Fig. 3: OTUB1 augments hypoxia signaling pathway independent of HIF-1α hydroxylation. | Cell Death & Disease

Fig. 3: OTUB1 augments hypoxia signaling pathway independent of HIF-1α hydroxylation.

From: OTUB1 augments hypoxia signaling via its non-canonical ubiquitination inhibition of HIF-1α during hypoxia adaptation

Fig. 3

A, B qPCR analysis of PDK1 (A) and PGK1 (B) mRNA in H1299 cells transfected with or without Flag-OTUB1 and cultured under normoxia (21% O2) for 24 h, followed by treatment with DMSO or DMOG (1 mM) for 8 h. Flag empty vector was used as a control. Two-way ANOVA analysis; Data show mean ± SD; Tukey’s multiple comparisons test; ns, not significant, *Adjusted p < 0.05, **Adjusted p < 0.01, ***Adjusted p < 0.001, ****Adjusted p < 0.0001; Data from 3 independent experiments. C, D qPCR analysis of PDK1 (C) and BNIP3 (D) mRNA in H1299 cells transfected with or without Flag-OTUB1 and cultured under normoxia (21% O2) for 24 h, followed by treatment with DMSO or FG4592 (100 μM) for 8 h. Flag empty vector was used as a control. Two-way ANOVA analysis; Data show mean ± SD; Tukey’s multiple comparisons test; ns, not significant, *Adjusted p < 0.05, **Adjusted p < 0.01, ***Adjusted p < 0.001, ****Adjusted p < 0.0001; Data from 3 independent experiments. E Western blot analysis of indicated proteins in VHL-deficient or wildtype H1299 cells (VHL−/− or VHL+/+) cultured under normoxia (21% O2). F qPCR analysis of PDK1 and PGK1 mRNA in VHL-deficient or wildtype H1299 cells (VHL−/− or VHL+/+) cultured under normoxia (21% O2). Data show mean ± SD; Student’s two tailed t-test; ***p < 0.001; Data from 3 independent experiments. G qPCR analysis of PDK1, PGK1, LDHA, VEGF and PKM2 mRNA in VHL-deficient H1299 cells (VHL−/−) transfected with or without Flag-OTUB1 and cultured under normoxia (21% O2) for 24 h. Flag empty vector was used as a control. Data show mean ± SD; Student’s two tailed t-test; ***p < 0.001; Data from 3 independent experiments. H Western blot analysis of indicated proteins in FIH-deficient or wildtype HEK293T cells (FIH−/− or FIH+/+) cultured under normoxia (21% O2). I–K qPCR analysis of PDK1 (I), PGK1 (J) and GLUT1 (K) mRNA in FIH-deficient HEK293T cells (FIH −/−) transfected with or without Flag-OTUB1 and cultured under normoxia (21% O2) or hypoxia (1% O2) for 24 h. Flag empty vector was used as a control. Two-way ANOVA analysis; Data show mean ± SD; Tukey’s multiple comparisons test; ns, not significant, ****Adjusted p < 0.0001; Data from 3 independent experiments.

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