Figure 6.

Mutations in PMR1 affect the response to induced accumulation of malfolded proteins in the ER. (A) The pmr1 mutant is sensitive to DTT. PMR1 (YR98) and pmr1 (YR122) cells were streaked onto YPD media, supplemented with 0.2% DTT (+) or without addition of DTT (−). Plates were photographed after 3 d incubation at 30°C. (B) The pmr1 mutant displays inositol auxotrophy and hypersensitivity to tunicamycin. Serial fivefold dilutions of saturated cultures were spotted onto synthetic media (left panel) containing (+) or lacking (−) inositol and onto YPD media containing (+) or lacking (−) tunicamycin (0.3 μg/ml, right panel). Plates were photographed after 3 d incubation at 30°C. Strains: YR98 (PMR1) and YR122 (pmr1). (C) Induction of Kar2 after treatment with tunicamycin. Tunicamycin (1 μg/ml) was added to cells growing in YPD (OD₆₀₀ = 0.8). After 1 h, 2 h, and 3 h, cell extracts were prepared and analyzed for the presence of Kar2 by SDS-PAGE and Western blotting. Each lane corresponds to 20 μg of total protein. Strains: YR98 (PMR1) and YR122 (pmr1). (D) PhosphoImager quantitation of the results shown in panel C. For each strain, signals are expressed as ratio of relative induction; the values at t = 0 were arbitrarily set to 1. Strains: YR98 (PMR1) and YR122 (pmr1).