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Allelic Exclusion of Antibody Gene Expression

Final Report Summary - KAPPA FLEX MOUSE (Allelic Exclusion of Antibody Gene Expression)

Immunoglobulin kappa (Igk) recombination creates a diverse and allelically excluded antibody light chain repertoire by joining a Vk gene segment with one of four Jk gene segments. This is controlled by two germline-transcript (GT) promoters located in the Vk-Jk region, but the mechanism that governs Jk choice is poorly understood. In the first part of this Marie-Curie Fellowship, the fellow showed in gene-targeted mice that the Jk proximal GT promoter contributes to targeting of primary rearrangements to Jk1 by preventing premature DNA breaks at Jk2. Therefore, the proximal GT promoter, which in contrast to the distal GT promoter is transcriptionally inactive during Igk recombination, controls the 5'- to-3'order of Jk rearrangements. Consequently, the removal of the proximal GT promoter allows use of more downstream Jk segments, thereby diminishing the potential for receptor editing in the Igk locus. The fellow's findings identify the first cis-element in the Igk locus that is critical for the internal order of antibody light chain rearrangements. This complements his collaborative studies on the role of CTCF-binding sites in controlling V
(d) J recombination of the antibody heavy chain locus.

In the second part of this Marie-Curie Fellowship, the fellow examined the importance of Igk allelic exclusion to B cell function and development. He successfully created a novel gene-targeted mouse model (kFlex) that was uniquely designed to address this question. kFlex can be induced through precisely timed Cre recombinase expression to produce about 12-13 % allelically included B cells with a diverse antibody repertoire. The fellow tested various hypotheses regarding the impact of allelically included B cells on the immune system. His results demonstrate that allelically included B cells were not counter-selected in vivo and that their presence leads to an increase of autoreactive antibodies in the serum. Even though, kFlex mice did not develop severe signs of lupus-like autoimmune disease, the observed increase in autoreactive antibodies mediated by allelically included B cells could be one of many contributing factors for this and other autoimmune diseases, which are notorious for their complex genetics and pathogenesis. Therefore, the molecular mechanisms that normally guarantee the allelic exclusion of antibody light chain genes could aid in maintaining central tolerance in the B cell branch of the human immune system.

The results of this fellowship will help biomedical researchers and medical doctors to better understand the regulation of antibody gene rearrangement, an important process that – if deregulated – can lead to autoimmune diseases and cancer. The fellow identified a key player in the ordered regulation of Igk gene recombination and demonstrated that ordered recombination of the Igk locus is critical for receptor editing, a process that prevents the generation of autoreactive antibodies. The fellow also found evidence supporting the idea that the allelic exclusion of Igk genes serves a similar function by limiting the secretion of autoreactive antibodies. These two novel mechanisms may represent safeguard strategies utilised by the murine and human immune system to preclude the initiation of autoimmune diseases.