FIGURE 3.

Tom1 and Dia2 control Cdc6 proteolysis in G₁. A, Cdc6 is stabilized in both tom1Δ and dia2Δ mutants in G₁. The indicated strains were arrested with α factor (αF) for 3 h and shifted to 37 °C for 30 min prior to performing stability assays as in Fig. 1B. Three independent experiments were used for quantification. Error bars indicate S.D. B, the catalytic activity of Tom1 Hect domain and the F-box domain of Dia2 is required for Cdc6 turnover. Wild type, tom1C3235A, dia2ΔF strains arrested with α factor were used in protein stability assays. Samples were collected at indicated times after cycloheximide (CHX) addition. Pgk1 was used as a loading control. Three independent experiments were used for quantification. C, the TPR mutant stabilizes Cdc6. Samples from wild type and TPR strains were prepared as in A. The asterisk indicates a nonspecific band. Error bars indicate S.D.