title: Cytolethal Distending Toxin FromCampylobacter jejuniRequires the Cytoskeleton for Toxic Activity.
abstract: BACKGROUND
  Campylobacter jejuniis one of the major causes of infectious diarrhea worldwide. The distending cytolethal toxin (CDT) ofCampylobacterspp. interferes with normal cell cycle progression. This toxic effect is considered a result of DNase activity that produces chromosomal DNA damage. To perform this event, the toxin must be endocytosed and translocated to the nucleus.
  
  
  OBJECTIVE
  The aim of this study was to evaluate the role of the cytoskeleton in the translocation of CDT to the nucleus.
  
  
  METHODS
  Campylobacter jejuniATCC 33291 and seven isolates donated from Instituto de Biotecnologia were used in this study. The presence of CDT genes inC. jejunistrains was determined by PCR. To evaluate the effect of CDT, HeLa cells were treated with bacterial lysate, and the damage and morphological changes were analyzed by microscopy, immunofluorescence staining, and flow cytometry. To evaluate the role of the cytoskeleton, HeLa cells were treated with either latrunculin A or by nocodazole and analyzed by microscopy, flow cytometry, and immunoquantification (ELISA).
  
  
  RESULTS
  The results obtained showed that the eight strains ofC. jejuni, including the reference strain, had the ability to produce the toxin. Usage of latrunculin A and nocodazole, two cytoskeletal inhibitors, blocked the toxic effect in cells treated with the toxin. This phenomenon was evident in flow cytometry analysis and immunoquantification of Cdc2-phosphorylated.
  
  
  CONCLUSIONS
  This work showed that the cytotoxic activity of theC. jejuniCDT is dependent on its endocytosis. The alteration in the microtubules and actin filaments caused a blockage transit of the toxin, preventing it from reaching the nucleus of the cell, as well as preventing DNA fragmentation and alteration of the cell cycle. The CDT toxin appears to be an important element for the pathogenesis of campylobacteriosis, since all clinical isolates showed the presence ofcdtA,cdtBandcdtCgenes.
authors.affiliation: Departamento de Producción Agrícola y Animal, and Doctorado en Ciencias Biológicas y de la Salud, Universidad Autónoma Metropolitana-Xochimilco, México City, México.
authors.forename: Estela T
authors.initials: ET
authors.lastname: Méndez-Olvera
authors.affiliation: Departamento de Atención a la Salud, Universidad Autónoma Metropolitana-Xochimilco, México City, México.
authors.forename: Jaime A
authors.initials: JA
authors.lastname: Bustos-Martínez
authors.affiliation: Departamento de Microbiología y Parasitología-Facultad de Medicina, Universidad Nacional Autónoma de México, México City, México.
authors.forename: Yolanda
authors.initials: Y
authors.lastname: López-Vidal
authors.affiliation: Departamento de Microbiología e Inmunología-Facultad de Medicina Veterinaria y Zootecnia, Universidad Nacional Autónoma de México, México City, México.
authors.forename: Antonio
authors.initials: A
authors.lastname: Verdugo-Rodríguez
authors.affiliation: Departamento de Producción Agrícola y Animal, Universidad Autónoma Metropolitana-Xochimilco, México City, México.
authors.forename: Daniel
authors.initials: D
authors.lastname: Martínez-Gómez
keywords: CDC2 Protein Kinase; Campylobacter jejuni; Cytolethal Distending Toxin; Cytoskeleton; Latrunculin A; Nocodazole