Epigenetic signatures mark early peripheral human B lineage bifurcation and differential transcriptional profiles in mature populations.

During human B cell maturation, immature transitional (T1) cells transit from bone marrow into the blood. At the subsequent immature T2 stage, a separation into IgMhi (T2Mhi) and IgMlo (T2Mlo) developmental trajectories has been proposed. Here, we isolate T1, T2Mhi and T2Mlo cells from human adult and cord blood for bulk and single-cell ATAC-seq, CUT&RUN, RNA-seq and CITE-seq to profile their transcriptomic and epigenetic differences. We identify accessible chromatin domains discriminating between T2Mhi and T2Mlo cells in peripheral B cell development, with signatures persisting during the differentiation of T2Mlo to naïve B cells. Similarly, memory and marginal zone B cells retain epigenetic hallmarks of their T2Mlo and T2Mhi precursors, coupled with transcriptional diversity. Imaging mass cytometry with RNAscope of spleen, appendix and tonsil further validates the spatial relationships of expressed genes. Our study thus provides insights into B lineage T2 bifurcation and describes epigenetic signatures of B cell developmental pathways.