scholarly journals CdtA, CdtB, and CdtC Form a Tripartite Complex That Is Required for Cytolethal Distending Toxin Activity

2001 ◽  
Vol 69 (7) ◽  
pp. 4358-4365 ◽  
Author(s):  
Marı́a Lara-Tejero ◽  
Jorge E. Galán
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Campylobacter Jejuni ◽  
Bacterial Pathogens ◽  
Target Cells ◽  
Cytolethal Distending Toxin ◽  
Toxic Activity ◽  
Cellular Toxicity ◽  
B Subunit ◽  
Cycle Arrest

ABSTRACT Campylobacter jejuni encodes a cytolethal distending toxin (CDT) that causes cells to arrest in the G2/M transition phase of the cell cycle. Highly related toxins are also produced by other important bacterial pathogens. CDT activity requires the function of three genes: cdtA, cdtB, andcdtC. Recent studies have established that CdtB is the active subunit of CDT, exerting its effect as a nuclease that damages the DNA and triggers cell cycle arrest. Microinjection of CdtB into target cells led to G2/M arrest and cytoplasmic distention, in a manner indistinguishable from that caused by CDT treatment. Despite this progress, nothing is known about the composition of the CDT holotoxin or the function of CdtA and CdtC. We show here that, when applied individually, purified CdtA, CdtB, or CdtC does not exhibit toxic activity. In contrast, CdtA, CdtB, and CdtC when combined, interact with one another to form an active tripartite holotoxin that exhibits full cellular toxicity. CdtA has a domain that shares similarity with the B chain of ricin-related toxins. We therefore proposed that CDT is a tripartite toxin composed of CdtB as the enzymatically active subunit and of CdtA and CdtC as the heterodimeric B subunit required for the delivery of CdtB.

scholarly journals The Aggregatibacter actinomycetemcomitans Cytolethal Distending Toxin Active Subunit CdtB Contains a Cholesterol Recognition Sequence Required for Toxin Binding and Subunit Internalization

2015 ◽  
Vol 83 (10) ◽  
pp. 4042-4055 ◽  
Author(s):  
Kathleen Boesze-Battaglia ◽  
Lisa P. Walker ◽  
Ali Zekavat ◽  
Mensur Dlakić ◽  
Monika Damek Scuron ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Consensus Sequence ◽  
Aggregatibacter Actinomycetemcomitans ◽  
Jurkat Cells ◽  
Membrane Microdomains ◽  
Target Cells ◽  
Cytolethal Distending Toxin ◽  
Toxin Binding ◽  
Cycle Arrest

Induction of cell cycle arrest in lymphocytes following exposure to theAggregatibacter actinomycetemcomitanscytolethal distending toxin (Cdt) is dependent upon the integrity of lipid membrane microdomains. Moreover, we have previously demonstrated that the association of Cdt with target cells involves the CdtC subunit which binds to cholesterol via a cholesterol recognition amino acid consensus sequence (CRAC site). In this study, we demonstrate that the active Cdt subunit, CdtB, also is capable of binding to large unilamellar vesicles (LUVs) containing cholesterol. Furthermore, CdtB binding to cholesterol involves a similar CRAC site as that demonstrated for CdtC. Mutation of the CRAC site reduces binding to model membranes as well as toxin binding and CdtB internalization in both Jurkat cells and human macrophages. A concomitant reduction in Cdt-induced toxicity was also noted, indicated by reduced cell cycle arrest and apoptosis in Jurkat cells and a reduction in the proinflammatory response in macrophages (interleukin 1β [IL-1β] and tumor necrosis factor alpha [TNF-α] release). Collectively, these observations indicate that membrane cholesterol serves as an essential ligand for both CdtC and CdtB and, further, that this binding is necessary for both internalization of CdtB and subsequent molecular events leading to intoxication of cells.

scholarly journals The Active Subunit of the Cytolethal Distending Toxin, CdtB, Derived From Both Haemophilus ducreyi and Campylobacter jejuni Exhibits Potent Phosphatidylinositol-3,4,5-Triphosphate Phosphatase Activity

