Light chain editing in κ-deficient animals: A potential mechanism of B cell tolerance

Abstract

The genetic organization of the κ and λ light chain loci permits multiple, successive rearrangement attempts at each allele. Multiple rearrangements allow autoreactive B cells to escape clonal deletion by editing their surface receptors. Editing may also facilitate efficient B cell production by salvaging cells with nonproductive light chain (L chain) rearrangements. To study receptor editing of κ L chains, we have characterized B cells from mice hemizygous for the targeted inactivation of κ (JCkD/wt) which have an anti-DNA heavy chain transgene, 3H9. Hybridomas from JCkD/wt mice exhibited an increased frequency of rearrangements to downstream Jk segments (such as Jk5) compared with most surveys from normal mice, consistent with receptor editing by sequential κ locus rearrangements in JCkD/wt. We observed an even higher frequency of rearrangements to Jk5 in 3H9 JCkD/wt animals compared with nontransgenic JCkD/wt, consistent with editing of autoreactive κ in 3H9 JCkD/wt. We also recovered a large number of 3H9 JCkD/wt lines with Vk12/13-Jk5 rearrangements and could demonstrate by PCR and Southern analysis that up to three quarters of these lines underwent multiple κ rearrangements. To investigate editing at the λ locus, we used homozygous κ-deficient animals (JCkD/JCkD and 3H9 JCkD/JCkD). The frequencies of Vλ1 and Vλ2 rearrangements among splenic hybridomas in 3H9 JCkD/JCkD were reduced by 75% whereas VλX was increased 5-10-fold, compared with nontransgenic JCkD/JCkD animals. This indicates that Vλ1 and Vλ2 are negatively regulated in 3H9 JCkD/JCkD, consistent with earlier studies that showed that the 3H9 heavy chain, in combination with λ1 binds DNA. As successive λ rearrangements to VλX do not inactivate Vλ1, the consequence of λ editing in 3H9 JCkD/JCkD would be failed allelic exclusion at λ. However, analysis of 18 3H9 JCkD/JCkD hybridomas with Vλ1 and VλX DNA rearrangements revealed that most of these lines do not have productive λ1 rearrangements. In sum, both κ and λ loci undergo editing to recover from nonproductive rearrangement, but only κ locus editing appears to play a substantial role in rescuing autoreactive B cells from deletion.