FIG 3.

Overexpression of YPC1 influences the growth of tetAUR1 cells depending on the mode of AUR1 repression. (A and B) Tenfold dilutions of cells carrying vectors for overexpression of YPC1 or the control vector were spotted on glucose- or galactose-containing medium. Two centromeric vectors having YPC1 either behind the strong constitutive promoter of ADH1 or behind the galactose-inducible GAL1 promoter were utilized. Growth media were supplemented with AbA or Doxy as indicated. In this experiment, cells plated on glucose or galactose had been precultured on that same carbon source before plating, but the same result was also obtained when all cells had been precultured on glucose (not shown). (C) The indicated strains were grown in SC medium to exponential phase (OD₆₀₀ = 0.5 to 0.7) and further treated for 3 h with either Calcofluor white (CFW) (5.0 μg/ml), AbA (3.0 μg/ml), or Doxy (10.0 μg/ml). Proteins were extracted and resolved by SDS-PAGE. Phosphorylated Slt2 (pSlt2) and Ypk1 (pYpk1, phosphorylated on Thr662) were detected as described previously (30). Sec63 was used as a loading control. For the quantifications of pSlt2 shown on the tops of lanes, data were normalized with regard to the amount of Sec63 and expressed as a fraction of results for nontreated cells (1.00).