Haploinsufficiency of Activation-Induced Deaminase for Antibody Diversification and Chromosome Translocations both In Vitro and In Vivo

Isora V Sernández,Yair Dorsett,Almudena R Ramiro,Virginia G De Yébenes,Marta Feldmesser

doi:10.1371/journal.pone.0003927

Abstract

The humoral immune response critically relies on the secondary diversification of antibodies. This diversification takes places through somatic remodelling of the antibody genes by two molecular mechanisms, Class Switch Recombination (CSR) and Somatic Hypermutation (SHM). The enzyme Activation Induced Cytidine Deaminase (AID) initiates both SHM and CSR by deaminating cytosine residues on the DNA of immunoglobulin genes. While crucial for immunity, AID-catalysed deamination is also the triggering event for the generation of lymphomagenic chromosome translocations. To address whether restricting the levels of AID expression in vivo contributes to the regulation of its function, we analysed mice harbouring a single copy of the AID gene (AID+/−). AID+/− mice express roughly 50% of normal AID levels, and display a mild hyperplasia, reminiscent of AID deficient mice and humans. Moreover, we found that AID+/− cells have an impaired competence for CSR and SHM, which indicates that AID gene dose is limiting for its physiologic function. We next evaluated the impact of AID reduction in AID+/− mice on the generation of chromosome translocations. Our results show that the frequency of AID-promoted c-myc/IgH translocations is reduced in AID+/− mice, both in vivo and in vitro. Therefore, AID is haploinsufficient for antibody diversification and chromosome translocations. These findings suggest that limiting the physiologic levels of AID expression can be a regulatory mechanism that ensures an optimal balance between immune proficiency and genome integrity.

Highlights

B cells are responsible for generating a repertoire of antibodies of virtually unlimited diversity in order to confront the antigenic universe
Mature B cells were isolated from spleens from Activation Induced Cytidine Deaminase (AID)+/+ and AID+/2 mice, stimulated in vitro in the presence of LPS and IL4 to promote AID transcriptional activation, and AID expression was quantified after 3 days of culture by real-time RT-PCR
We found that AID mRNA levels are reduced roughly to 50% in AID+/2 as compared to wild type AID+/+ B cells (Figure 1a)

Summary

Introduction

B cells are responsible for generating a repertoire of antibodies of virtually unlimited diversity in order to confront the antigenic universe. The first one is antigen- independent and takes place during B cell generation in the bone marrow through a site-specific recombination named V(D)J recombination, which gives rise to B cells expressing a primary repertoire of Iow affinity IgM antibodies (reviewed in [1]). Upon antigen encounter B cells have as yet another chance to further diversify their antibody repertoire in germinal centers by two independent molecular mechanisms called somatic hypermutation (SHM) and class switch recombination (CSR). B cells where SHM gives rise to antibodies with higher affinity for their cognate antigen are positively selected, a process referred to as affinity maturation (reviewed in [2] and [3]). CSR takes place between highly repetitive sequences that precede the Cm, Cc, Ce and Ca genes, called switch regions, through the generation of double strand breaks (DSBs), ligation, and concomitant excision of the intervening sequence from the locus (reviewed in [4,5])

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Conclusion