Fig. 2

Hypoxic mRNA stability does not require HIF transcriptional activity. HEK293T cells were simultaneously transfected with siRNAs targeting ARNT1 and 2 (ARNT KD), HIF-1α and 2α (HIF KD) or negative control siRNA (Neg) for 48 h before being exposed to 24 h of Normoxia or Hypoxia (1% O₂) in the presence of 1 g/L glucose. Cells were treated with actinomycin D for 0, 1 or 2 h. RT-PCR (a, d) and western blotting (b, e) of 0 h time point were used to assess effectiveness of knockdown. c, f mRNA levels of VEFG, MYC and CYR61 were determined via qPCR, normalized to GAPDH and 0 h time point and half-lives determined. Data represents average of N = 3 ± SD for each condition. *P ≤ 0.05; @P ≤ 0.1 from respective control