Extended Data Fig. 2: Cysteine depletion triggers metabolic rewiring without altering protein synthesis and iron-sulfur cluster maintenance.
From: Cysteine depletion triggers adipose tissue thermogenesis and weight loss
a, b) Volcano plot of a) serum and b) subcutaneous adipose tissue (SFAT) metabolites identified by LC-MS/MS in Cth−/− mice after 6 days of CTRL or CysF diet (n = 4 CTRL and n = 5 CysF for serum, n = 6/group for SFAT). Transsulfuration pathway related metabolites are highlighted in red. Significantly increased or decreased metabolites (-log10(pvalue)>1.3 and ∣log2(FC)∣>1) are highlighted in blue and listed on the right. Cys: cysteine. Met: methionine. SAH: S-adenosyl homocysteine. SAM: S-adenosyl methionine. Cysteine and cysteine were not detectable in SFAT samples. See Supplementary Table 1 and 2 for the full list of metabolites. c) Serum L-methionine, L-homocysteine, glutamic acid quantified by mass spectrometry in Cth−/− mice fed with CTRL (n = 4) or CysF (n = 5) diet for 6 days. AU: arbitrary units. d) Total GSSG content in SFAT, brown adipose tissue (BAT) and liver samples of Cth−/− mice fed CTRL or CysF diet for 5 days, determined by colorimetric assay (n = 7/group). e) Surface sensing of translation (SUnSET) assay was performed to assess protein synthesis. Immunoblot detection of puromycin incorporation in neosynthesized proteins in SFAT, visceral adipose tissue (VFAT), BAT and liver samples of Cth−/− mice fed with CTRL or CysF diet for 5 days (n = 6/group). f) Heatmaps of RNA-seq based gene expression of genes involved in Fe-S cluster assembly in SFAT, VFAT and BAT of Cth+/+ and Cth−/− mice, fed with CTRL or CysF diet (n = 4/group). g) Representative EPR spectra of POBN-lipid radical adducts measured in Folch extracts of SFAT, VFAT and BAT tissues of Cth−/− mice fed with either CTRL or CysF diet. The six-line spectrum (red arrows) is consistent with carbon-centred lipid-derived radicals, indicative of lipid peroxidation (identified through hyperfine coupling constants aN = 15.75 ± 0.06 G and aβH = 2.77 ± 0.07 G). Data are expressed as mean±SEM. Box plots represent median value and extend to the 25th and 75th percentiles. Whiskers are plotted down to the minimum and up to the maximum value. Statistical differences were calculated by 2-way ANOVA with Sidak’s correction for multiple comparisons, or by unpaired two-tailed t-tests.
