Fig. 3: RNA-seq reveals cooperative perturbation of intra-GC transcriptional transitions by CREBBP and KMT2D haploinsufficiency. | Nature Communications

Fig. 3: RNA-seq reveals cooperative perturbation of intra-GC transcriptional transitions by CREBBP and KMT2D haploinsufficiency.

From: Loss of CREBBP and KMT2D cooperate to accelerate lymphomagenesis and shape the lymphoma immune microenvironment

Fig. 3

a Experimental scheme for RNA-seq profiling of CB and CC (results shown in b–h). 8-weeks, female C57BL/6J background mice were used. b PCA of RNA-seq datasets. WT/C/K/CK: n = 4/4/3/3 mice (CB), n = 4/4/4/3 mice (CC). c Dendrogram showing the hierarchical clustering result of RNA-seq datasets using the top 10% most variable genes, the Manhattan distance and ward.D2 linkage. d, e Heatmap showing the relative expression levels of the union differentially expressed genes (DEGs) as log2FC (vs mean WT expression) for each genotype in (d) CB and (e) CC. Union DEGs include DEGs defined in at least one pair-wise comparison using WT as control with a significance cut-off of padj < 0.01, |log2FC| > 0.58. Scale factors, based on single-cell RNA-seq UMI counts, were applied to account for total mRNA difference. Each column represents one mouse dataset. f Pathway enrichment analysis for CK vs WT DEGs using Parametric Analysis of Gene Set Enrichment (PAGE). g Fuzzy c-means clustering of RNA-seq datasets identified 8 clusters (named as Traj_1 to Traj_8) with distinct trajectory patterns, Traj_3 and Traj_4 are shown: line plot (left) and heatmap (right) of log2FC expression (vs mean of WT CB). Black lines in the line plot are cluster centroid; genes are colored by the degree of cluster membership. A linear regression model was fit for log2FC expression compared to WT as a function of cell type (CB or CC), genotype (C, K, or CK), and interaction of cell type and genotype. The corresponding p values for the coefficients are shown. h Pathway enrichment analysis using PAGE for Traj_3 and Traj_4 genes. i, RT-qPCR of indicated genes in sorted GC B cells for each genotype (n = 4 mice per genotype). qPCR signal was normalized as log2FC (vs mean WT) and presented as mean ± SEM. P values were calculated by one-tailed hypergeometric test (f, h) and ordinary one-way ANOVA followed by Tukey–Kramer’s multiple comparisons test (i). Source data are provided as a Source Data file.

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