Fig. 6: miR-433-3p was the downstream target of NORAD to regulate autophagy. | Cell Death & Disease

Fig. 6: miR-433-3p was the downstream target of NORAD to regulate autophagy.

From: Oxidative stress activates NORAD expression by H3K27ac and promotes oxaliplatin resistance in gastric cancer by enhancing autophagy flux via targeting the miR-433-3p

Fig. 6

A LC3B I/II expression in NORAD knockdown SGC-7901-R and KATO III-R cells: LC3B I/II 16/14 kDa, p62 62 kDa, GAPDH 37 kDa. B Autophagy flux was decreased by knocking down NORAD (SGC-7901-R, t = 4.241, P = 0.043; KATO III-R, t = 3.992, P = 0.036) C Expression of autophagy-related genes in NORAD knockdown SGC-7901-R and KATO III-R cell line: ATG5(ATG5-ATG12 complex) 55 kDa, ATG12(free ATG12) 16 kDa, ATG 78 kDa, GAPDH 37 kDa, ATG10 25 kDa, β-Tubulin 55 kDa. D Starbase prediction for the binding site of NORAD to miR-433-3p; and Starbase prediction for the binding site of miR-433-3p to ATG5 and ATG12. E qRT-PCR to detect the expression of miR-433-3p in gastric cancer compared with GES-1. One-way ANOVA: GES-1 vs SGC-7901, q = 10.37, P < 0.000; GES-1 vs AGS, q = 10.26, P < 0.000; GES-1 vs SNU-1, q = 6.274, P = 0.000; GES-1 vs HGC-27, q = 5.816, P = 0.000; GES-1 vs KATO III, q = 12.27, P < 0.000. F qRT-PCR to detect the expression of miR-433-3p in SGC-7901-R compared with SGC-7901 and KATO III-R compared with KATO III (SGC-7901 vs SGC-7901-R, t = 40.75, P = 0.000; KATO III vs KATO III-R, t = 23.00, P = 0.000). G qRT-PCR to detect the influence of NORAD knockdown on the expression of miR-433-3p in SGC-7901, SGC-7901-R, KATO III, and KATO III-R. (SGC-7901, t = 10.72, P < 0.000; SGC-7901-R, t = 17.31, P < 0.000; KATO III, t = 6.778, P < 0.000; KATO III-R, t = 12.98, P < 0.000). H Dual-luciferase reporter gene assay for NORAD and miR-433-3p (NORAD-Wt vs NORAD-Mut in miR-433-3p group: SGC-7901, t = 10.76, P < 0.000; SGC-7901-R, t = 7.986, P < 0.000; KATO III, t = 6.778, P < 0.000; KATO III-R, t = 12.98, P < 0.000). I RNA pull-down assay for the expression of NORAD in miR-433-3p-Wt and miR-433-3p-Mut group in SGC-7901, SGC-7901-R, KATO III, and KATO III-R (miR-433-3p-Wt vs miR-433-3p -Mut: SGC-7901, t = 9.321, P = 0.001; SGC-7901-R, t = 8.962, P = 0.001; KATO III, t = 10.60, P = 0.000; KATO III-R, t = 19.07, P = 0.000). J RIP assay to detect the enrichment of miR-433-3p (NORAD-Wt vs NORAD-Mut: SGC-7901, t = 32.111, P = 0.000; SGC-7901-R, t = 15.12, P = 0.000; KATO III, t = 24.21, P = 0.000; KATO III-R, t = 13.22, P = 0.000).

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