Fig. 3.

AICAR treatment increases FSP27 ubiquitination in 3T3-L1 adipocytes. FSP27-FLAG-overexpressed cells were pretreated with 10 μM MG-132 (A and B), 1 μM epoxomycin (C and D), or 100 μM chloroquine (E and F) for 30 min prior to the addition of 2 mM AICAR. After 3 h of AICAR treatment, cells were lysed for immunoblotting (IB) analysis. G: cells were infected with Ad-null or Ad-FSP27-FLAG for 2 days. After 1.5 h of treatment with 2 mM of AICAR, immunoprecipitation (IP) was performed by anti-FLAG affinity gel. Immunoprecipitates were subject to immunoblotting analysis using anti-ubiquitin and -FLAG antibodies. The experiments were repeated 3 times, and data are shown as means ± SD. **P < 0.01 and ***P < 0.001 vs. single AICAR treatment.