Figure 5.

SLC7A11-AS1 Prevents NRF2 Proteasomal Degradation to Defend ROS

(A) Western blot analysis of NRF2 in PDAC cell lines. (B and C) Effects of SLC7A11-AS1 knockdown on NRF2 expression were determined by qRT-PCR (B) and western blot (C) in SLC7A11-AS1-overexpressed PDAC cells (n = 3). (D) Effect of SLC7A11-AS1 knockdown on half-life of NRF2 was determined by western blot in PANC-1 cells treated with CHX (100 μg/mL). (E) Quantification of NRF2 was normalized to the loading control and expressed relative to 0 h. (F) NRF2 protein levels in the nucleus and cytosol in PANC-1 cells with SLC7A11-AS1 knockdown or sh-Ctrl were determined by western blot. (G) Quantifications of nuclear and cytoplasmic NRF2 were normalized to the loading control and expressed relative to sh-Ctrl. (H) Western blot analysis of nuclear and cytoplasmic fractions from SLC7A11-AS1-knockdown and control PANC-1 cells treated with or without MG132 (1 μM) for 24 h. (I) Quantification of nuclear NRF2 as in (H) (n = 3). (J–L) PANC-1 cells with or without siRNA-mediated NRF2 knockdown were transfected with SLC7A11-AS1 or empty vector for 48 h, followed by ROS detection using flow cytometry (J), qRT-PCR analysis of NRF2 target genes, HMOX1 (K), and GCLM (L). *p < 0.05, **p < 0.01, ***p < 0.001. ns, not significant.