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. 2015 Jan 29;6(9):7195–7208. doi: 10.18632/oncotarget.3131

Figure 6. NexrutineR induces oxidative stress and inhibits mTOR signaling in a RAPTOR/RICTOR-independent manner.

Figure 6

(A) Effect of NexrutineR (vehicle, 10, 20 μg/ml; 18 h) and NAC pre-treatment (5 mM; 1 h) on disruption of mTOR complex 1 and 2 formation in 1205Lu cells by co-IP using mTOR antibody followed by western blotting with anti-RICTOR, anti-RAPTOR, and anti-mTOR (1:1000) antibodies. 1205Lu whole cell lysate used as input control and pull-down with no antibody as negative control. (B) Evaluation of total ROS (% cells) in 1205Lu cells following NexrutineR treatment (10 μg/ml; 18 h), after transient transfection with siRNA targeting RAPTOR, RICTOR, or both RAPTOR and RICTOR. TBHP is the positive control for ROS induction. (C) Evaluation of total ROS by fluorescence microscopy (carboxy-H2DCFDA) following NexrutineR treatment (10 μg/ml; 18 h) and with NAC pre-treatment (5 mM; 1 h) in 1205Lu cells after transient transfection with siRNA targeting RAPTOR and RICTOR. TBHP is the positive control for ROS induction. Representative images of 3 independent experiments are shown. Significance values; *indicates p ≤ 0.05.