Figure 3

SPC activates Akt and mTOR in Mel-Ab cells. (A) After serum starvation for 24 h, Mel-Ab cells were treated with 10 μM SPC for the indicated times. Whole cell lysates were analyzed by Western blotting with antibodies against phospho-Akt, Akt, phospho-mTOR, mTOR, and actin (loading control). (B) Cells were incubated with 10 μM SPC for the indicated time periods and lysed. The levels of LC3 II were analyzed by Western blotting. (C) Cells were pretreated with 100 nM rapamycin for 1 h prior to the addition of 10 μM SPC. Cells were incubated for an additional 4 d and then the cellular melanin contents were analyzed. Data are expressed as the mean ± SD of triplicate assays. * P < 0.05 compared to the SPC-treated cells. (D) Cells were preincubated with 100 nM rapamycin for 1 h prior to addition of 10 μM SPC and then incubated for another 3 d. Whole cell lysates were analyzed by Western blotting with antibodies against LC3 II and actin (loading control).