title: Uptake of clostridium botulinum C3 exoenzyme into intact HT22 and J774A.1 cells.
abstract: The Clostridium botulinum C3 exoenzyme selectively ADP-ribosylates low molecular weight GTP-binding proteins RhoA, B and C. This covalent modification inhibits Rho signaling activity, resulting in distinct actin cytoskeleton changes. Although C3 exoenzyme has no binding, the translocation domain assures that C3 enters cells and acts intracellularly. C3 uptake is thought to occur due to the high concentration of the C3 enzyme. However, recent work indicates that C3 is selectively endocytosed, suggesting a specific endocytotic pathway, which is not yet understood. In this study, we show that the C3 exoenzyme binds to cell surfaces and is internalized in a time-dependent manner. We show that the intermediate filament, vimentin, is involved in C3 uptake, as indicated by the inhibition of C3 internalization by acrylamide, a known vimentin disruption agent. Inhibition of C3 internalization was not observed by chemical inhibitors, like bafilomycin A, methyl-β-cyclodextrin, nocodazole or latrunculin B. Furthermore, the internalization of C3 exoenzyme was markedly inhibited in dynasore-treated HT22 cells. Our results indicate that C3 internalization depends on vimentin and does not depend strictly on both clathrin and caveolae.
authors: Institute of Toxicology, Hannover Medical School, Carl-Neuberg-Str. 1, Hannover D-30625, Germany. rohrbeck.astrid@mh-hannover.de.
authors: Astrid
authors: A
authors: Rohrbeck
authors: Institute of Toxicology, Hannover Medical School, Carl-Neuberg-Str. 1, Hannover D-30625, Germany. vonElsner.Leonie@mh-hannover.de.
authors: Leonie
authors: L
authors: von Elsner
authors: Institute of Toxicology, Hannover Medical School, Carl-Neuberg-Str. 1, Hannover D-30625, Germany. hagemann.sandra@mh-hannover.de.
authors: Sandra
authors: S
authors: Hagemann
authors: Institute of Toxicology, Hannover Medical School, Carl-Neuberg-Str. 1, Hannover D-30625, Germany. Just.ingo@mh-hannover.de.
authors: Ingo
authors: I
authors: Just
mesh_terms: D036002:ADP Ribose Transferases / Q000378:metabolism*; D000199:Actins / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D001905:Botulinum Toxins / Q000378:metabolism*; D002460:Cell Line; D003014:Clostridium botulinum / Q000201:enzymology; D034281:Dynamins / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004705:Endocytosis; D051379:Mice; D018613:Microscopy, Confocal; D011485:Protein Binding; D011956:Receptors, Cell Surface / Q000378:metabolism; D013997:Time Factors; D014746:Vimentin / Q000378:metabolism*; D020741:rho GTP-Binding Proteins / Q000378:metabolism; D020742:rhoA GTP-Binding Protein
chemical_list: D000199:Actins; D011956:Receptors, Cell Surface; D014746:Vimentin; D036002:ADP Ribose Transferases; C063007:exoenzyme C3, Clostridium botulinum; D001905:Botulinum Toxins; C526380:RhoA protein, mouse; D020741:rho GTP-Binding Proteins; D020742:rhoA GTP-Binding Protein; D034281:Dynamins
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