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. 2020 Aug 6;11(8):649. doi: 10.1038/s41419-020-02825-y

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 3 — a Cisplatin-induced MTHFD2 K88 acetylation. Flag-tagged MTHFD2 was expressed, which were then treated with NAM (5 μm) for 4 or 6 h and cisplatin (40 μm) for 12 or 24 h. Flag-MTHFD2 was immunoprecipitated with Flag beads and immunoblotting was performed with the antibodies indicated. Relative acetylation was normalized by Flag protein. b Cisplatin treatment increases MTHFD2 WT acetylation levels but not K88R/K88Q mutant acetylation levels. Flag-tagged WT MTHFD2 or mutants (K88R and K88Q) were expressed in HCT116 cells, followed by treatments with or without 40 μm NAM for 24 h. Flag-MTHFD2 was immunoprecipitated and its acetylation was examined with α-AcK. c, d Cisplatin treatment increases MTHFD2 acetylation in a dose and time-dependent manner. Acetylation levels were detected of ectopically expressed MTHFD2(left) and endogenous MTHFD2 (right) after treated with cisplatin for the duration indicated at different concentrations for 24 h c or at a concentration of 40 μm for different time d. e, f Western blot analysis of SIRT3 protein levels in control and cisplatin-treated HCT116 cells. Cisplatin treatment decreases SIRT3 protein levels in a dose and time-dependent manner. HCT116 cells were treated with cisplatin for the duration indicated at different concentrations for 24 h e or at a concentration of 40 μm for different time f. g, h Real-time PCR analysis of the expression of SIRT3 mRNA. Cisplatin treatment decreases SIRT3 mRNA levels in a dose and time-dependent manner. HCT116 Cells were treated with cisplatin for the duration indicated at different concentrations for 24 h g or at a concentration of 40 μm for different time h, followed by statistical analysis. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001; n.s. not significant.