Trafficking of ganglioside GD3 to mitochondria by tumor necrosis factor-alpha.
The interaction of mitochondria with proapoptotic proteins activates apoptosis pathways. Previous findings have identified ganglioside GD3 (GD3) as an emerging apoptotic lipid intermediate that targets mitochondria in response to death signals. Using immunoelectron and laser scanning confocal microscopy, we characterize the trafficking of GD3 to mitochondria in response to tumor necrosis factor-alpha (TNF-alpha) in rat hepatocytes. In control hepatocytes, GD3 is present predominantly at the plasma membrane as well as in the endosomal/Golgi network, as verified by its colocalization with the asialoglycoprotein receptor. Following TNF-alpha exposure, GD3 undergoes a rapid cellular redistribution with a gradual loss from the plasma membrane before its colocalization with mitochondria. This process is mimicked by acidic sphingomyelinase and ionizing radiation but not by neutral sphingomyelinase or staurosporin. TNF-alpha stimulated the colocalization of GD3 with early and late endosomal markers, Rab 5 and Rab 7, whereas perturbation of plasma membrane cholesterol or actin cytoskeleton or inhibition of glucosylceramide synthase prevented the trafficking of GD3 to mitochondria. Finally, prevention of the TNF-alpha-stimulated neosynthesis of GD3, cyclosporin A, and latrunculin A or filipin protected sensitized hepatocytes from TNF-alpha-mediated cell death. Thus, the intracellular redistribution and mitochondrial targeting of GD3 during TNF-alpha signaling occurs through actin cytoskeleton vesicular trafficking and contributes to TNF-alpha-mediated hepatocellular cell death.
Liver Unit, Instituto de Malalties Digestives, Hospital Clinic i Provincial, Barcelona, 08036, Spain.
Carmen
C
García-Ruiz
Anna
A
Colell
Albert
A
Morales
María
M
Calvo
Carlos
C
Enrich
José C
JC
Fernández-Checa
D000199:Actins / Q000378:metabolism; D000818:Animals; D017209:Apoptosis; D001692:Biological Transport; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000494:pharmacology; D016923:Cell Death; D002462:Cell Membrane / Q000378:metabolism; D002784:Cholesterol / Q000378:metabolism; D016572:Cyclosporine / Q000494:pharmacology; D003599:Cytoskeleton / Q000378:metabolism; D011992:Endosomes / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005372:Filipin / Q000494:pharmacology; D005732:Gangliosides / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism; D022781:Hepatocytes / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D018613:Microscopy, Confocal; D016253:Microscopy, Immunoelectron; D008928:Mitochondria / Q000378:metabolism*; D011485:Protein Binding; D051381:Rats; D015398:Signal Transduction; D013844:Thiazoles / Q000494:pharmacology; D053778:Thiazolidines; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*; D020691:rab GTP-Binding Proteins / Q000378:metabolism; D020696:rab5 GTP-Binding Proteins / Q000378:metabolism; D000090802:rab7 GTP-Binding Proteins
D000199:Actins; D019086:Bridged Bicyclo Compounds, Heterocyclic; D004791:Enzyme Inhibitors; D005732:Gangliosides; D013844:Thiazoles; D053778:Thiazolidines; D014409:Tumor Necrosis Factor-alpha; D000090802:rab7 GTP-Binding Proteins; C026226:ganglioside, GD3; D016572:Cyclosporine; D005372:Filipin; D002784:Cholesterol; D020691:rab GTP-Binding Proteins; D020696:rab5 GTP-Binding Proteins; C037067:latrunculin A