Novel Plasma Cell developmental intermediates produce graded expression of IgM secretory transcripts
Differentiation into plasma cells (PC) enables secretion of ~10,000 antibody molecules per second. This extraordinary capacity requires the upregulation of PC transcriptional determinants that increase immunoglobulin mRNA synthesis, coordinate alternative 3'end processing of the heavy chain transcript from the distal to proximal poly-adenylation site (PAS), and remodel the secretory pathway. Based on the difficulty of prospectively identifying secretory cells, we developed a dual-fluorescent protein reporter mouse to study the post-transcriptional-level transition from membrane-anchored to secretory immunoglobulin M (µM PAS and µS PAS, respectively), in single cells. We observed (i) dynamic acquisition of graded µS PAS usage during PC differentiation (ii) IRF4 and Blimp-1 functioned hierarchically to increase µ abundance as well as µS PAS usage and (iii) graded µS populations did or did not express Blimp-1. Interestingly, the low and high µS and Blimp-1-expressing populations arose from distinct developmental intermediates that exhibited dissimilar endoplasmic reticulum features revealing unpredicted complexity of PC differentiation. The novel cell and µS PAS fate trajectories may have implications for discrete specializations of the secretory pathway. Overall design: To investigate the relationship between uS poly-adenylation site (PAS) usage and its relationship to Plasma cell (PC) differentiation, we created a mouse with an engineered allele of the u locus that reports on uS expression and bred that mouse to Blimp-1:YFP mice that marks PC. B cells from such mice were used to generate emergent PC using in vitro systems and we observed 4 populations of graded uS expression, uS INT and uS HI, that were Blimp-1:YFP negative (pre-PC) and uS INT and uS HI Blimp-1:YFP positive (PC). Comparative gene expression analysis of RNA-seq data was used to establish the transcriptomic states of the populations.