Convergent alterations in the phosphoproteome status after resveratrol and/or spermidine treatment. (A–C) Human colon carcinoma HCT 116 cells were treated for 2 h with vehicle (Co, control), 100-µM resveratrol (Resv), and 100-µM spermidine (Spd), alone or in combination (Resv + Spd). (A) Representative phosphoprotein arrays are shown. (B) Clustering analysis for the effects on protein kinase phosphorylation. (C) Representative immunoblots of selected kinases whose phosphorylation status was unaffected (PRKAA1, RPS6KB1, and acetyl-CoA carboxylase a [ACACA]) or affected by resveratrol or/and spermidine treatment (PTKB, AKT1, MAPK8, and CDKN1B), validating phosphoprotein array data. (D) Human colorectal carcinoma HCT 116 cells were transfected with a GFP-LC3–encoding plasmid, cultured in complete medium for 24 h, and then treated with either vehicle or the indicated dose of resveratrol or spermidine, alone or in combination, for 2 h. Quantitative data. Bars depict the percentages (means ± SD; n = 3; *, P < 0.05) of cells showing the accumulation of GFP-LC3 in puncta (GFP-LC3^vac). GAPDH, glyceraldehyde 3-phosphate dehydrogenase.