Fig 1: Both AAV.ULK1.DN and the ULK1 inhibitor SBI-0206965 enhance mTOR activation.Lysates were obtained from E18 rat cortical neurons on DIV 8 after transduction with AAV.ULK1.DN or AAV.CTRL. Alternatively, cells were treated with the ULK1 inhibitor SBI-0206965 (SBI, 5 µM) or DMSO as control for 30 min before lysis. RAP addition of rapamycin (750 nM) 24 h before lysis. a–h Top: Representative immunoblots of mTOR, p-mTOR, S6, p-S6, 4E-BP1, p-4E-BP1, AMPK, p-AMPK, and the corresponding bands of the loading controls tubulin or GAPDH after transduction with AAV.ULK1.DN or AAV.CTRL are shown. Bottom: Quantifications of the band intensities of mTOR, p-mTOR, p-S6, AMPK (all n = 6 independent cultures), p-4E-BP1 (n = 5 independent cultures), p-AMPK (n = 4–5 independent cultures), 4E-BP1 (n = 4 independent cultures), and S6 (n = 3 independent cultures) normalized to tubulin or GAPDH as loading controls. CTR control, RAP treated with rapamycin. i–p Top: Representative immunoblots of mTOR, p-mTOR, S6, p-S6, 4E-BP1, p-4E-BP1, AMPK, p-AMPK, and the corresponding bands of the loading control GAPDH after treatment with SBI or DMSO as control are shown. Bottom: Quantifications of the band intensities of mTOR, p-mTOR, p-S6, AMPK, p-AMPK, 4E-BP1, p-4E-BP1 (all n = 5 independent cultures), and S6 (n = 4 independent cultures) normalized to GAPDH as loading control. CTR control, RAP treated with rapamycin. Data are presented as single data points and means ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, N.S. no significant difference, according to one-way ANOVA and Sidak’s multiple comparisons test.