TLR7- and TLR9-responsive human B cells share phenotypic and genetic characteristics

B cells activated by nucleic acid-sensing TLR7 and TLR9 proliferate and secrete immune globulins. Memory B cells are presumably more responsive due to higher TLR expression levels, but selectivity and differential outcomes remain largely unknown. In this study, peripheral blood human B cells were stimulated by TLR7 or TLR9 ligands, with or without IFN-α, and compared with activators CD40L plus IL-21, to identify differentially responsive cell populations, defined phenotypically and by BCR characteristics. Whereas all activators induced differentiation and Ab secretion, TLR stimulation expanded IgM(+) memory and plasma cell lineage committed populations, and favored secretion of IgM, unlike CD40L/IL-21, which drove IgM and IgG more evenly. Patterns of proliferation similarly differed, with CD40L/IL-21 inducing proliferation of most memory and naive B cells, in contrast with TLRs that induced robust proliferation in a subset of these cells. On deep sequencing of the IgH locus, TLR-responsive B cells shared patterns of IgHV and IgHJ usage, clustering apart from CD40L/IL-21 and control conditions. TLR activators, but not CD40L/IL-21, similarly promoted increased sharing of CDR3 sequences. TLR-responsive B cells were characterized by more somatic hypermutation, shorter CDR3 segments, and less negative charges. TLR activation also induced long positively charged CDR3 segments, suggestive of autoreactive Abs. Testing this, we found culture supernatants from TLR-stimulated B cells to bind HEp-2 cells, whereas those from CD40L/IL-21-stimulated cells did not. Human B cells possess selective sensitivity to TLR stimulation, with distinctive phenotypic and genetic signatures.