Extended Data Fig. 6: NNMT upregulation in fibroblasts reduces H3K27me3 methylation.
From: NNMT inhibition in cancer-associated fibroblasts restores antitumour immunity
a, scRNA-seq data analysis of CAFs from Luo et al.21, including CAFs from ovarian, breast, colon, and other cancers. Pearson correlation coefficients of log-normalized gene expression values are shown for genes and NNMT across approximately 800,000 single CAFs (visualized using RAWGraphs 2.0). NNMT expression positively correlates with 1,265 genes, with complement factors highlighted in red. b, Fibroblasts were isolated from the ovaries of tumour-naïve Nnmt−/− mice, and NNMT expression was restored via lentiviral transduction. Western blot analysis confirmed NNMT expression in NNMT-expressing ovarian fibroblasts and Nnmt−/− control cells. c, Expression of several CAF markers in NNMT-expressing 3T3 fibroblasts (3T3-NNMT) and control (CTRL) cells was analysed by flow cytometry (n = 3 technical replicates) (two-tailed unpaired Student’s t-test). MFI = mean fluorescence intensity, FMO = fluorescence minus one. d, Integrative Genomics Viewer visualization of H3K27me3 peaks for the CAF markers C3, Mylk, Fbln1, Saa3, and their adjacent genes (Genome Reference Consortium GRCm38, y-axis scale 0-36). e, Cell size (FSC-A) of 3T3-NNMT and 3T3-CTRL cells was measured by flow cytometry (n = 3 technical replicates) (two-tailed unpaired Student’s t-test). f, 3T3-NNMT and 3T3-CTRL cells were grown in either low- or high- L-methionine media for 24 h before RNA extraction, and C3 and Cfb expression levels were measured by qPCR (n = 3 technical replicates) (ordinary one-way ANOVA with Tukey’s multiple comparisons test). Data are shown as mean ± SEM. One experiment representative of at least two independent experiments is shown for panels b, c, e, and f.
