Extended Data Fig. 6: Loss of basal ATF4 levels in ΔuORF1 mouse ES Cells impairs cellular bioenergetics.
From: Plasticity of the mammalian integrated stress response
(a) Schematic of [3-13C]-pyruvate tracing in the citric acid cycle. Black-filled circles indicated [13C]-labelled carbons. Citrate m+1, Glutamine m+1, Succinate m+1, Malate m+1, and Aspartate m+1 refer to isotopomers of metabolites after incorporation of one [13C]-labelled carbon following one turn of the citric acid cycle. Citrate m+2, Glutamine m+2, Succinate m+2, Malate m+2, and Aspartate m+2, refer to isotopomers of metabolites after incorporation of two [13C]-labelled carbons after two turns of the citric acid cycle. (b, c) Fractional enrichment of the indicated metabolites (m+1, b and m+2, c) in WT and ΔuORF1 mouse ES Cells. p-values, two-tailed Student’s t-test. Data are presented as mean ± SEM (n = 4 independent experiments). (d) Protein synthesis rates in WT and ΔuORF1 mouse ES Cells were measured using the incorporation of [35S]-cysteine and methionine. No statistical significance (n.s.) according to the two-tailed Student’s t-test (n = 3 independent experiments). (e) Differential expression analysis (DESeq2) of comparing mRNA expression quantified by RNA sequencing in WT versus ΔuORF1 mouse ES Cells. Genes belonging to the ATF4-regulated signature20 are shown in red. Selected differentially expressed genes are indicated in green. All regulated genes can be found in Supplementary Table 1. (f) Steady state levels of metabolites (serine, glycine, PEP) in ΔuORF1 mouse ES Cells compared to WT (n = 3 independent experiments). Data depicts fold change of ΔuORF1 mouse ES Cells relative to WT.
