Involvement of Raft Aggregates Enriched in Fas/CD95 Death-Inducing Signaling Complex in the Antileukemic Action of Edelfosine in Jurkat Cells
Figure 1
DISC formation in lipid rafts following Jurkat cell incubation with edelfosine.
(A) Untreated control Jurkat cells (Control) and Jurkat cells treated with 10 µM edelfosine (EDLF) for 9 h were analyzed for lipid raft isolation on a discontinuous sucrose density gradient. Raft and non-raft fractions were analyzed by Western blotting for the indicated proteins using specific antibodies. The migration positions of the 55-kDa procaspase-8 as well as of the cleavage product p18 are denoted. Location of GM1-containing lipid rafts was determined using CTx B subunit conjugated to horseradish peroxidase. (B) Fas/CD95 was immunoprecipitated from the raft fraction of edelfosine-treated Jurkat cells. Immunoprecipitates were subjected to SDS-PAGE and immunoblotted with Fas/CD95, FADD- and procaspase-8 specific antibodies, respectively. Raft fraction was also immunoprecipitated with P3X63 (X63) myeloma supernatant as a negative control. Experiments shown are representative of three performed.