Extended Data Figure 7: tRNA modification status in ∆SHMT2 and effects of 5-taurinomethyluridine modification loss caused by human disease gene MTO1.
From: Mitochondrial translation requires folate-dependent tRNA methylation
a, Total ion chromatogram of 5-formylcytidine monophosphate in digested mitochondrial tRNAs upon loss of SHMT2. The same samples were analysed for 5-taurinomethyluridine monophosphate (p-τm5U) in Fig. 4b. The combined data demonstrate that SHMT2 deletion causes loss of τm5U but not 5-formylcytidine. b, Levels of τm5U, 5-taurinomethyl-2-thiouridine monophosphate (p-τm5s2U) and 2-thiouridine monophosphate (p-s2U) in wild-type HCT116 and SHMT2 deletion lines normalized to 5-formylcytidine monophosphate (p-f5C) (n = 3). c, Taurine levels in HCT116 wild-type and SHMT2-knockout cells (n = 3). d, τm5U levels in digested mitochondrial tRNAs upon re-expression of SHMT2 (n = 1). e, τm5U, τm5s2U and s2U levels normalized to f5C in HCT116 SHMT2/MTHFD2 knockout lines after sarcosine supplementation and HCT116 upon loss of MTO1 (n = 2). For all panels, data are mean ± s.e.m. or individual data points only. f, Labelling pattern of 5-taurinomethyluridine and 5-formylcytidine monophosphate extracted from mitochondrial tRNAs after growth in media containing either [3-13C]serine or [U-13C]methionine. g, Mean cumulative count of ribosome protected fragments (RPF) mapping to mitochondrial protein coding transcripts upon ribosome profiling in HCT116 MTO1-knockout cell lines. Data were normalized to RPM (n = 2); n indicates the number of biological replicates. *P < 0.01, two-tailed Student’s t-test (see Supplementary Table 7 for exact P values).
