Fig 3. Gnp1 and Agp1 are essential for the uptake of exogenous serine.

(a) WT cells (black line), gnp1Δ cells (red line), agp1Δ cells (green line) and gnp1Δagp1Δ cells (purple line) were grown in SDC medium. Cells were washed and incubated with [¹⁴C]-serine. The uptake rate of [¹⁴C]-serine was measured. Error bars correspond to standard deviations. n = 3. (b) Schematic outline of the main serine consuming and producing processes. (c) WT cells (black line), gnp1Δ cells (red line), agp1Δ cells (green line), gnp1Δagp1Δ cells (purple line) and ser2Δ cells (blue line) were grown in the presence of stable isotope labelled, [¹³C₃¹⁵N₁]-serine in SDC medium. Graphs represent the density function of the rates of [¹³C₃¹⁵N₁]-serine incorporation into all peptides containing a single serine. (d) Cellular concentrations of free [¹³C₃¹⁵N₁]-serine in WT, gnp1Δ cells, apg1Δ cells, gnp1Δagp1Δ cells and ser2Δ cells grown in SDC medium with [¹³C₃¹⁵N₁]-serine were analyzed by mass spectrometry. Error bars represent standard deviations. n = 3. *, P-value <0.05, calculated from t-test. (e) Proteome comparison of lysine labeled WT cells and [¹³C₆¹⁵N₂]-lysine labeled gnp1Δ cells grown in SILAC medium. Protein intensities are plotted against normalized SILAC ratios of heavy (gnp1Δ) to light (WT). Significant outliers are colored in red (p < 1e^-11), orange (p<1e^-4) or dark blue (p < 0.05); other proteins are shown in light blue.