RNA-seq analysis of quiescent human splenic B cell populations

Memory B cells represent one of the pillar of humoral protection and are found in the circulation and secondary lymphoid organs several decades after initial pathogen or vaccine encounter. The exact cellular and biological mechanisms allowing long-term survival of quiescent cells in such a fluctuating microenvironment remain poorly understood. scRNA-seq analysis of long-lived Vaccinia-specific and total IgG+ memory B cells had helped us identify two subsets of quiescent switched memory B cells in the spleen of adult donors, separated by the expression of CR2(CD21). To better understand the relationship between these two resting splenic memory B cell subsets, we sorted CD21hiCD20hi and CD21intCD20int IgG+ memory B cells from the live IgD-CD27+IgG+CD20+CD3-CD14-CD16- compartment of the spleen of four organ donors, collected between 2015 and 2019. For all donors, both IgG+ memory B cell subsets were sorted alongside naïve B cells (IgD+CD27-CD38-CD24-CD20+CD3-CD14-CD16-). Assay for Transposase-Accessible Chromatin with high-throughput sequencing (ATAC-seq) was performed on 50000 cells of each sorted population, according to published methods (Buenrostro, Nature Methods, 2013, PMID: 24097267) and sent for library sequencing by Integragen (Evry, France) (See linked experiment). RNA was extracted from remaining cells and sent for library preparation and sequencing by Integragen (Evry, France).