Figure 3.

Intracellular distribution of Rpn5 in rpn11-m1. (A) WT (left) and rpn11-m1 mutant cells (right) expressing a genomic copy of Rpn5–GFP and plasmids expressing either mCherry-VHL (top) or mCherry-Rnq1 (bottom). Cells were grown at indicated temperatures for 5 h and images of living cells were taken by ×100 oil immersion objective using a confocal microscope (Zeiss LSM 510). Z stacked images were captured at 0.5 μM intervals and images were processed using Zen Lite software. (B) Total cell extracts from rpn11-m1 constitutively expressing Rpn5-TAP under ADH1 promoter or expressing Rpn5 under the control of RPN10 promoter were resolved in polyacrylamide gel and blotted with anti-yCul1 and anti-Rpn5.