Figure 3. tRNA uridine thiolation regulates amino acid biosynthesis and sugar metabolism.
(A) Schematic of the SILAC quantitative proteomic approach used to assess global changes in protein levels in thiolation-deficient cells (Supplemental Information).
(B) tRNA uridine thiolation is responsible for most uba4Δ phenotypes. Correlation plot comparing normalized log2 H/L ratios from WT/ubaΔ (X-axis) and WT/ncs2Δ (Y-axis) samples, for all proteins detected in both samples (~1900 total). ncs2Δ and uba4Δ strains showed similar profiles (slope=0.87, r=0.83). See also Table S2 for complete data, Table S3 and S4 for GO analysis data.
(C) tRNA uridine thiolation regulates sugar and carbohydrate metabolism as well as sulfur and lysine amino acid metabolism. Proteins increased (top panel) or decreased (bottom panel) by at least 1.4 fold in thiolation-deficient strains (compared to WT cells) were analyzed by GO term grouping, to find significantly enriched terms (significance threshold p<0.0001). Consolidated GO-term groups were visualized using the descriptive, scaled TreeMaps shown (Supplemental Information).
(D) Proteins related to sulfur amino acid metabolism or lysine metabolism that increase or decrease in uba4Δ or ncs2Δ mutant cells were compared using log2 H/L ratio plots. WT/uba4Δ (X-axis) ratios for these proteins are compared against WT/ncs2Δ (Y-axis). Both samples showed a nearly 1:1 correlation (r=0.95, slope=1).
(E) Sulfur amino acid metabolism-related proteins that increase in tRNA uridine thiolation-deficient mutants. Proteins involved in methionine or cysteine biosynthesis or salvage that specifically increased in both uba4Δ and ncs2Δ mutants are indicated in magenta. See also Figure S3.
(F) Lysine metabolism-related proteins that increased in tRNA uridine thiolation-deficient mutants (uba4Δ, ncs2Δ) compared to WT are indicated in magenta.