(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.