Fig. 3: Methionine restrains PD-1 via H3K79 methylation.

a Heat maps showing mRNA levels of antitumoral immunity-related proteins in CD4 T cells cultured in CM and DM alone or DM supplemented with methionine (n = 2 biological samples). b GSEA plot showing enriched PD-1 neighborhood genes in DM-cultured CD4 T cells. Enriched PD-1 neighborhood genes in CD4 T cells were downregulated by methionine supplementation (n = 2 biologically independent samples). c Volcano plot representing metabolic changes in CD4 T cells cultured in TM alone or TM supplemented with methionine (n = 3 independent samples per group). d Analysis of intracellular methionine, S-adenosyl methionine (SAM), and S-adenosyl homocysteine (SAH) concentrations via Liquid chromatography-mass spectrometry (LC-MS) in CD4 T cells cultured in CM, TM alone, and TM supplemented with methionine (n = 3 independent samples per group). e Immunoblots of PD-1 expression in CD4 T cells cultured for 72 h in CM, DM, or DM supplemented with methionine and its metabolites. The experiment was repeated two times. f Effects of TM and methionine treatments on the histone methylation profiles of cultured CD4 T cells. The experiment was repeated three times. g Immunoblots for H3K79me2 in CD4 T cells cultured in TM with methionine and its metabolites. The experiment was repeated two times. h Effects of the TME and methionine treatment on histone methylation profiles. Immunoblots of CD4 T cells isolated from tumor-free (n = 6), PBS-treated tumor-bearing (n = 6), and methionine-treated tumor-bearing mice (n = 6). The experiment was repeated two times. i Histone methylation profiles of tumor-infiltrated CD4 T cells isolated from SLC43A2-intact and SLC43A2 KO tumor-bearing mice (n = 6 in each group). The experiment was repeated two times. j PD-1 and H3K79me2 immunoblots in CD4 T cells with DOT1L (EPZ004777) inhibitor treatment. The experiment was repeated two times. PD-1 MFI in CD4 T cells via FACS after DOT1L inhibitor treatment (n = 6 per group). Statistical analyses were performed using two-tailed Student’s t test & TMM + CPM normalized method for correction (b), one-way analysis of variance (d, j) and Tukey HSD test (posthoc) after ANOVA (c). Data are presented as mean ± standard error of the mean. Source data are provided as a Source Data file.