Panel A: A U M A P plot displays clusters of I g D-G C 1, I g D-G C 2, I g D-M E 1, I g D-M E 2, I g D-P C 1, and I g D-P C 2. The trajectory inference graph illustrates the developmental trajectories between these clusters. Below the UMAP plot, dot plots show the differential gene expression for each subcluster, with average expression levels and percentage expression indicated by the size and color of the dots. Panel B: A heatmap visualizes the differential expression of genes involved in somatic hypermutation (S H M) between tonsillar I g D-G C subcluster 1 and 2 versus I g G-G C B cells. Genes with significant differential expression are marked with an asterisk. Panel C: A box plot compares the normalized heights of lineage trees for different antibody contents (I g D, I g M, I g G, I g A) across four donors. Panel D: A box plot shows the average stepwise mutation count per tree edge for the same antibody contents and donors. The median values for each group are provided below the plots.
Tonsillar IgD-ME B cells give rise to different IgD-PC subpopulations via distinct developmental trajectories, and evidence a long developmental history. (A) Trajectory inference of IgD-ME B cells, IgD-GC B cells, and IgD-PCs. UMAP, trajectory inference (upper panel); dot plots showing differential gene expression by GC B cell, ME B cell, and PC subclusters (lower panel). (B) Heatmap visualizing the DE of a custom panel of genes involved in SHM from scRNA-seq analysis of tonsillar IgD-GC subcluster 1 and 2 vs. IgG-GC B cells. DEGs with |log2FC| >1 and adj. P <0.05 are marked with an asterisk. (C and D) Phylogenetic topology analysis of lineage trees’ normalized heights (C) and average stepwise mutation count (D). The numbers below plots in C and D represent the median value per group. Data show results from one experiment with multiple biological replicates. Statistical significance in C and D was assessed with pairwise Mann–Whitney tests. ****P < 0.0001.