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. Author manuscript; available in PMC: 2022 Jun 29.
Published in final edited form as: Nat Metab. 2022 Jun 27;4(6):739–758. doi: 10.1038/s42255-022-00593-x

Extended Data Fig. 5. VHL restores cellular oxygen consumption rate.

Extended Data Fig. 5

(a) Seahorse XF-96 analysis of oxygen consumption rate (OCR). Mitochondrial respiration reflected by OCR levels was detected in 786-O cells with indicated genotype. The rates of basal respiration and maximal respiratory capacity were respectively quantified by normalization of amount of cells. One way ANOVA Tukey's Multiple Comparison Test. ****p <0.0001. (b) Seahorse XF-96 analysis of oxygen consumption rate (OCR) of 786-O cells with indicated VHL status transduced with lentiviral pL.KO shRNA targeting EGLN3 or no targeting control. The rates of basal respiration and maximal respiratory capacity were respectively quantified as described above. One way ANOVA Tukey's Multiple Comparison Test. ****p <0.0001. (c) Seahorse XF-96 analysis of oxygen consumption rate (OCR) of primary EGLN3+/+ and EGLN3-/- MEFs. The rates of basal respiration and maximal respiratory capacity were respectively quantified by normalization of amount of cells. One way ANOVA Tukey's Multiple Comparison Test. ***p <0.001, ****p <0.0001. (d) Seahorse XF-96 analysis of oxygen consumption rate (OCR) of primary EGLN3-MEFs of indicated genotype stably transduced with lentivirus encoding EGLN3 WT, catalytic death mutant or empty control. The rates of basal respiration and maximal respiratory capacity were respectively quantified as described above. ***p <0.001, ****p <0.0001. a-d, data are presented as mean values ± SD. n = 3 biological independent experiments. (e) Crystal violet staining of 786-O cells with indicated VHL status treated with high glucose (25 mM) or no glucose respectively for 36 hours. (f) Crystal violet staining of primary EGLN3+/+ and EGLN3-/- MEFs treated with 100 μM 3-bromopyruvic acid (3-BP) for 4 hours. (g) Crystal violet staining of primary EGLN3+/+ and EGLN3-/- MEFs treated with high glucose (25μM) or no glucose (0μM) respectively for 48 hours. (h) Crystal violet staining of 786-O cells with indicated VHL status transduced with lentiviral pL.KO shRNA targeting EGLN3 or no targeting control, treated with 100 μM 3-bromopyruvic acid (3-BP) for 4 hours.