title: Structure-dependent pseudoreceptor intracellular traffic of adamantyl globotriaosyl ceramide mimics.
abstract: The verotoxin (VT) (Shiga toxin) receptor globotriaosyl ceramide (Gb(3)), mediates VT1/VT2 retrograde transport to the endoplasmic reticulum (ER) for cytosolic A subunit access to inhibit protein synthesis. Adamantyl Gb(3) is an amphipathic competitive inhibitor of VT1/VT2 Gb(3) binding. However, Gb(3)-negative VT-resistant CHO/Jurkat cells incorporate adaGb(3) to become VT1/VT2-sensitive. CarboxyadaGb(3), urea-adaGb(3), and hydroxyethyl adaGb(3), preferentially bound by VT2, also mediate VT1/VT2 cytotoxicity. VT1/VT2 internalize to early endosomes but not to Golgi/ER. AdabisGb(3) (two deacyl Gb(3)s linked to adamantane) protects against VT1/VT2 more effectively than adaGb(3) without incorporating into Gb(3)-negative cells. AdaGb(3) (but not hydroxyethyl adaGb(3)) incorporation into Gb(3)-positive Vero cells rendered punctate cell surface VT1/VT2 binding uniform and subverted subsequent Gb(3)-dependent retrograde transport to Golgi/ER to render cytotoxicity (reduced for VT1 but not VT2) brefeldin A-resistant. VT2-induced vacuolation was maintained in adaGb(3)-treated Vero cells, but vacuolar membrane VT2 was lost. AdaGb(3) destabilized membrane cholesterol and reduced Gb(3) cholesterol stabilization in phospholipid liposomes. Cholera toxin GM1-mediated Golgi/ER targeting was unaffected by adaGb(3). We demonstrate the novel, lipid-dependent, pseudoreceptor function of Gb(3) mimics and their structure-dependent modulation of endogenous intracellular Gb(3) vesicular traffic.
authors.affiliation: Research Institute, Program in Molecular Structure and Function, Hospital for Sick Children, Toronto, Ontario M5G 1X8, Canada.
authors.forename: Mitsumasa
authors.initials: M
authors.lastname: Saito
authors.forename: Murugespillai
authors.initials: M
authors.lastname: Mylvaganum
authors.forename: Patty
authors.initials: P
authors.lastname: Tam
authors.forename: Anton
authors.initials: A
authors.lastname: Novak
authors.forename: Beth
authors.initials: B
authors.lastname: Binnington
authors.forename: Clifford
authors.initials: C
authors.lastname: Lingwood
mesh_terms: D000218:Adamantane / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D001693:Biological Transport, Active / Q000187:drug effects; D016466:CHO Cells; D002462:Cell Membrane / Q000378:metabolism; D002522:Chlorocebus aethiops; D002784:Cholesterol / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D011992:Endosomes / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D022622:Shiga Toxin 1 / Q000494:pharmacology*; D022641:Shiga Toxin 2 / Q000494:pharmacology*; D013329:Structure-Activity Relationship; D014281:Trihexosylceramides / Q000378:metabolism* / Q000494:pharmacology*; D014709:Vero Cells
chemical_list: D022622:Shiga Toxin 1; D022641:Shiga Toxin 2; D014281:Trihexosylceramides; C514155:adamantyl-globotriaosyl ceramide; C018549:globotriaosylceramide; D002784:Cholesterol; D000218:Adamantane