Figure 1. Effect of the 26S proteasome inhibitor on protein synthesis in COS-7 cells.

To determine the protein synthesis rate after MG132 treatment (50 μM) or 0.1% DSMO for 4 h, cells were labeled by addition of either 0.5 μCi/ml [³H]proline or 0.4 μCi/ml [³H]methionine to the complete media for 20 min. Cells were washed extensively with PBS, and with 10% TCA for 10 min to remove unincorporated radioactive precursor. Cells were washed with 10% TCA twice more for 5 min each. After a final wash with absolute MeOH, cells were lysed in 0.3N NaOH, 1% SDS for 30 min at room temperature. Aliquots were counted by liquid scintillation counting in 10 ml BCS. Values are means±SEM (n = 4 in [³H]proline experiment; n = 3 in [³H]methionine experiment). The p values significantly different in comparison with the control are indicated (Student’s t-test). The SEM is small and not seen in the MG132-treated group in [³H]methionine incorporation experiment (plotted on the right Y-axis).