Fig. 3

H3.3K27M Mutations Cause an Overall Repression of Biological Pathways while the Effect on Gene Expression is Balanced. Differential gene expression and pathway responses in K27M and PRC2 mutants. (A) Venn diagram illustrating unique and shared down-regulated and (B) up-regulated genes in SF8628 cells with K27M mutation, and EZH1/2KO (PRC2 loss). DEGs were identified using DESeq2 with the following comparisons: K27M-associated: SF8628-H3.3K27M-EZH1/2WT vs. SF8628-H3.3WT-EZH1/2WT. EZH1/2KO associated: SF8628-H3.3WT-EZH1/2KO vs. SF8628-H3.3WT-EZH1/2WT. K27M + EZH1/2KO effects: SF8628-H3.3K27M-EZH1/2KO vs. SF8628-H3.3WT-EZH1/2WT. These comparisons allowed us to delineate distinct transcriptional programs uniquely regulated by H3.3K27M mutation versus PRC2 loss. (C–I) Gene ontology (GO) pathway enrichment analysis of genes unique to each condition or common across conditions, based on the intersections identified in panels A and B. Blue and red indicate down-regulated and up-regulated pathways, respectively. These analyses reveal that genes uniquely affected by K27M mutation are predominantly associated with pathway repression, whereas genes commonly regulated by PRC2 loss tend to drive pathway upregulation, consistent with the repressive function of PRC2