Figure 2.

Deletions of components of the NMD pathway suppress tra1-L3733A. A. Haploid strains CY4353 (TRA1 UPF1), CY4103 (tra1-L3733A UPF1), CY5932 (TRA1 upf1Δ), CY5972 (tra1-L3733A upf1Δ), CY5936 (TRA1 upf3Δ), CY5983 (tra1-L3733A upf3Δ), CY5934 (TRA1 upf2Δ), and CY5996 (tra1-L3733A upf2Δ) were grown in YPD media to stationary phase, and 10-fold serial dilutions plated onto YPD, YPD containing 6% ethanol, or YPD grown at 37°C. B. Expression of Flag⁵-Tra1-L3733A. Yeast strains CY5940 (Flag⁵-TRA1; lane 1), CY6004 (Flag⁵-tra1-L3733A; lanes 2-4), CY6005 (Flag⁵-tra1-L3733A upf1Δ0; lanes 5-7) and BY4741 (TRA1, not applicable; lane 8) were grown to stationary phase then diluted 1:20 into YPD and grown for 8 hours at 30°C. Protein extracts were prepared by lysis with glass beads. The indicated amount of protein was separated by SDS-PAGE (5%) and either Western blotted with anti-Flag antibody or stained with Coomassie Brilliant Blue (CBB).