis and autophagic features may play a protective role against toxin-induced cell death. EVs from CDT-treated Caco-2 cells represent reliable CDT carriers, potentially suitable in colorectal cancer treatments. Our data present a potential bacterial-related biotherapeutic supporting a multidrug anticancer protocol. Full article

The Cytolethal Distending Toxin (CDT) is produced by many Gram-negative pathogenic bacteria responsible for major foodborne diseases worldwide. CDT induces DNA damage and cell cycle arrest in host-cells, eventually leading to senescence or apoptosis. According to structural and sequence comparison, the catalytic subunit CdtB is suggested to possess both nuclease and phosphatase activities, carried by a single catalytic site. However, the impact of each activity on cell-host toxicity is yet to be characterized. Here, we analyze the consequences of cell exposure to different CDT mutated on key CdtB residues, focusing on cell viability, cell cycle defects, and DNA damage induction. A first class of mutant, devoid of any activity, targets putative catalytic (H160A), metal binding (D273R), and DNA binding residues (R117A-R144A-N201A). The second class of mutants (A163R, F156-T158, and the newly identified G114T), which gathers mutations on residues potentially involved in lipid substrate binding, has only partially lost its toxic effects. However, their defects are alleviated when CdtB is artificially introduced inside cells, except for the F156-T158 double mutant that is defective in nuclear addressing. Therefore, our data reveal that CDT toxicity is mainly correlated to CdtB nuclease activity, whereas phosphatase activity may probably be involved in CdtB intracellular trafficking. Full article

Bacterial genotoxins (BTGX) induce DNA damage, which results in senescence or apoptosis of the target cells if not properly repaired. Three BTGXs have been identified: the cytolethal distending toxin (CDT) family produced by several Gram-negative bacteria, the typhoid toxin produced by several Salmonella enterica serovars, and colibactin, a peptide-polyketide, produced mainly by the phylogenetic group B2 Escherichia coli. The cellular responses induced by BTGXs resemble those of well-characterized carcinogenic agents, and several lines of evidence indicate that bacteria carrying genotoxin genes can contribute to tumor development under specific circumstances. Given their unusual mode of action, it is still enigmatic why these effectors have been acquired by microbes and what is their role in the context of the biology of the producing bacterium, since it is unlikely that their primary purpose is to induce/promote cancer in the mammalian host. In this review, we will discuss the possibility that the DNA damage induced by BTGX modulates the host immune response, acting as immunomodulator, leading to the establishment of a suitable niche for the producing bacterium. We will further highlight open questions that remain to be solved regarding the biology of this unusual family of bacterial toxins. Full article

The Cytolethal Distending Toxin (CDT) is a bacterial virulence factor produced by several Gram-negative pathogenic bacteria. These bacteria, found in distinct niches, cause diverse infectious diseases and produce CDTs differing in sequence and structure. CDTs have been involved in the pathogenicity of the associated bacteria by promoting persistent infection. At the host-cell level, CDTs cause cell distension, cell cycle block and DNA damage, eventually leading to cell death. All these effects are attributable to the catalytic CdtB subunit, but its exact mode of action is only beginning to be unraveled. Sequence and 3D structure analyses revealed similarities with better characterized proteins, such as nucleases or phosphatases, and it has been hypothesized that CdtB exerts a biochemical activity close to those enzymes. Here, we review the relationships that have been established between CdtB structure and function, particularly by mutation experiments on predicted key residues in different experimental systems. We discuss the relevance of these approaches and underline the importance of further study in the molecular mechanisms of CDT toxicity, particularly in the context of different pathological conditions. Full article

Binary toxins are produced by several pathogenic bacteria. Examples are the C2 toxin from Clostridium botulinum, the iota toxin from Clostridium perfringens, and the CDT from Clostridium difficile. All these binary toxins have ADP-ribosyltransferases (ADPRT) as their enzymatically active component that modify monomeric actin in their target cells. The binary C2 toxin was intensively described as a tool for intracellular delivery of allogenic ADPRTs. Here, we firstly describe the binary toxin CDT from C. difficile as an effective tool for heterologous intracellular delivery. Even 60 kDa glucosyltransferase domains of large clostridial glucosyltransferases can be delivered into cells. The glucosyltransferase domains of five tested large clostridial glucosyltransferases were successfully introduced into cells as chimeric fusions to the CDTa adapter domain (CDTaN). Cell uptake was demonstrated by the analysis of cell morphology, cytoskeleton staining, and intracellular substrate glucosylation. The fusion toxins were functional only when the adapter domain of CDTa was N-terminally located, according to its native orientation. Thus, like other binary toxins, the CDTaN/b system can be used for standardized delivery systems not only for bacterial ADPRTs but also for a variety of bacterial glucosyltransferase domains. Full article

