title: HA-33 facilitates transport of the serotype D botulinum toxin across a rat intestinal epithelial cell monolayer.
abstract: A large size botulinum toxin complex (L-TC) is composed of a single neurotoxin (BoNT), a single nontoxic nonhaemagglutinin (NTNHA) and a haemagglutinin (HA) complex. The HA complex is comprised of three HA-70 molecules and three arm structures of HA-33/HA-17 that consist of two HA-33 and a single HA-17. In addition to the mature L-TC, smaller TCs are present in cultures: M-TC (BoNT/NTNHA), M-TC/HA-70 and immature L-TCs with fewer HA-33/HA-17 arms than mature L-TC. Because L-TC displays higher oral toxicity than pure BoNT, it was presumed that nontoxic proteins are critical for food poisoning. In this study, the absorption of TCs across intestinal epithelial cells was assessed by examining the cell binding and monolayer transport of serotype D toxins in the rat intestinal epithelial cell line IEC-6. All TCs, including pure BoNT, displayed binding and transport, with mature L-TC showing the greatest potency. Inhibition experiments using antibodies revealed that BoNT, HA-70 and HA-33 could be responsible for the binding and transport. The findings here indicate that all TCs can transport across the cell layer via a sialic acid-dependent process. Nonetheless, binding and transport markedly increased with number of HA-33/HA-17 arms in the TC. We therefore conclude that the HA-33/HA-17 arm is not necessarily required for, but facilitates, transport of botulinum toxin complexes.
authors.affiliation: Department of Food and Cosmetic Science, Faculty of Bioindustry, Tokyo University of Agriculture, Abashiri, Japan.
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Ito
authors.forename: Yoshimasa
authors.initials: Y
authors.lastname: Sagane
authors.forename: Keita
authors.initials: K
authors.lastname: Miyata
authors.forename: Ken
authors.initials: K
authors.lastname: Inui
authors.forename: Tomohito
authors.initials: T
authors.lastname: Matsuo
authors.forename: Ryohta
authors.initials: R
authors.lastname: Horiuchi
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Ikeda
authors.forename: Tomonori
authors.initials: T
authors.lastname: Suzuki
authors.forename: Kimiko
authors.initials: K
authors.lastname: Hasegawa
authors.forename: Hirokazu
authors.initials: H
authors.lastname: Kouguchi
authors.forename: Keiji
authors.initials: K
authors.lastname: Oguma
authors.forename: Koichi
authors.initials: K
authors.lastname: Niwa
authors.forename: Tohru
authors.initials: T
authors.lastname: Ohyama
authors.forename: Toshihiro
authors.initials: T
authors.lastname: Watanabe
mesh_terms: D000818:Animals; D001905:Botulinum Toxins / Q000378:metabolism*; D002460:Cell Line; D004847:Epithelial Cells / Q000378:metabolism*; D021381:Protein Transport; D051381:Rats
chemical_list: C118973:hemagglutinin 33 protein, Clostridium botulinum; C096325:botulinum toxin type D; D001905:Botulinum Toxins