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. Author manuscript; available in PMC: 2022 Jan 12.
Published in final edited form as: Nat Metab. 2021 Jul 12;3(8):1091–1108. doi: 10.1038/s42255-021-00422-7

Fig. 4. Mitochondrial defects are a common feature of cells eliminated by cell competition.

Fig. 4

a, Metabolic enrichment analysis of the TCA cycle and intermediate metabolites obtained using Metabolon platform for defective cells (Bmpr1a-/-, left bar and 4n, right bar), in comparison to wild-type cells during differentiation. Bars indicate compound levels relative to wild-type cells. Blue bars indicate compounds that are significantly depleted (p<0.05) and light blue bars indicate compounds that are almost significantly depleted (0.05≤p≤0.1). Black bars indicate compounds that are depleted although not statistically significant in comparison to the levels found in wild-type cells. The enzymes on the pathway are represented as boxes and labelled by their canonical names. b-e, Metabolic flux analysis of wild-type and BMP-defective cells during differentiating conditions. Analysis of oxygen consumption rate (OCR) as a measure of mitochondria function (mitochondria stress test) (b). Detail of metabolic parameters found changed from the analysis of the mitochondria stress test (c). Analysis of extracellular acidification rate (ECAR) as a measure of glycolytic function (glycolysis stress test) (d). Detail of metabolic parameters found changed from the analysis of the glycolysis stress test (e). f-g, Δψm in defective mESCs undergoing differentiation in separate or co-culture conditions. Representative histograms of TMRM fluorescence and quantification for wild-type and Bmpr1a-/- (f) and wild-type and 4n (g). h, Representative micrographs of wild-type and Bmpr1a-/- cells co-cultured during differentiation and stained for a reporter of Δψm (MitoTracker Red, top panel) or mitochondria mass (TOMM20, bottom panel). Nuclei are stained with Hoechst. Scale bar = 10 μm. i-j, Western blot analysis of mitochondria mass markers TOMM20 (i) and mt-CO1 (j) for wild-type and Bmpr1a-/- cells during differentiation. k, Analysis of Δψm for wild-type, Bmpr1a-/- and Bmpr1a-/-;p53-/- cells during differentiation. Representative histogram of TMRM fluorescence and quantification. Data shown as mean ± SEM. Extracellular flux Seahorse data was obtained from 3 (d,e) or 4 independent experiments (b,c), with 5 replicates per cell type in each assay. Remaining was data obtained from 3 (g,j), or 5 (a,f,i,k) independent experiments. See methods for details on statistical analysis.