Identification of the cytolinker plectin as a major early in vivo substrate for caspase 8 during CD95- and tumor necrosis factor receptor-mediated apoptosis.
Caspase 8 plays an essential role in the execution of death receptor-mediated apoptosis. To determine the localization of endogenous caspase 8, we used a panel of subunit-specific anti-caspase 8 monoclonal antibodies in confocal immunofluorescence microscopy. In the human breast carcinoma cell line MCF7, caspase 8 predominantly colocalized with and bound to mitochondria. After induction of apoptosis through CD95 or tumor necrosis factor receptor I, active caspase 8 translocated to plectin, a major cross-linking protein of the three main cytoplasmic filament systems, whereas the caspase 8 prodomain remained bound to mitochondria. Plectin was quantitatively cleaved by caspase 8 at Asp 2395 in the center of the molecule in all cells tested. Cleavage of plectin clearly preceded that of other caspase substrates such as poly(ADP-ribose) polymerase, gelsolin, cytokeratins, or lamin B. In primary fibroblasts from plectin-deficient mice, apoptosis-induced reorganization of the actin cytoskeleton, as seen in wild-type cells, was severely impaired, suggesting that during apoptosis, plectin is required for the reorganization of the microfilament system.
Stegh
A H
AH
Tumor Immunology Program, German Cancer Research Center, D-69120 Heidelberg, Germany.
Herrmann
H
H
Lampel
S
S
Weisenberger
D
D
Andrä
K
K
Seper
M
M
Wiche
G
G
Krammer
P H
PH
Peter
M E
ME
D000199:Actins / Q000378:metabolism / Q000648:ultrastructure; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D001692:Biological Transport; D001943:Breast Neoplasms; D002277:Carcinoma; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000276:immunology / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D004792:Enzyme Precursors / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D018260:Gelsolin / Q000378:metabolism; D006801:Humans; D007381:Intermediate Filament Proteins / Q000235:genetics / Q000378:metabolism*; D007633:Keratins / Q000378:metabolism; D034921:Lamin Type B; D034882:Lamins; D051379:Mice; D008817:Mice, Mutant Strains; D008928:Mitochondria / Q000378:metabolism; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D051190:Plectin; D018124:Receptors, Tumor Necrosis Factor / Q000378:metabolism*; D013379:Substrate Specificity; D014407:Tumor Cells, Cultured; D019014:fas Receptor / Q000378:metabolism*
D000199:Actins; D000911:Antibodies, Monoclonal; D004792:Enzyme Precursors; D018260:Gelsolin; D007381:Intermediate Filament Proteins; D034921:Lamin Type B; D034882:Lamins; D009687:Nuclear Proteins; C495332:PLEC protein, human; C495333:Plec protein, mouse; D051190:Plectin; D018124:Receptors, Tumor Necrosis Factor; D019014:fas Receptor; D007633:Keratins; C505530:CASP8 protein, human; C506433:CASP9 protein, human; C505526:Casp8 protein, mouse; C506434:Casp9 protein, mouse; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases