Figure S4.
Changes in ECAR in MEFs after mitochondrial inhibitor treatments. (A) ECAR (± SD) upon 100 µM CK666, 1 µM CCCP or 1 µM CCCP + 100 µM CK666 addition (15 min), followed by 50 mM 2-DG (59 min) in 25 mM glucose medium without serum. n = 3 individual well measurements for CCCP and CK666; 4 for CCCP + CK666. Pink arrow indicates drug treatment and blue arrow indicates 2-DG treatment. (B) ECAR (± SD) upon DMSO, 100 µM CK666, 2.5 µM antimycin A or 2.5 µM antimycin A + 100 µM CK666 addition (33 min), then 50 mM 2-DG (258 min) in 25 mM glucose medium without serum. n = 5 individual well measurements per condition. Pink arrow indicates drug treatment and blue arrow indicates 2-DG treatment. (C) ECAR (± SD) upon DMSO, 100 µM CK666, 5 µM rotenone or 5 µM rotenone + 100 µM CK666 addition (33 min), then 50 mM 2-DG (258 min) in 25 mM glucose medium without serum. n = 5 individual well measurements per condition. Pink arrow indicates drug treatment and blue arrow indicates 2-DG treatment. (D) P values for comparisons between individual curves in Fig. 3 F. Statistical significance was calculated by two-way ANOVA using Tukey’s multiple comparisons test. (E) Effect of glucose concentration (± SD) on prolonged ECAR increase (after 40 min) induced by 1 µM CCCP or 1 µM CCCP + 100 µM CK666 in MEFs. n = 9 individual well measurements for CCCP and CK666; 12 for CCCP + CK666. P values graphed in F. (F) P values for comparisons between individual curves in E. Statistical significance was calculated by two-way ANOVA using Tukey’s multiple comparisons test. Number of experiments, statistical tests, and sample sizes are provided in Table S1.

Changes in ECAR in MEFs after mitochondrial inhibitor treatments. (A) ECAR (± SD) upon 100 µM CK666, 1 µM CCCP or 1 µM CCCP + 100 µM CK666 addition (15 min), followed by 50 mM 2-DG (59 min) in 25 mM glucose medium without serum. n = 3 individual well measurements for CCCP and CK666; 4 for CCCP + CK666. Pink arrow indicates drug treatment and blue arrow indicates 2-DG treatment. (B) ECAR (± SD) upon DMSO, 100 µM CK666, 2.5 µM antimycin A or 2.5 µM antimycin A + 100 µM CK666 addition (33 min), then 50 mM 2-DG (258 min) in 25 mM glucose medium without serum. n = 5 individual well measurements per condition. Pink arrow indicates drug treatment and blue arrow indicates 2-DG treatment. (C) ECAR (± SD) upon DMSO, 100 µM CK666, 5 µM rotenone or 5 µM rotenone + 100 µM CK666 addition (33 min), then 50 mM 2-DG (258 min) in 25 mM glucose medium without serum. n = 5 individual well measurements per condition. Pink arrow indicates drug treatment and blue arrow indicates 2-DG treatment. (D) P values for comparisons between individual curves in Fig. 3 F. Statistical significance was calculated by two-way ANOVA using Tukey’s multiple comparisons test. (E) Effect of glucose concentration (± SD) on prolonged ECAR increase (after 40 min) induced by 1 µM CCCP or 1 µM CCCP + 100 µM CK666 in MEFs. n = 9 individual well measurements for CCCP and CK666; 12 for CCCP + CK666. P values graphed in F. (F) P values for comparisons between individual curves in E. Statistical significance was calculated by two-way ANOVA using Tukey’s multiple comparisons test. Number of experiments, statistical tests, and sample sizes are provided in Table S1.

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