XRN1 knockdown in SAOS-2 cells does not result in observable phenotypes. (A) Successful knockdown of XRN1 in SAOS-2 cells using RNAi 24 h post transfection. Scr samples treated with 20 pmol scrambled siRNA and KD cells treated with 20 pmol XRN1 siRNA. Error bars represent SEM, (***) P = 0.0008. (B) Quantification and representative images (40× objective) of the BrdU proliferation assay. Error bars represent SEM, n ≥ 25, P = 0.7938, scale bar = 50 µM. (C) WST-1 assay at 24 h time intervals following transfection with either Scrambled (Scr) or XRN1 (KD) siRNA. Error bars represent SEM, n = 3. (D) Caspase Glo 3/7 assay at 24 h time intervals following transfection with either Scrambled (Scr) or XRN1 (KD) siRNA. Error bars represent SEM, n = 3. (E) Quantification and representative images (20× objective) of transwell migration assay 6, 24, or 30 h post seeding. Seeding was performed 24 h post transfection with either Scrambled (Scr) or XRN1 (KD) siRNA. Error bars represent SEM, n = 4, P > 0.05, scale bar = 100 µM. (F) Knockdown of XRN1 does not affect nascent translation rates. Quantification of Puromycin incorporation or XRN1 expression (normalized to GAPDH relative to its own scrambled partner) 24 h post transfection in cells treated with either Scrambled (Scr) or XRN1 (KD) siRNA. Error bars represent SEM, (***) P = 0.0003, ns = P = 0.7432, n = 5.
