Figure 3. Decreased chymotrypsin-like activity of proteasomes in U87MG cells that express PTEN.

The same cells from Figure 2 were cultured in KH without (A) or with (B) amino acids and insulin. The chymotrypsin-like peptidase activity of proteasomes was determined using the fluorogenic substrate N-Suc-LLVY-AMC as described in Materials and Methods. As a control, the specific proteasome inhibitor lactacystin (40 µM) was added and the activity of proteasomes was determined as the difference obtained in the absence and in the presence of lactacystin, which was about 12-15% of the activity measured without the inhibitor. Results are expressed in percentage relative to the fluorescence values of the mock-treated cells at 3.5 h and are means ± SD of five independent experiments, each of them run in duplicate.