Fig. 5

SHMT2 acetylation inhibits cell proliferation and tumor growth. a SHMT2-K95Q is compromised to support cell proliferation. SW480 cells stably knockdown SHMT2 with re-express the shRNA-resistant wild-type or K95Q mutant were established. SHMT2 knockdown efficiency and re-expression were determined by western blot. SHMT2 WT or SHMT2 K95Q cells were seeded in 12-well plates. Cell numbers were counted every 24 h. b SHMT2 K95Q increases intracellular serine level and serine/glycine ratio. Intracellular serine and glycine amount were measured by GC-MS. c SHMT2 K95Q is defective in supporting tumor growth in vivo. Xenograft was performed using the SHMT2-knockdown SW480 cells with reexpressing wild-type or K95Q mutant SHMT2 as indicated. ***P < 0.001. d SHMT2 K95Q decreases cellular NADPH level. SHMT2 WT or K95Q-rescued HCT116-SHMT2-knockout cells were established and confirmed by western blot. Intracellular NADPH was determined by using NAD(P)H-Glo Detection System. e SHMT2 K95Q increases cellular ROS level. SHMT2 WT or K95Q-rescued HCT116-SHMT2-knockout cells were seeded in confocal dishes and ROS level was measured by adding 10 μM H2DCF-DA. Fluorescent strength per unit area was quantified using the ImageJ software, followed by statistical analysis. f Sirt3-knockout mice developed less tumors in the AOM-DSS CRC mouse models. Sirt3 WT and KO mice were subjected to AOM-DSS colitis-associated cancer model. The representative images of colon tumors are shown. Tumor number of each group were shown. n = 8 (WT), n = 9 (Sirt3_KO), P = 0.0027 by student’s t-test. The expression of Sirt3 and Shmt2 in the colon tumors from WT and Sirt3_KO mice were detected by IHC. For a, d, and e, mean values ± s.d. of triplicate experiments are reported. *P < 0.05; **P < 0.001; ***P < 0.001