Cytolethal distending toxin (CDT) is found in Gram-negative bacteria, especially in certain Proteobacteria such as the Pasteurellaceae family, including Haemophilus ducreyi and Aggregatibacter (Actinobacillus) actinomycetemcomitans, in the Enterobacteriaceae family and the Campylobacterales order, including the Campylobacter and Helicobacter species. In vitro and in vivo studies have clearly shown that this toxin has a strong effect on cellular physiology (inflammation, immune response modulation, tissue damage). Some works even suggest a potential involvement of CDT in cancers. In this review, we will discuss these different aspects. Full article

The cytolethal distending toxin (CDT) is a well characterized bacterial genotoxin encoded by several Gram-negative bacteria, including Salmonella enterica (S. enterica). The CDT produced by Salmonella (S-CDT) differs from the CDT produced by other bacteria, as it utilizes subunits with homology to the pertussis and subtilase toxins, in place of the traditional CdtA and CdtC subunits. Previously, S-CDT was thought to be a unique virulence factor of S. enterica subspecies enterica serotype Typhi, lending to its classification as the “typhoid toxin.” Recently, this important virulence factor has been identified and characterized in multiple nontyphoidal Salmonella (NTS) serotypes as well. The significance of S-CDT in salmonellosis with regards to the: (i) distribution of S-CDT encoding genes among NTS serotypes, (ii) contributions to pathogenicity, (iii) regulation of S-CDT expression, and (iv) the public health implication of S-CDT as it relates to disease severity, are reviewed here. Full article

The pathogenic bacteria Clostridium difficile, Clostridium perfringens and Clostridium botulinum produce the binary actin ADP-ribosylating toxins CDT, iota and C2, respectively. These toxins are composed of a transport component (B) and a separate enzyme component (A). When both components assemble on the surface of mammalian target cells, the B components mediate the entry of the A components via endosomes into the cytosol. Here, the A components ADP-ribosylate G-actin, resulting in depolymerization of F-actin, cell-rounding and eventually death. In the present study, we demonstrate that 4-bromobenzaldehyde N-(2,6-dimethylphenyl)semicarbazone (EGA), a compound that protects cells from multiple toxins and viruses, also protects different mammalian epithelial cells from all three binary actin ADP-ribosylating toxins. In contrast, EGA did not inhibit the intoxication of cells with Clostridium difficile toxins A and B, indicating a possible different entry route for this toxin. EGA does not affect either the binding of the C2 toxin to the cells surface or the enzyme activity of the A components of CDT, iota and C2, suggesting that this compound interferes with cellular uptake of the toxins. Moreover, for C2 toxin, we demonstrated that EGA inhibits the pH-dependent transport of the A component across cell membranes. EGA is not cytotoxic, and therefore, we propose it as a lead compound for the development of novel pharmacological inhibitors against clostridial binary actin ADP-ribosylating toxins. Full article

Bacterial genotoxins are unique among bacterial toxins as their molecular target is DNA. The consequence of intoxication or infection is induction of DNA breaks that, if not properly repaired, results in irreversible cell cycle arrest (senescence) or death of the target cells. At present, only three bacterial genotoxins have been identified. Two are protein toxins: the cytolethal distending toxin (CDT) family produced by a number of Gram-negative bacteria and the typhoid toxin produced by Salmonella enterica serovar Typhi. The third member, colibactin, is a peptide-polyketide genotoxin, produced by strains belonging to the phylogenetic group B2 of Escherichia coli. This review will present the cellular effects of acute and chronic intoxication or infection with the genotoxins-producing bacteria. The carcinogenic properties and the role of these effectors in the context of the host-microbe interaction will be discussed. We will further highlight the open questions that remain to be solved regarding the biology of this unusual family of bacterial toxins. Full article

