Fig. 6: Arginine activates mTOR pathway to regulate the level of acetyl-CoA and modulate histone acetylation.

a After starvation overnight, cells were treated with arginine and harvested at different time points as indicated. qPCR analysis shows the gene expression of ACLY and ACSS2, but not of ACSS1, were upregulated after arginine stimulation in a time-dependent manner. b After arginine starvation, cells were treated with arginine for 24 h. Western blotting shows that arginine stimulation (+) induced the protein expression of ACLY and ACSS2. c Cells were grown either in regular media supplemented with rapamycin or in arginine-free media for 24 h. ELISA data show that inhibition of mTOR activity via rapamycin treatment or arginine deprivation decreased the level of acetyl-CoA (blue) and histone H3 acetylation (purple). d Cells were first transduced with mTOR shRNA for 24 h, following an antibiotic selection for 1 week. Western blotting shows that the protein expression of ACLY and ACSS2 were downregulated after silencing of mTOR. e Cells were first transduced with ACLY or ACSS2 shRNA for 24 h, following an antibiotic selection for 1 week. Western blotting data show that the ratio of acetyl-histone H3 was decreased after silencing of ACLY or ACSS2. f After starvation overnight, cells were treated with arginine and harvested at different time points as indicated. qPCR analysis shows that arginine stimulation globally induced the histone acetyltransferase (HAT) expression. The color key indicates the relative gene expression to vehicle control (at time 0 h). The expression of ACLY and ACSS2 serves as an internal positive control. g After arginine starvation, cells were treated with arginine for 24 h. ELISA data show that arginine stimulation induced the histone H3 acetylation, specifically on lysine residues 9, 18, and 27, as well as H4 acetylation on lysine residues 5 and 12. h Cells were grown in regular media treated with KAT2B inhibitor (KAT2Bi), Garcinol, or in arginine-free media for 24 h. Seahorse data show that inhibition of KAT2B significantly suppressed the mitochondrial respiration activities. O oligomycin, F FCCP, R/A rotenone/antimycin. i Cells were first transduced with KAT2B shRNA for 24 h, following an antibiotic selection for 1 week. ChIP-qPCR shows the TEAD4 recruitment on OXPHOS promoter regions after silencing of KAT2B. j Seahorse data show that silencing of KAT7 significantly suppressed the mitochondrial respiration activities. O oligomycin, F FCCP, R/A rotenone/antimycin. k Cells were first transduced with KAT7 shRNA for 24 h, following an antibiotic selection for 1 week. ChIP-qPCR shows the TEAD4 recruitment on OXPHOS promoter regions after silencing of KAT7. l Diagram of hypothetical model. Data are presented as mean value ± SEM of independent experiments (n = 3 in a, c, f–k). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, using unpaired two-tailed Student’s t test. Source data are provided as a Source data file.