FIGURE 5:

The effects of polarisome mutations on Slt2p activation level and cell growth in sec3∆ cells. (A) Yeast lysates were extracted separately from strains wt, ptc1∆, sec3∆, and myo4∆ (lanes 1–4) or from strains wt, sec3∆, sec3∆bud6∆, sec3∆spa2∆, and sec3∆pea2∆ (lanes 5–9) and blotted using antibody that specifically recognizes dually phosphorylated Slt2p (top) or the total Slt2p (middle). Pgk1p was used as a loading control. (B) The relative activation level of Slt2p was quantified from the blots. Error bars, SEM from three independent experiments. (C) Tetrad dissection analysis with sec3∆. Tetrad dissection plates from the indicated crosses are shown with representative double-deletion mutant spores circled and representative single sec3∆ spores squared. Replica plating to infer genotype later showed that the inviable spores were double mutants. In the cross of sec3∆ to spa2∆, spores were dissected onto two different plates, and empty regions of the plates were deleted as indicated by the white borders.