Fig. 8.
The ESCRT system is required for sorting of ubiquitylated LDLR. A: A431 cells were transfected with 20 nM of control or TSG101 siRNA and subsequently incubated for 16 h with sterol-depletion medium followed by treatment with vehicle (−) or 1 μM GW3965 (+) for 5 h. B: HEK293T cells were transfected with expression plasmids for LDLR, FLAG-IDOL, GFP-VPS4WT, GFP-VPS4E228Q, or GFP. C: HEK293T cells were transfected with expression plasmids for LDLR, Myc-IDOL, FLAG-AMSH, FLAG-USP8, and GFP to monitor for transfection efficiency. In (A–C) total cell lysates were analyzed by immunoblotting as indicated. Immunoblots are representative of three independent experiments.