title: Cholesterol-rich membrane microdomains mediate cell cycle arrest induced by Actinobacillus actinomycetemcomitans cytolethal-distending toxin.
abstract: We have previously shown that Actinobacillus actinomycetemcomitans cytolethal-distending toxin (Cdt) is a potent immunosuppressive agent that induces G2/M arrest in human lymphocytes. In this study, we explored the possibility that Cdt-mediated immunotoxicity involves lipid membrane microdomains. We first determined that following treatment of Jurkat cells with Cdt holotoxin all three Cdt subunits localize to these microdomains. Laser confocal microscopy was employed to colocalize the subunits with GM1-enriched membrane regions which are characteristic of membrane rafts. Western blot analysis of isolated lipid rafts also demonstrated the presence of Cdt peptides. Cholesterol depletion, using methyl beta-cyclodextrin, protected cells from the ability of the Cdt holotoxin to induce G2 arrest. Moreover, cholesterol depletion reduced the ability of the toxin to associate with Jurkat cells. Thus, lipid raft integrity is vital to the action of Cdt on host cells. The implications of our observations with respect to Cdt mode of action are discussed.
authors.affiliation: Department of Biochemistry, University of Pennsylvania School of Dental Medicine, Philadelphia, PA, USA. battagli@biochem.dental.upenn.edu
authors.forename: Kathleen
authors.initials: K
authors.lastname: Boesze-Battaglia
authors.forename: Dave
authors.initials: D
authors.lastname: Besack
authors.forename: Terry
authors.initials: T
authors.lastname: McKay
authors.forename: Ali
authors.initials: A
authors.lastname: Zekavat
authors.forename: Linda
authors.initials: L
authors.lastname: Otis
authors.forename: Kelly
authors.initials: K
authors.lastname: Jordan-Sciutto
authors.forename: Bruce J
authors.initials: BJ
authors.lastname: Shenker
mesh_terms: D016976:Aggregatibacter actinomycetemcomitans / Q000378:metabolism*; D015703:Antigens, CD / Q000378:metabolism; D001427:Bacterial Toxins / Q000494:pharmacology*; D002453:Cell Cycle; D002784:Cholesterol / Q000378:metabolism*; D016537:Gangliosidosis, GM1 / Q000378:metabolism; D006801:Humans; D019169:Jurkat Cells; D021962:Membrane Microdomains / Q000502:physiology*; D021122:Protein Subunits / Q000494:pharmacology; D011990:Receptors, Transferrin / Q000378:metabolism; D047392:beta-Cyclodextrins / Q000494:pharmacology
chemical_list: D015703:Antigens, CD; D001427:Bacterial Toxins; C068126:CD71 antigen; D021122:Protein Subunits; D011990:Receptors, Transferrin; D047392:beta-Cyclodextrins; C447141:cytolethal distending toxin, Actinobacillus actinomycetemcomitans; C108732:methyl-beta-cyclodextrin; D002784:Cholesterol