Extended Data Fig. 8: Immunity following PRMT9 inhibition requires cGAS activity.
a, Ifit1 expression in AML cells from BM scRNA-seq. b, GSEA analysis of BM T cells. Normalized enrichment scores and family-wise error rate P-value was determined by the GSEA permutation method. c, d, RNA-seq of common DEG in PRMT9 KD versus Ctrl cell lines. Heatmap (c) and GSEA (d) show upregulation of IFNα response genes upon PRMT9 KD. The color code represents z scores for differential gene expression. For (b) and (d), the normalized enrichment scores and family-wise error rate P-value was determined by the GSEA permutation method. The Normalized Enrichment Score is calculated by dividing the Enrichment Score from the actual ranking by the means of the random permutations. An enrichment P-value is calculated by comparing the observed frequency of an annotation term with the frequency expected by chance; individual terms beyond cut-off (p-value ≤ 0.05) are deemed enriched. e, IFI44 expression in Molm13 engineered as indicated (n = 5 independent experiments). Data were mean ± SD. P value was determined by one-way ANOVA. f, g, Levels of selected ISG genes (f) in AML and B-NHL lines after 2 days LD2 treatment (n = 3 independent experiments). Data were presented as mean ± SD. h, Lucia activity of engineered THP1 after LD2 treatment (n = 5 independent experiments). Data were mean ± SD. P value was determined by one-way ANOVA. i, Levels of PRMT9 and Flag-tagged ENPP1 in engineered THP1 cells, (n = 1). j, Levels of γH2AX in THP1 cells, (n = 1). k, dsDNA by immunostaining in THP1 cells after LD2 treatment for 72 hr. Violin plots (right) summarized dsDNA intensity (n = 50 cells/group). Scale bar, 10 μm. l, Immunostaining for γH2AX in Ctrl and PRMT9 KD THP1 cells. Violin plots (right) summarized γH2AX intensity (n = 100 cells/group). Scale bar, 10 μm. m, dsDNA by immunostaining in Ctrl and PRMT9 KD THP1 cells. Violin plots (right) summarized dsDNA intensity (n = 50 cells/group). Scale bar, 10 μm. P values (k,l,m) were calculated by unpaired two-sided t test. n, Levels of cGAS in Ctrl and cGAS-KO MA9 cells (n = 2 independent experiments). o, ENPP1 mRNA levels in the BEAT AML (n = 451 AML cases; n = 19 healthy donors). The P value was by unpaired two-sided t test. p-q, cGAS and ENPP1 expression in AML and B-NHL lines compared with other cancer cell lines. AML (n = 14), B-NHL (n = 9), LC (n = 77), COAD (n = 30), PAAD (n = 20), BRCA (n = 30), PRAD (n = 5), LIHC (n = 14), GBM (n = 12), BLCA, (n = 11) and ALL (n = 8). P values were by unpaired two-sided t test. ‘n’ represents number of cell lines. r, Cd80, Cd86, H2-ab1 expression in monocytes/macrophages from BM, from scRNA-seq. Ctrl, n = 631 cells; Prmt9 KD, n = 469 cells. Data were mean ± SEM. P values were determined by unpaired two-sided t-test. s, Representative spectrum of XRN2 R946 methylation in SILAC-based methyl-peptide quantitative LC-MS/MS. t, 293 T were transfected with Flag-tagged XRN2, then for immunoprecipitation and the interactor detected by immunoblot, (n = 1). γH2AX was detected in the input lysate.
