FIGURE 9:

Overexpression of Gpa1 perturbs surface recycling. (A) Confocal microscopy of wild-type cells expressing Ste3-GFP-DUb with and without Gpa2-mCherry. (B) Localization of Ste3-GFP-DUb in wild-type cells transformed with vector or Gpa2-mCherry at mid–log phase was quantified from >35 cells (n = 3). Unpaired Student’s t tests were performed, with asterisk (*) indicating p < 0.000001. (C) FM4-64 efflux assay was measured from wild-type cells expressing either Gpa2-GFP or Mup1-GFP. Cells were loaded with rich media containing 40 µM FM4-64 for 8 min before washing and dye efflux measured over time by flow cytometry and expressed a % of the initial 10 s fluorescence. (D) Wild-type cells expressing the copper-inducible Gpa2-mCherry construct, using 50 µM copper chloride, and coexpressing either Mup1-GFP, Can1-GFP, or Ste3-GFP were imaged using confocal microscopy. Scale bar, 5 µm.