Figure 4.

Proteolytic turnover mediated by the Hac1p degron requires components of the SCF^Cdc4 pathway. (A) Hac1p protein is stabilized in MT670 cells temperature sensitive for the E2 ubiquitin-conjugating enzyme Ubc3 (Cdc34p). Synthesis shutoff assays on MT670 and parental W303 cells were performed as described in the legend to Figure 1A. (B) The Cdc53 cullin and Skp1 core components of the SCF^Cdc4 E3 ubiquitin ligase complex are required for rapid turnover of Hac1p. Synthesis shutoff assays on MT871 (cdc53^ts), Y552 (skp1-11^ts), and Y554 (skp1-12^ts) cells and their respective parental W303 and Y80 strains were performed as described in the legend to Figure 1A. (C) The Cdc4 F-box protein is required for rapid turnover of Hac1p. Synthesis shutoff assays on MT668 (cdc4^ts) and PY283 (met30^ts) and their respective parental W303 and PY1 strains were performed as described in the legend to Figure 1A. Assays on deletion strain grr1Δ and its BY4742 parent strain were performed as described in the legend to Figure 2B.