TABLE 2.
Calculated half-lives of 6myc-Hmg2p and CPY*a
| Strain | Temp (°C) | 6myc-Hmg2p | CPY* (min) | Remark |
|---|---|---|---|---|
| KFY99 (WT CDC48) | 30 | 70 min | ND | 6myc-Hmg2p on plasmid pRH244 |
| 37 | 50 min | |||
| KFY100 (WT CDC48) | 30 | 65 ± 10 min | 23 ± 1.2 | Integrated CPY* (plasmid bMK150) 6myc-Hmg2p on plasmid pRH244 |
| 20 | 69 ± 4 min | 22 ± 1.4 | ||
| 37 | 76 ± 6 min | 24 ± 1.6 | ||
| KFY116 cdc48-1cs | 30 | 89 ± 6 min | 23 ± 1.2 | Integrated CPY* (plasmid bMK150) 6myc-Hmg2p on plasmid pRH244 |
| 20 | 4.1 ± 1 h | 99 ± 6.9 | ||
| KFY194 cdc48-10ts | 30 | 2.6 ± 0.6 h | 22 ± 1.6 | Integrated CPY* (plasmid bMK150) 6myc-Hmg2p on plasmid pRH244 |
| 37 | 10.4 ± 1.1 h | 83 ± 5.3 | ||
| KFY188 cdc48-7ts | 30 | 5.2 h | ND | 6myc-Hmg2p on plasmid pRH244 |
| 37 | 17.0 h | |||
| RHY696 (WT CDC48) | 30 | 40 min | ND | Integrated 6myc-Hmg2p |
| 20 | 75 min | |||
| RHY965 (hrd1Δ) | 30 | 7.2 h | ND | Integrated 6myc-Hmg2p |
| 37 | 7.6 h | |||
| 20 | >10.0 h |
a
The stability of CPY* and 6myc-Hmg2p was measured by cycloheximide-chase assays in strains expressing integrated CPY* or 6myc-Hmg2p (RHY696 and RHY965) or nonintegrated 6myc-Hmg2p on plasmid pRH244. Strains KFY99, KFY100, RHY696, and RHY965 express wild-type (WT) Cdc48p, strain KFY116 carries the cold-sensitive cdc48-1 mutation, and strains KFY188 and KFY194 are two temperature-sensitive cdc48 mutants. Strain RHY965 is hrd1Δ. Blots, represented in Fig. 3 and 4, were quantified by densitometry, and values of t½ were calculated from slopes of semi-logarithmic curves, also represented in Fig. 3 and 4. Values are the means ± standard deviations of eight independent experiments. ND, not done.