2021 ◽  
Vol 11 ◽  
Author(s):  
Grace Huang ◽  
Kathleen Boesze-Battaglia ◽  
Lisa P. Walker ◽  
Ali Zekavat ◽  
Zachary P. Schaefer ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Phosphatase Activity ◽  
Campylobacter Jejuni ◽  
Human Lymphocytes ◽  
Haemophilus Ducreyi ◽  
Cell Protein ◽  
Cytolethal Distending Toxin ◽  
Induce Cell Cycle Arrest ◽  
Cycle Arrest

Human lymphocytes exposed to Aggregatibacter actinomycetemcomitans (Aa) cytolethal distending toxin (Cdt) undergo cell cycle arrest and apoptosis. In previous studies, we demonstrated that the active Cdt subunit, CdtB, is a potent phosphatidylinositol (PI) 3,4,5-triphosphate phosphatase. Moreover, AaCdt-treated cells exhibit evidence of PI-3-kinase (PI-3K) signaling blockade characterized by reduced levels of PIP3, pAkt, and pGSK3β. We have also demonstrated that PI-3K blockade is a requisite of AaCdt-induced toxicity in lymphocytes. In this study, we extended our observations to include assessment of Cdts from Haemophilus ducreyi (HdCdt) and Campylobacter jejuni (CjCdt). We now report that the CdtB subunit from HdCdt and CjCdt, similar to that of AaCdt, exhibit potent PIP3 phosphatase activity and that Jurkat cells treated with these Cdts exhibit PI-3K signaling blockade: reduced levels of pAkt and pGSK3β. Since non-phosphorylated GSK3β is the active form of this kinase, we compared Cdts for dependence on GSK3β activity. Two GSK3β inhibitors were employed, LY2090314 and CHIR99021; both inhibitors blocked the ability of Cdts to induce cell cycle arrest. We have previously demonstrated that AaCdt induces increases in the CDK inhibitor, p21CIP1/WAF1, and, further, that this was a requisite for toxin-induced cell death via apoptosis. We now demonstrate that HdCdt and CjCdt also share this requirement. It is also noteworthy that p21CIP1/WAF1 was not involved in the ability of the three Cdts to induce cell cycle arrest. Finally, we demonstrate that, like AaCdt, HdCdt is dependent upon the host cell protein, cellugyrin, for its toxicity (and presumably internalization of CdtB); CjCdt was not dependent upon this protein. The implications of these findings as they relate to Cdt’s molecular mode of action are discussed.

scholarly journals Campylobacter jejuni Cytolethal Distending Toxin Causes a G2-Phase Cell Cycle Block

1998 ◽  
Vol 66 (5) ◽  
pp. 1934-1940 ◽  
Author(s):  
Chris A. Whitehouse ◽  
Paul B. Balbo ◽  
Everett C. Pesci ◽  
Daniel L. Cottle ◽  
Peter M. Mirabito ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Campylobacter Jejuni ◽  
Hela Cells ◽  
Chromatin Condensation ◽  
Phase Cell ◽  
Specific Cell ◽  
Cytolethal Distending Toxin ◽  
Cycle Arrest ◽  
M Phase

ABSTRACT Cytolethal distending toxin (CDT) from the diarrheagenic bacteriumCampylobacter jejuni was shown to cause a rapid and specific cell cycle arrest in HeLa and Caco-2 cells. Within 24 h of treatment, CDT caused HeLa cells to arrest with a 4N DNA content, indicative of cells in G2 or early M phase. Immunofluorescence studies indicated that the arrested cells had not entered M phase, since no evidence of tubulin reorganization or chromatin condensation was visible. CDT treatment was also shown to cause HeLa cells to accumulate the inactive, tyrosine-phosphorylated form of CDC2. These results indicated that CDT treatment results in a failure to activate CDC2, which leads to cell cycle arrest in G2. This mechanism of action is novel for a bacterial toxin and provides a model for the generation of diarrheal disease byC. jejuni and other diarrheagenic bacteria that produce CDT.

