BCR-signalling synergizes with TLR-signalling for induction of AID and immunoglobulin class-switching through the non-canonical NF-κB pathway

By diversifying antibody biological effector functions, class switch DNA recombination has a central role in the maturation of the antibody response. Here we show that BCR-signalling synergizes with Toll-like receptor (TLR) signalling to induce class switch DNA recombination. BCR-signalling activates the non-canonical NF-κB pathway and enhances the TLR-dependent canonical NF-κB pathway, thereby inducing activation-induced cytidine deaminase (AID), which is critical for class switch DNA recombination. Escherichia coli lipopolysaccharide (LPS) triggers dual TLR4/BCR-signalling and induces hallmarks of BCR-signalling, including CD79a phosphorylation and Ca2+ mobilization, and activates both the NF-κB pathways to induce AID and class switch DNA recombination in a PI(3)K p85α-dependent fashion. CD40-signalling activates the two NF-κB pathways to induce AID and class switch DNA recombination independent of BCR-signalling. Finally, dual BCR/TLR-engaging NP–lipopolysaccharide effectively elicits class-switched NP-specific IgG3 and IgG2b in mice. Thus, by integrating signals of the non-canonical and canonical NF-κB pathways, BCR and TLRs synergize to induce AID and T-cell-independent class switch DNA recombination

14-3-3 adaptor proteins recruit AID to 5′-AGCT-3′–rich switch regions for class switch recombination

Class switch DNA recombination (CSR) is the mechanism that diversifies the biological effector functions of antibodies. Activation-induced cytidine deaminase (AID), a key CSR player, targets IgH switch (S) regions, which contain 5′-AGCT-3′ repeats in their core. How AID is recruited to S regions remains unclear. Here we show that 14-3-3 adaptor proteins play an important role in CSR. 14-3-3 proteins specifically bind 5′-AGCT-3′ repeats, are upregulated in B cells undergoing CSR and are recruited together with AID to the S regions involved in CSR events (Sμ→Sγ1, Sμ→Sγ3 or Sμ→Sα). Moreover, blocking 14-3-3 by difopein, deficiency in 14-3-3γ or expression of a dominant negative 14-3-3σ mutant impaired recruitment of AID to S regions and decreased CSR. Finally, 14-3-3 proteins interact directly with AID and enhance AID-mediated in vitro DNA deamination, further emphasizing the important role of these adaptors in CSR