Figure 2.

Substrate-induced transporter recycling. (A) Cartoon of substrate-induced degradation (left) and recycling (right) of Mup1 triggered by modulation of extracellular methionine. (B) Time-lapse Airyscan microscopy of cells expressing Mup1-GFP before and after 2-min methionine (2 µg/ml, upper and 40 µg/ml, lower) pulse-chase incubations. (C) Cells expressing Mup1-GFP were incubated with 20 µg/ml methionine for 0, 1, 5, and 60 min followed by three times washes and further incubation in SC-Met up to 60 min before lysates were generated and immunoblotted. (D) Quantification of average intensity of Mup1-GFP (left) and vacuolar processed GFP (right) from methionine pulse-chase experiments from C. *, P < 0.01. (E and F) Yellow regions from cells expressing Mup1-mEos and Fur4-mEos were exposed to 405-nm laser at 0.5% to photoconvert molecules (left) and mEOS fluorescence-tracked over time before (right). (G) Time-lapse microscopy of cells expressing Mup1-mEOS following three times pulse with 0.1% 405-nm laser followed by substrate-induced recycling stimulated by 2 µg/ml methionine for 30 s. Scale bar, 5 µm (white); 1 µm (yellow). Source data are available for this figure: SourceData F2.