Differential activities of plant polyphenols on the binding and internalization of cholera toxin in vero cells.
Plant polyphenols, RG-tannin, and applephenon had been reported to inhibit cholera toxin (CT) ADP-ribosyltransferase activity and CT-induced fluid accumulation in mouse ileal loops. A high molecular weight fraction of hop bract extract (HBT) also inhibited CT ADP-ribosyltransferase activity. We report here the effect of those polyphenols on the binding and entry of CT into Vero cells. Binding of CT to Vero cells or to ganglioside GM1, a CT receptor, was inhibited in a concentration-dependent manner by HBT and applephenon but not RG-tannin. These observations were confirmed by fluorescence microscopy using Cy3-labeled CT. Following toxin binding to cells, applephenon, HBT, and RG-tannin suppressed its internalization. HBT or applephenon precipitated CT, CTA, and CTB from solution, creating aggregates larger than 250 kDa. In contrast, RG-tannin precipitated CT poorly; it formed complexes with CT, CTA, or CTB, which were demonstrated with sucrose density gradient centrifugation and molecular weight exclusion filters. In agreement, CTA blocked the inhibition of CT internalization by RG-tannin. These data suggest that some plant polyphenols, similar to applephenon and HBT, bind CT, forming large aggregates in solution or, perhaps, on the cell surface and thereby suppress CT binding and internalization. In contrast, RG-tannin binding to CT did not interfere with its binding to Vero cells or GM1, but it did inhibit internalization.
Morinaga
Naoko
N
Department of Molecular Infectiology, Graduate School of Medicine, Chiba University, Japan. nmorinaga@faculty.chiba-u.jp
Iwamaru
Yoshifumi
Y
Yahiro
Kinnosuke
K
Tagashira
Motoyuki
M
Moss
Joel
J
Noda
Masatoshi
M
D000818:Animals; D002470:Cell Survival; D002499:Centrifugation, Density Gradient; D002522:Chlorocebus aethiops; D002772:Cholera Toxin / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D004797:Enzyme-Linked Immunosorbent Assay; D005419:Flavonoids / Q000378:metabolism / Q000494:pharmacology*; D005638:Fruit / Q000378:metabolism; D005677:G(M1) Ganglioside / Q000378:metabolism; D027582:Humulus; D047348:Hydrolyzable Tannins / Q000737:chemistry / Q000494:pharmacology*; D008856:Microscopy, Fluorescence; D008970:Molecular Weight; D010636:Phenols / Q000302:isolation & purification / Q000494:pharmacology* / Q000633:toxicity*; D010936:Plant Extracts / Q000378:metabolism / Q000494:pharmacology; D059808:Polyphenols; D011485:Protein Binding; D013395:Sucrose / Q000494:pharmacology; D013997:Time Factors; D014709:Vero Cells
D005419:Flavonoids; D047348:Hydrolyzable Tannins; D010636:Phenols; D010936:Plant Extracts; D059808:Polyphenols; D005677:G(M1) Ganglioside; D013395:Sucrose; D002772:Cholera Toxin