The cytolethal distending toxin (Cdt) is a heterotrimeric holotoxin produced by a diverse group of Gram-negative pathogenic bacteria. The Cdts expressed by the members of this group comprise a subclass of the AB toxin superfamily. Some AB toxins have hijacked the retrograde transport pathway, carried out by the Golgi apparatus and endoplasmic reticulum (ER), to translocate to cytosolic targets. Those toxins have been used as tools to decipher the roles of the Golgi and ER in intracellular transport and to develop medically useful delivery reagents. In comparison to the other AB toxins, the Cdt exhibits unique properties, such as translocation to the nucleus, that present specific challenges in understanding the precise molecular details of the trafficking pathway in mammalian cells. The purpose of this review is to present current information about the mechanisms of uptake and translocation of the Cdt in relation to standard concepts of endocytosis and retrograde transport. Studies of the Cdt intoxication process to date have led to the discovery of new translocation pathways and components and most likely will continue to reveal unknown features about the mechanisms by which bacterial proteins target the mammalian cell nucleus. Insight gained from these studies has the potential to contribute to the development of novel therapeutic strategies. Full article

The cytolethal distending toxin (CDT) is produced by many pathogenic Gram-negative bacteria and is considered as a virulence factor. In human cells, CDT exposure leads to a unique cytotoxicity associated with a characteristic cell distension and induces a cell cycle arrest dependent on the DNA damage response (DDR) triggered by DNA double-strand breaks (DSBs). CDT has thus been classified as a cyclomodulin and a genotoxin. Whereas unrepaired damage can lead to cell death, effective, but improper repair may be detrimental. Indeed, improper repair of DNA damage may allow cells to resume the cell cycle and induce genetic instability, a hallmark in cancer. In vivo, CDT has been shown to induce the development of dysplastic nodules and to lead to genetic instability, defining CDT as a potential carcinogen. It is therefore important to characterize the outcome of the CDT-induced DNA damage and the consequences for intoxicated cells and organisms. Here, we review the latest results regarding the host cell response to CDT intoxication and focus on DNA damage characteristics, cell cycle modulation and cell outcomes. Full article

The cytolethal distending toxins (CDTs) are a family of exotoxins produced by a wide range of Gram-negative bacteria. They are known for causing genotoxic stress to the cell, resulting in growth arrest and eventually apoptotic cell death. Nevertheless, there is evidence that CDTs can also perturb the innate immune responses, by regulating inflammatory cytokine production and molecular mediators of bone remodeling in various cell types. These cellular and molecular events may in turn have an effect in enhancing local inflammation in diseases where CDT-producing bacteria are involved, such as Aggregatibacter actinomycetemcomitans, Haemophilus ducreyi, Campylobacter jejuni and Helicobacter hepaticus. One special example is the induction of pathological bone destruction in periodontitis. The opportunistic oral pathogen Aggregatibatcer actinoycemetemcomitans, which is involved in the aggressive form of the disease, can regulate the molecular mechanisms of bone remodeling in a manner that favors bone resorption, with the potential involvement of its CDT. The present review provides an overview of all known to-date inflammatory or bone remodeling responses of CDTs produced by various bacterial species, and discusses their potential contribution to the pathogenesis of the associated diseases. Full article

The cytolethal distending toxins (CDTs), produced by a variety of Gram-negative pathogenic bacteria, are the first bacterial genotoxins described, since they cause DNA damage in the target cells. CDT is an A-B₂ toxin, where the CdtA and CdtC subunits are required to mediate the binding on the surface of the target cells, allowing internalization of the active CdtB subunit, which is functionally homologous to the mammalian deoxyribonuclease I. The nature of the surface receptor is still poorly characterized, however binding of CDT requires intact lipid rafts, and its internalization occurs via dynamin-dependent endocytosis. The toxin is retrograde transported through the Golgi complex and the endoplasmic reticulum, and subsequently translocated into the nuclear compartment, where it exerts the toxic activity. Cellular intoxication induces DNA damage and activation of the DNA damage responses, which results in arrest of the target cells in the G1 and/or G2 phases of the cell cycle and activation of DNA repair mechanisms. Cells that fail to repair the damage will senesce or undergo apoptosis. This review will focus on the well-characterized aspects of the CDT biology and discuss the questions that still remain unanswered. Full article