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. 2002 Sep 20;99(20):12853–12858. doi: 10.1073/pnas.202115499

Figure 2.

Figure 2

Separation of wild-type Pma1 and Pma1-D378N in lcb1-100 cells. Cells (RH3804) bearing GAL-HA-PMA1 (pND542) and GAL-myc-pma1-D378N (pWQ4) were shifted to galactose-containing medium for 4 h. Lysates were prepared and fractionated. (A) Distribution of HA and myc epitopes and Sec61 was assayed by Western blotting. HA-Pma1 is predominantly found in fractions 9 and 10, whereas myc-Pma1-D378N is colocalized with Sec61 in fractions 1, 2, and 3. (B) Reversibility by phytosphingosine (PHS). Phytosphingosine (50 μM) was added on shift to galactose, and the distributions of HA and myc epitopes and Sec61 were assayed after fractionation. (C) lcb1-100 is a suppressor of pma1-D378N. Growth of cells bearing GAL-pma1-D378N (pWQ4) at 30°C on plates with synthetic complete medium without uracil containing glucose or galactose.