a, Protein from indicated brain lysates are shown, with NRF1 levels normalized to actin below. b,
Nrf1 and Nrf2 and c,
PSM gene transcript levels from brain tissues in a are shown as mean ± s.e.m. of triplicate samples relative to Tsc2+/+ sample 1. d, Mice fasted overnight were refed (6 h) following 30-min pretreatment with vehicle or rapamycin (10 mg/kg). Protein from liver lysates are shown, with NRF1 levels normalized to actin graphed as mean ± s.e.m. relative to fasted mice (n=4 per condition). *P<0.05, ##P<0.01. e, Transcript levels from liver tissues in d are shown as mean ± s.e.m. relative to fasted mice. *P<0.05, #P<0.05. f,
Tsc2−/− MEFs expressing TSC2 or empty vector transfected with Nrf1 (a and b) or control siRNAs were serum starved 16 h with vehicle or 20 nM rapamycin or treated 1 h with 100 nM bortezomib. Amino acid levels are shown as mean ± s.e.m. of triplicate samples relative to TSC2-expressing cells. *P<0.05 compared to TSC2-expressing cells, ##P<0.01 or ###P<0.001 compared to vehicle-treated vector-expressing cells. g, Rates of protein synthesis in cells treated as in f are shown as mean ± s.e.m relative to TSC2-expressing cells (n=3). **P<0.01 compared to TSC2-expressing cells; #P<0.05 or ##P<0.01 compared to vehicle-treated vector-expressing cells. h, Cells treated as in f were switched to low or high amino acid media overnight, and rates of protein synthesis are shown as the mean ± s.e.m relative to vehicle-treated cells (n=3). ***P<0.001 compared to vehicle-treated low AA cells; #P<0.05 or ##P<0.01 compared to vehicle-treated high AA cells. d–h, Statistical significance for pairwise comparisons evaluated with a two-tailed Student’s t test.