Pathways and consequences of antibody heavy chain rearrangement in B cells with a site-directed heavy chain transgene.

摘要

Antibody heavy chain gene rearrangement occurs in some B cells that harbor a productive heavy chain gene. The cause of these additional rearrangements is unknown, although it has been proposed that they occur in order to correct or edit the specificity of an autoreactive B cell receptor. Here, the genetic pathways, developmental timing and functional consequences of ongoing heavy chain rearrangements have been analyzed in a mouse model that harbors a site-directed anti-dsDNA heavy chain called 56R.;H chain rearrangements in B cells from 56R mice were characterized by sequencing DNA. These sequences revealed a bias for rearrangement of gene segments at the 3' end of the immunoglobulin variable locus. The sequences of H chain rearrangements in 56R B cells harbored N-nucleotide additions, characteristic of rearrangements occurring at the pro-B cell stage of development, when H chain rearrangement normally occurs, rather than when light chain receptor editing occurs. These data suggest that H chain rearrangement in 56R B cells occurs without regard for contribution of the H chain transgene to the specificity of the BCR.;The consequences of H chain rearrangement in 56R B cells were also analyzed. 56R B cells that had rearranged the endogenous H chain allele (IgM b) exhibited several characteristics that are likely to be advantageous compared to 56R-expressing B cells. Hybridomas created from IgMb+ B cells from spleen of 56R mice were less likely to secrete anti-dsDNA antibodies than 56R-expressing hybridomas. 56R negative hybridomas also exhibited a more diverse repertoire of light chains compared to those expressing 56R. Intriguingly, B cells that successfully replaced 56R via rearrangement of a VH gene segment on that allele show features that are suggestive of counter-selection, including long CDR3 length and a lower in-frame frequency, compared to rearrangements on the endogenous allele and in non-transgenic mice.;The work presented herein suggests that H chain rearrangement does not occur in response to BCR reactivity, but rather that continued H chain rearrangement can have a tremendous impact in the context of an autoimmune repertoire. The 56R model can be further used to determine the frequency of H chain rearrangement in B cells with a functional H chain gene, and to study the mechanisms of regulating autoreactive B cells.