CD95 death-inducing signaling complex formation and internalization occur in lipid rafts of type I and type II cells.
We investigated the membrane localization of CD95 in type I and type II cells, which differ in their ability to recruit and activate caspase-8. We found that CD95 was preferentially located in lipid rafts of type I cells, while it was present both in raft and non-raft plasma membrane sub-domains of type II cells. After stimulation, CD95 located in phospholipid-rich plasma membrane was recruited to lipid rafts in both types of cells. Similarly, CD95 cross-linking resulted in caspase-independent translocation of FADD/MORT1 and caspase-8 to the lipid rafts, which was prevented by a death domain-defective receptor. CD95 internalization was then rapid in type I and delayed in type II cells and showed a substantial correlation with the kinetics of Fas-associated death domain (FADD)and caspase-8 recruitment to lipid rafts. Finally, electron microscopy analysis showed that after CD95 stimulation lipid rafts aggregated in large clusters that were internalized in endosomal vesicles, where caspase-8 underwent massive processing. Taken together, our data demonstrate that CD95 death-inducing signaling complex formation and internalization in type I and type II cells occur in lipid rafts, which are a major site of caspase-8 activation.
Eramo
Adriana
A
Department of Hematology, Oncology and Molecular Medicine, Istituto Superiore di Sanità, Rome, Italy.
Sargiacomo
Massimo
M
Ricci-Vitiani
Lucia
L
Todaro
Matilde
M
Stassi
Giorgio
G
Messina
Carlo G M
CG
Parolini
Isabella
I
Lotti
Fiorenza
F
Sette
Giovanni
G
Peschle
Cesare
C
De Maria
Ruggero
R
D017209:Apoptosis / Q000502:physiology*; D020169:Caspases / Q000378:metabolism; D045744:Cell Line, Tumor; D002784:Cholesterol / Q000378:metabolism; D053418:Death Domain Receptor Signaling Adaptor Proteins; D004705:Endocytosis; D006801:Humans; D021962:Membrane Microdomains / Q000378:metabolism*; D011485:Protein Binding; D021381:Protein Transport; D018124:Receptors, Tumor Necrosis Factor / Q000378:metabolism; D015398:Signal Transduction; D013601:T-Lymphocytes / Q000166:cytology / Q000378:metabolism; D019014:fas Receptor / Q000378:metabolism*
D053418:Death Domain Receptor Signaling Adaptor Proteins; D018124:Receptors, Tumor Necrosis Factor; D019014:fas Receptor; D002784:Cholesterol; D020169:Caspases