Fig. 5: Arginine modulates OXPHOS pathway and TEAD4 recruitment is mTOR dependent.

a Gene set enrichment assay (GSEA, v4.1.0) of microarray data was performed using hallmark gene set collection. Normalized enrichment scores (NES) for pathways indicate the significant difference in the arginine-stimulation group. b GSEA shows that the genes associated with oxidative phosphorylation (upper panel) and mTORC1 pathway (lower panel) are enriched in the arginine-stimulation Arg (+) group. c After starvation overnight, cells were treated with arginine and harvested at different time points as indicated. Western blotting shows that arginine stimulation induced the phosphorylation of mTORC1 (T2446 and S2448) and its downstream target, phosphor-p70 S6K. d qPCR analysis of OXPHOS gene expression after mTOR activation by arginine stimulation and mTOR inactivation by rapamycin treatment (24 h) or shRNA knockdown. The color key indicates the relative gene expression to vehicle or scramble control. e–g Seahorse assay shows that activation of mTOR by 24-h arginine stimulation (e) increases the mitochondrial respiration activity. By contrast, inactivation of mTOR by 24-h rapamycin treatment (f) or shRNA knockdown (g) suppressed the mitochondrial respiration activity. O oligomycin, F FCCP, R/A rotenone/antimycin. h ChIP-qPCR of TEAD4 on OXPHOS gene promoter region shows that silencing of mTOR disrupted the TEAD4 recruitment after arginine stimulation. i Immunofluorescent staining of TEAD4 shows that inhibition of mTOR via rapamycin treatment induced TEAD4 cytosolic translocation (scale bar = 20 μm). The quantification of immunofluorescent staining is shown in j. Data are presented as mean value ± SEM of independent experiments (n = 3 in d–h, j). *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.