Fig 2: GR-mediated SETD1A/COMPASS recruitment to enhancers shows locus-specific histone methylation(A) Normalized SETD1A, CXXC1, H3K4me1/me2/me3, and H3K27 acetylation (H3K27ac) ChIP-seq signals in LPS- and Dex + LPS-treated macrophages, ±2 kb around the GBS. Loci are sorted by clusters identified in Figure S4C. Invariant: GBSs with unchanged SETD1A (p > 0.1, -1.5 < FC < 1.5) occupancy, 588 bp around the GBS. ? represents the coverage difference between Dex + LPS and LPS treatments.(B) Log2FC in SETD1A, CXXC1, H3K4me1/me2/me3, and H3K27ac ChIP-seq signals at intergenic regions. mRNA expression of the genes closest to the enhancers from cluster 1, cluster 2, and cluster 3 is displayed together with those showing invariant SETD1A occupancy (inv., p > 0.1, -1.5 < FC < 1.5; gray). Black lines indicate the distribution mean. (ChIP-seq: Kruskal-Wallis test with post hoc Dunn’s test; RNA-seq: one-way ANOVA with post hoc pairwise two-sided t test, Benjamini-Hochberg adj). n = 2.(C) Normalized median genome browser tracks for H3K4me1/me2/me3 and H3K27ac ChIP-seq (n = 2, LPS- (red) versus Dex + LPS-treated (blue) BMDMs) plus GR. Arrowheads point at treatment-dependent changes; lines below GR peaks indicate primer locations for (D).(D) Normalized spike-in ChIP-qPCR for selected loci; mean Z scores of total H3 normalized % inputs. Error bars represent standard deviation (n = 3, two-sided Wilcoxon-Mann-Whitney test).(E) Correlation plot of log2FC in SETD1A, CXXC1, H3K4me1/me2/me3, and H3K27ac at intergenic GBSs with differential SETD1A occupancy (-1.5 > FC > 1.5, p < 0.1) and the mRNA expression of the closest gene. Pearson correlation coefficients with p < 0.001 are displayed as numbers.(F) Log2FC in mRNA expression of genes with nearby GBS over changes in SETD1A occupancy and/or H3K27ac levels (+, gain; -, invariant or lost). Each dot reflects one enhancer. Red: mean and 95% confidence intervals (Kruskal-Wallis test with post hoc Dunn’s test, Benjamini-Hochberg adj). Enhancer numbers are in parentheses.

Fig 3: Knockdown of SETD1A reduces GC cell metastasis. (A) Downregulation of SETD1A in BGC-823 cells reduces the number of tumors in the lung surface. Data from six mice are shown as the means ± SD. ***p < 0.001. (B) Downregulation of SETD1A in BGC-823 cells reduces tumor growth in the lung. Data from six mice are shown as the means ± SD. ****p < 0.0001 by Student’s t-test.

Fig 4: SETD1A Prevents Excessive ssDNA Formation after Replication Stress by Stabilizing RAD51(A–C) HeLa cells were transfected with the indicated siRNAs for 72 hr; exposed to 50 ng/mL MMC for 24 hr; immunostained with antibodies to RPA2 (A), phospho-RPA2 S4/S8 (B), or RAD51 (C). Focus formation was analyzed by fluorescence microscopy.(D) Cells from (A) were exposed to 4 mM HU for 5 hr and immunostained with antibodies to RAD51, and focus formation was analyzed as above.(E) Nuclear fluorescence intensity of cells from (A) was quantified using ImageJ. Lines denote mean values.(F) Double-positive (RPA- and phospho-RPA2) cells from (A) and (B) were enumerated and are displayed as a percentage of total cells.(G) The mean percentage of cells from (C) and (D) exhibiting RAD51 foci was enumerated as above.(H) Quantification of PLA signals between EdU and RAD51 in U-2-OS cells transfected with the indicated siRNAs. Where denoted, cells were exposed to 4 mM HU for 5 hr (as shown in the schematic). The mean ± SD of the number of biotin/biotin PLA signal-positive cells (below) indicates the number of S phase cells in each condition.The plots in (E)–(H) represent mean data ± SEM from 3 independent experiments. Scale bars, 10 µm. *p < 0.05, **p < 0.01, and ***p < 0.001. See also Figure S2.

Fig 5: Restoration of snail-associated EMT recovers GC cell migration and invasion inhibited by SETD1A knockdown. (A,B) Restoration of snail resulted in recovering EMT, cell migration, and invasion suppressed by SETD1A knockdown in BGC-823 (A) and AGS (B) cells. Data from three replicates are shown as the means ± SD. **p < 0.01, ***p < 0.001 by Student’s t-test.