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This Article Full Text Full Text (PDF) All Versions of this Article: 293/3/F927    most recent 00199.2007v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in ISI Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via ISI Web of Science (1) Citing Articles via Google Scholar Google Scholar Articles by Chassin, C. Articles by Vandewalle, A. Search for Related Content PubMed PubMed Citation Articles by Chassin, C. Articles by Vandewalle, A.

Pore-forming epsilon toxin causes membrane permeabilization and rapid ATP depletion-mediated cell death in renal collecting duct cells

¹Institut National de la Santé et de la Recherche Médicale, U773, Centre de Recherche Biomédicale Bichat-Beaujon CRB3, Paris; ²Université Paris 7-Denis Diderot, Paris; ³Institut des Neurosciences Cellulaires et Intégratives, UMR7168-LC2 CNRS-Université L. Pasteur, Département de Neurotransmission et Sécrétion Neuroendocrine, Strasbourg; and ⁴Unité des Bactéries Anaérobies et Toxines, Institut Pasteur, Paris, France

Submitted 26 April 2007 ; accepted in final form 9 June 2007

Clostridium perfringens epsilon toxin (ET) is a potent pore-forming cytotoxin causing fatal enterotoxemia in livestock. ET accumulates in brain and kidney, particularly in the renal distal-collecting ducts. ET binds and oligomerizes in detergent-resistant membranes (DRMs) microdomains and causes cell death. However, the causal linkage between membrane permeabilization and cell death is not clear. Here, we show that ET binds and forms 220-kDa insoluble complexes in plasma membrane DRMs of renal mpkCCD_cl4 collecting duct cells. Phosphatidylinositol-specific phospholipase C did not impair binding or the formation of ET complexes, suggesting that the receptor for ET is not GPI anchored. ET induced a dose-dependent fall in the transepithelial resistance and potential in confluent cells grown on filters, transiently stimulated Na⁺ absorption, and induced an inward ionic current and a sustained rise in [Ca²⁺]_i. ET also induced rapid depletion of cellular ATP, and stimulated the AMP-activated protein kinase, a metabolic-sensing Ser/Thr kinase. ET also induced mitochondrial membrane permeabilization and mitochondrial-nuclear translocation of apoptosis-inducing factor, a potent caspase-independent cell death effector. Finally, ET induced cell necrosis characterized by a marked reduction in nucleus size without DNA fragmentation. DRM disruption by methyl--cyclodextrin impaired ET oligomerization, and significantly reduced the influx of Na⁺ and [Ca²⁺]_i, but did not impair ATP depletion and cell death caused by the toxin. These findings indicate that ET causes rapid necrosis of renal collecting duct cells and establish that ATP depletion-mediated cell death is not strictly correlated with the plasma membrane permeabilization and ion diffusion caused by the toxin.

ion diffusion; lipid raft; oncosis