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This Article Full Text Full Text (PDF) All Versions of this Article: 287/2/G436    most recent 00019.2004v1 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 HighWire Citing Articles via ISI Web of Science (32) Citing Articles via Google Scholar Google Scholar Articles by Werneburg, N. Articles by Gores, G. J. Search for Related Content PubMed PubMed Citation Articles by Werneburg, N. Articles by Gores, G. J.

LIVER AND BILIARY TRACT

TNF--mediated lysosomal permeabilization is FAN and caspase 8/Bid dependent

¹Division of Gastroenterology and Hepatology, Mayo Clinic College of Medicine, Rochester, Minnesota 55905; and ²Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15260

Submitted 15 January 2004 ; accepted in final form 29 March 2004

TNF- cytotoxic signaling involves lysosomal permeabilization with release of the lysosomal protease cathepsin B (ctsb) into the cytosol. However, the mechanisms mediating lysosomal breakdown remain unclear. Because caspase-8 and factor associated with neutral sphingomyelinase activation (FAN) have been implicated as proximal mediators of TNF--associated apoptosis, their role in lysosomal permeabilization was examined. Cellular distribution of ctsb-green fluorescent protein (ctsb-GFP) in a rat hepatoma cell line was imaged by confocal microscopy. ctsb-GFP fluorescence was punctate under basal conditions but became diffuse after treatment with TNF-/actinomycin D. This cellular redistribution of ctsb-GFP was blocked by transfection with a vector expressing a dominant-negative Fas-associated protein with death domain (FADD), cytokine response modifier A, or a pharmacological caspase-8 inhibitor, IETD-fmk. Consistent with the concept that caspase 8-mediated apoptosis is also Bid-dependent in hepatocytes, ctsb-GFP release from lysosomes was reduced in hepatocytes from Bid^–/– mice. Interestingly, transfection with a vector expressing a dominant-negative FAN (FAN) also blocked ctsb-GFP release and caspase-8 activation. Paradigms that inhibited ctsb-GFP release from lysosomes also reduced apoptosis as assessed by morphology and biochemical criteria. In conclusion, these studies suggest FAN is upstream of caspase-8/Bid in a signaling cascade culminating in lysosomal permeabilization.

cathepsin B; dominant-negative Fas-associated protein with death domain; dominant-negative factor associated with neutral sphingomyelinase activation; green fluorescent protein

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