A. TSC2-/- p53-/- MEFs were deprived of glucose, amino acids, or both with or without rapamycin.
B. Cell viability of cell with Branched chain amino acids (BCAAs), L-glutamine, or total amino depletion.
C. Total cellular ATP levels were determined after 24 hours of glucose deprivation.
D. S6K-1 (T389) phosphorylation after 24 hours of deprivation.
E. L-glutamine was added to glucose and total amino acid deprived conditions at the time of deprivation, and viability was measured at 60 H post withdrawal.
F. A time course measuring the viability of TSC2-/- p53-/- MEFs grown in complete, (-) glucose, (-)Glu (-)AA, and (-)glu (-)AA plus 4mM L-glutamine media. The samples labeled with X were replenished with only glutamine (2mM) starting at 48 hours and replenished every 24 hours. The Rap X sample also got 20nM of rapamycin at 0 and 72 hours.
G. Phase images were taken at 3 days and 6 days with or without L-glutamine (2mM) retreatment in (-) glucose (-) amino acids conditions.
Shown is an average (+ SEM) of 3 independent experiments.