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. 2021 May 15;41(7):560–575. doi: 10.1002/cac2.12158

FIGURE 6.

FIGURE 6

HIF‐1α up‐regulates ALYREF expressions in BCa cells. (A) Bioinformatic analysis identifies HIF‐1α as the potential transcription factor and ‘ACGTGC’ as the consensus hypoxia response element for ALYREF. (B) ALYREF and PKM2 mRNA levels are up‐regulated in 1% O2 group as compared with 20% O2 group by qRT‐PCR. 5637 and T24 cells were exposed to 20% or 1% O2 for 24 h. (C) HIF‐1α, ALYREF, and PKM2 proteins are up‐regulated in 5637 and T24 cells exposed to 1% O2 as compared with those exposed to 20% O2 for 48 h by Western blotting. (D) Relative luciferase activity was measured in 293T cells transfected with reporter plasmids containing either wild‐type or mutant ALYREF promoter fragments. Then cells were exposed to 20% or 1% O2 for 48 h. (E) Relative luciferase activity was measured in 293T cells transfected with reporter plasmids containing PKM2 promoter fragments. Then cells were exposed to 20% or 1% O2 for 48 h. The ratio of luciferase activity was determined. * P < 0.05, ** P < 0.01, *** P < 0.001. Abbreviations: HRE, hypoxia‐responsive element. Abbreviations: BCa, bladder cancer; HIF‐1α, hypoxia‐ inducible factor‐ 1‐alpha; ALYREF, Aly/REF export factor; PKM2, Pyruvate kinase muscle isozyme M2; Wt, wild type; Mut, mutation; ALY‐WT, Aly/REF export factor wild type; ALY‐MUT, Aly/REF export factor mutation; hLuc, fireflyer luciferase; Rluc, Renillar luciferase.