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. 2024 Jun 18;12:RP91611. doi: 10.7554/eLife.91611

Figure 3. PPM1D-mutant cells have altered mitochondrial function.

(A) Mitochondrial mass of wild-type (WT) and PPM1D-mutant leukemia cells was determined using MitoTracker Green (100 nM) and the mean fluorescence intensity was analyzed by flow cytometry. Data represents mean ± SD of triplicates. At least three independent experiments were conducted with similar findings; unpaired t-tests. (B) Immunoblot of WT and PPM1D-mutant cell lysates probed with the human OXPHOS antibody cocktail (1:1000) and vinculin (1:2000). (C) Measurement of mitochondrial oxygen consumption rate (OCR) by seahorse assay in WT and PPM1D-mutant OCI-AML2 cells after treatment with oligomycin (1.5 µM), FCCP (0.5 µM), and rot/AA (0.5 µM). Quantification of basal, maximal, and ATP-linked respiration are shown. Data shown are the mean ± SD of technical triplicates. (D) Mitochondrial membrane potential of WT and PPM1D-mutant OCI-AML2 cells was measured using MitoTracker CMXRos (400 nM). The mean fluorescence intensity (MFI) was measured and analyzed by flow cytometry. Data represents mean ± SD of triplicates, unpaired t-test, ns = non-significant (p>0.05), *p<0.05, **p<0.01.

Figure 3—source data 1. Western blot of mitochondrial proteins in WT and PPM1D-mutant cells.

Figure 3.

Figure 3—figure supplement 1. PPM1D-mutant cells have altered mitochondrial function.

Figure 3—figure supplement 1.

(A,B) Measurement of mitochondrial oxygen consumption rate (OCR) by seahorse assay in wild-type (WT) vs. PPM1D-mutant MOLM-13 (A) and OCI-AML3 (B) cells after treatment with oligomycin (1.5 µM), FCCP (0.5 µM), and rot/AA (0.5 µM). Quantification of basal, maximal, and ATP-linked respiration shown. Each cell line was performed in technical triplicates, Student’s t-test. (C) Growth curves of WT and PPM1D-mutant leukemia cell lines at 24, 48, and 72 hr. Cell counts were normalized to day 0. ns = non-significant (p>0.05), *p<0.05, ***p<0.001.