Analysis of A
R expression and membrane expansion in the Δpah1 strain. A. Western blot analysis of A2AR in 10 μg of total membrane proteins of wild type, Δpah1, HAC1 co-overexpression and Δpah1/Hac1p strains. B. Radioligand binding studies on membranes expressing the A2AR. Deletion of the PAH1 gene leads to a strong increase (over 20-fold) in production of ligand-binding receptor. Artificial induction of the UPR reduces the quantity of receptor produced, but strongly enhances the radioligand binding activity, indicating enhanced receptor protein quality. C. EM pictures were taken after 48 hours growth on oleic acid of both wild type and Δpah1 cells. When the knockout cells are grown on oleic acid, whirls and stacked layers of proliferated membrane become apparent. Also, few or no lipid droplets are present in the knockout strain, while big droplets are present in the wild type cells (arrows). D. HPLC-LSD analysis of the major cellular lipid classes derived from the strains grown on oleic acid. Lipid levels were quantified relative to those in the wild type parental strain (represented as 100%, broken horizontal line). The values derived from two independent experiments are shown, with their average represented by the short horizontal lines.