FIGURE 6.

Effect of Siah2 knockdown on Nrf2 accumulation and transcription of Nrf2-responsive genes. A, WT Hep3B, MOCK (GFP shRNA control), or Siah2 knockdown Hep3B, HEK293 and HeLa cells were grown under normoxia or hypoxia for 6 h. Two independent Siah2 knockdown clones were used in each cell line. Total cell lysates were immunoblotted using anti-Nrf2, anti-HIF-1α, or anti-β-actin antibodies. Four dots with cross, effect of transfection with the Siah2 pBAsi plasmid are indicated by HIF-1α blots. B, MOCK (GFP shRNA control) or Siah2 knockdown Hep3B cells were exposed to hypoxia for 6 h. Total RNA was isolated and Siah2, HO-1, or NQO-1 mRNAs were determined using qRT-PCR. Values are expressed as the mean ± S.D. of three replicates. **, p < 0.01; *, p < 0.05 significantly different from MOCK.