Cells were homogenized in cold buffer (20 mM Tris–HCl pH 7.5, 2 mM EDTA) and centrifuged at 15,000 g for 10 min at 4°C. Protein concentration in supernatants was determined using the BCA protein assay (Thermo Fisher Scientific). All samples were assayed in triplicate using 10 μg of freshly prepared protein extracts. Proteasomal activity was measured using the CHEMICON Proteasome Activity Assay Kit (APT280, Millipore), as described by the manufacturer. The extracts were incubated (2 h at 37°C) with a labeled substrate, LLVY‐7‐amino‐4‐methyl‐coumarin, and the proteolysis activity was monitored by detection of the free fluorophore 7‐amino‐4‐methyl‐coumarin, using a SpectraMax M2e (Molecular devices) at 380/460 nm.