scholarly journals Cholesterol Depletion Reduces Entry of Campylobacter jejuni Cytolethal Distending Toxin and Attenuates Intoxication of Host Cells

2011 ◽  
Vol 79 (9) ◽  
pp. 3563-3575 ◽  
Author(s):  
Chia-Der Lin ◽  
Cheng-Kuo Lai ◽  
Yu-Hsin Lin ◽  
Jer-Tsong Hsieh ◽  
Yu-Ting Sing ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Lipid Rafts ◽  
Campylobacter Jejuni ◽  
Host Cells ◽  
Membrane Cholesterol ◽  
Cholesterol Depletion ◽  
Cytolethal Distending Toxin ◽  
Content Type ◽  
Cycle Arrest

ABSTRACTCampylobacter jejuniis a common cause of pediatric diarrhea worldwide. Cytolethal distending toxin, produced byCampylobacter jejuni, is a putative virulence factor that induces cell cycle arrest and apoptosis in eukaryotic cells. Cellular cholesterol, a major component of lipid rafts, has a pivotal role in regulating signaling transduction and protein trafficking as well as pathogen internalization. In this study, we demonstrated that cell intoxication byCampylobacter jejunicytolethal distending toxin is through the association of cytolethal distending toxin subunits and membrane cholesterol-rich microdomains. Cytolethal distending toxin subunits cofractionated with detergent-resistant membranes, while the distribution reduced upon the depletion of cholesterol, suggesting that cytolethal distending toxin subunits are associated with lipid rafts. The disruption of cholesterol using methyl-β-cyclodextrin not only reduced the binding activity of cytolethal distending toxin subunits on the cell membrane but also impaired their delivery and attenuated toxin-induced cell cycle arrest. Accordingly, cell intoxication by cytolethal distending toxin was restored by cholesterol replenishment. These findings suggest that membrane cholesterol plays a critical role in theCampylobacter jejunicytolethal distending toxin-induced pathogenesis of host cells.

Delivery of cytolethal distending toxin B induces cell cycle arrest and apoptosis in gingival squamous cell carcinoma in vitro

2004 ◽  
Vol 112 (5) ◽  
pp. 445-451 ◽  
Author(s):  
Kozo Yamamoto ◽  
Kazuhiro Tominaga ◽  
Miho Sukedai ◽  
Toshinori Okinaga ◽  
Kenjiro Iwanaga ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Squamous Cell Carcinoma ◽  
Cell Carcinoma ◽  
Cell Cycle Arrest ◽  
Squamous Cell ◽  
Cytolethal Distending Toxin ◽  
Toxin B ◽  
Cycle Arrest ◽  
Gingival Squamous Cell Carcinoma

scholarly journals Escherichia coli CdtB Mediates Cytolethal Distending Toxin Cell Cycle Arrest

2001 ◽  
Vol 69 (5) ◽  
pp. 3418-3422 ◽  
Author(s):  
Cherilyn Elwell ◽  
Kinlin Chao ◽  
Kamlesh Patel ◽  
Lawrence Dreyfus
Keyword(s):  
Escherichia Coli ◽  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Hela Cells ◽  
Functional Relationship ◽  
Type I ◽  
Cytolethal Distending Toxin ◽  
E Coli ◽  
Cycle Arrest ◽  
Dna Nicking

ABSTRACT We previously reported that the CdtB polypeptide ofEscherichia coli cytolethal distending toxin (CDT) shares significant pattern-specific homology with mammalian type I DNases. In addition, the DNase-related residues of CdtB are required for cellular toxicity. Here we demonstrate that purified CdtB converts supercoiled plasmid DNA to relaxed and linear forms and promotes cell cycle arrest when combined with an E. coli extract containing CdtA and CdtC. CdtB alone had no effect on HeLa cells, however; introduction of the polypeptide into HeLa cells by electroporation resulted in cellular distension, chromatin fragmentation, and cell cycle arrest, all of which are consequences of CDT action. In contrast to these findings, purified CdtBH154A lacked both DNA-nicking and cell cycle arrest activities. These results suggest a functional relationship between DNase-related residues in CdtB and CDT biological activity.

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