A temporal map of B cell diversification mechanisms in mice.

Naive B cells diversify via clonal expansion, immunoglobulin isotype switching, phenotypic variation and somatic hypermutation (SHM). Diversity in antigenic targets, functional classes and the production kinetics of antibodies affects immunity to malaria. Here we show that individual clones diversify over time during Plasmodium infection. During the first week, amid widespread bystander activation, isotype switching initiates soon after Myc upregulation and overlaps with clonal expansion, resulting in isotype variegation among clones. During the second week, expanded clones seeding germinal centers (GC) bifurcate into extrafollicular plasmablasts, exhibit isotype variegation and initiate SHM, indicating substantial intraclonal diversification. Over the following month, GC clones exhibit SHM at approximately four mutations per week. Antimalarial intervention does not impede SHM, instead exerting quantitative limits on GC size, plasma cell emergence, circulating antibody levels and protection against reinfection. Finally, contemporaneous B cell development relocates from bone marrow to spleen. Thus, multiple temporally overlapping mechanisms combine in vivo to diversify and safeguard humoral immune responses.