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. 2007 Mar 1;21(5):519–524. doi: 10.1101/gad.1490207

Figure 2.

Figure 2.

Protein product of nonstop mRNA is destabilized by the proteasome. (A) The product of nonstop mRNA is unstable. (Top panels) Samples of W303 cells harboring an indicated plasmid were prepared at indicated times after the addition of cycloheximide (0.1 mg/mL). The levels of the remaining proteins were determined by Western blotting. The samples of cells harboring pGPDp-GFP-2A-HIS3 (WT) were applied after 10-fold dilution. (Bottom panels) The relative levels of proteins were quantified and shown as a function of time after the inhibition of translation by cycloheximide (CHX). Relative amounts are shown as the mean values of three independent experiments with standard deviations. (B) MG132, a proteasome-specific inhibitor, stabilizes the products of nonstop mRNA. (Top panels) W303erg6Δ cells harboring an indicated plasmid were cultured and samples were prepared 2 h after the addition of MG132 at the indicated concentration. The samples of cells harboring pGPDp-GFP-2A-HIS3 (WT) were applied after 10-fold dilution. (Bottom panels) The qualification of GFP-2A-His3 and GFP-2A-His3-NS levels. The relative numbers are shown as the mean values of three independent experiments with standard deviations.