FIGURE 5.

TORC2-dependent post-translational modification of Ypk2 is affected in avo3^ts mutants. A, the control vector (V) or AVO3-expressing plasmid was transformed into WT or avo3^ts cells expressing Ypk2-Myc. Log phase cultures were shifted to 37 °C for 3 h and treated with Zymolyase for 1 h before lysis. The lysates were subjected to SDS-PAGE and Western analysis for Ypk2-Myc using anti-Myc antibodies. Pgk1 serves as a loading control. B, lysates from wild-type or avo3^ts cells were subjected to immunoprecipitation (IP) with anti-Myc antibodies. Immunoprecipitates were incubated with or without alkaline phosphatase for 1 h and analyzed by Western blotting using anti-Myc antibodies. Ypk2-Myc levels in the lysates detected by Western blotting are also shown. C, two avo3^ts mutants carrying the control vector (V) or a plasmid expressing AVO1, AVO2, or AVO3 were subjected to the same assay as in A.