RRC ID 18530
Author Nagahama M, Itohayashi Y, Hara H, Higashihara M, Fukatani Y, Takagishi T, Oda M, Kobayashi K, Nakagawa I, Sakurai J.
Title Cellular vacuolation induced by Clostridium perfringens epsilon-toxin.
Journal FEBS J
Abstract The epsilon-toxin of Clostridium perfringens forms a heptamer in the membranes of Madin-Darby canine kidney cells, leading to cell death. Here, we report that it caused the vacuolation of Madin-Darby canine kidney cells. The toxin induced vacuolation in a dose-dependent and time-dependent manner. The monomer of the toxin formed oligomers on lipid rafts in membranes of the cells. Methyl-β-cyclodextrin and poly(ethylene glycol) 4000 inhibited the vacuolation. Epsilon-toxin was internalized into the cells. Confocal microscopy revealed that the internalized toxin was transported from early endosomes (early endosome antigen 1 staining) to late endosomes and lysosomes (lysosomal-associated membrane protein 2 staining) and then distributed to the membranes of vacuoles. Furthermore, the vacuolation was inhibited by bafilomycin A1, a V-type ATPase inhibitor, and colchicine and nocodazole, microtubule-depolymerizing agents. The early endosomal marker green fluorescent protein-Rab5 and early endosome antigen 1 did not localize to vacuolar membranes. In contrast, the vacuolar membranes were specifically stained by the late endosomal and lysosomal marker green fluorescent protein-Rab7 and lysosomal-associated membrane protein 2. The vacuoles in the toxin-treated cells were stained with LysoTracker Red DND-99, a marker for late endosomes and lysosomes. A dominant negative mutant of Rab7 prevented the vacuolization, whereas a mutant form of Rab5 was less effective. These results demonstrate, for the first time, that: (a) oligomers of epsilon-toxin formed in lipid rafts are endocytosed; and (b) the vacuoles originating from late endosomes and lysosomes are formed by an oligomer of epsilon-toxin.
Volume 278(18)
Pages 3395-407
Published 2011-9-1
DOI 10.1111/j.1742-4658.2011.08263.x
PMID 21781280
MeSH Animals Bacterial Toxins / antagonists & inhibitors Bacterial Toxins / toxicity* Biomarkers / metabolism Cell Line Dogs Endocytosis / drug effects Endosomes / drug effects Endosomes / metabolism Endosomes / ultrastructure Kidney / drug effects* Kidney / metabolism Kidney / ultrastructure* Kinetics Lysosomal-Associated Membrane Protein 2 / metabolism Lysosomes / drug effects Lysosomes / metabolism Lysosomes / ultrastructure Membrane Microdomains / drug effects Membrane Microdomains / ultrastructure Microtubules / drug effects Mutant Proteins / genetics Mutant Proteins / metabolism Polyethylene Glycols / pharmacology Tubulin Modulators / pharmacology Vacuoles / drug effects* Vacuoles / metabolism Vacuoles / ultrastructure* Vesicular Transport Proteins / metabolism beta-Cyclodextrins / pharmacology rab GTP-Binding Proteins / genetics rab GTP-Binding Proteins / metabolism rab7 GTP-Binding Proteins
IF 4.392
Times Cited 24
Human and Animal Cells MDCK(RCB0995)