Dynamin is involved in human epithelial cell vacuolation caused by the Helicobacter pylori-produced cytotoxin VacA.
The Helicobacter pylori-produced cytotoxin VacA induces intracellular vacuolation. To elucidate the molecular mechanism of vacuole formation by VacA, we examined the participation of dynamin, a GTPase functioning in intracellular vesicle formation, in human HeLa cells. Immunocytochemistry revealed that endogenous dynamin was localized to vacuoles induced by VacA. In cells transiently transfected with a GTPase-defective (dominant-negative) dynamin mutant, VacA failed to induce vacuolation. In contrast, VacA did induce vacuolation in cells transiently transfected with wild-type dynamin. Furthermore, under VacA treatment, neutral red dye uptake, a parameter of VacA-induced vacuolation, was inhibited in cells stably transfected with the dominant-negative dynamin mutant. In contrast, uptake was markedly enhanced in cells stably transfected with wild-type dynamin. Moreover, VacA cytopathic effects on the viability of HeLa cells were inhibited in cells stably transfected with dominant-negative dynamin-1. Sequential immunocytochemical observation confirmed that expression of dominant-negative dynamin did not affect VacA attachment to or internalization into HeLa cells. We suggest that dynamin is involved in the intracellular vacuolation induced by VacA.
Suzuki
J
J
Department of Internal Medicine, University of Tokyo School of Medicine, Tokyo, Japan.
Ohnsihi
H
H
Shibata
H
H
Wada
A
A
Hirayama
T
T
Iiri
T
T
Ueda
N
N
Kanamaru
C
C
Tsuchida
T
T
Mashima
H
H
Yasuda
H
H
Fujita
T
T
D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D002470:Cell Survival; D003603:Cytotoxins / Q000096:biosynthesis*; D004378:Duodenal Diseases / Q000382:microbiology; D034284:Dynamin I; D034281:Dynamins; D004847:Epithelial Cells / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D020558:GTP Phosphohydrolases / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D016480:Helicobacter pylori / Q000378:metabolism*; D006801:Humans; D010957:Plasmids; D014162:Transfection
D001426:Bacterial Proteins; D003603:Cytotoxins; C074689:VacA protein, Helicobacter pylori; D034284:Dynamin I; D020558:GTP Phosphohydrolases; D034281:Dynamins