AID-Targeting and Hypermutation of Non-Immunoglobulin Genes Does Not Correlate with Proximity to Immunoglobulin Genes in Germinal Center B Cells

Abstract

Upon activation, B cells divide, form a germinal center, and express the activation induced deaminase (AID), an enzyme that triggers somatic hypermutation of the variable regions of immunoglobulin (Ig) loci. Recent evidence indicates that at least 25% of expressed genes in germinal center B cells are mutated or deaminated by AID. One of the most deaminated genes, c-Myc , frequently appears as a translocation partner with the Ig heavy chain gene ( Igh ) in mouse plasmacytomas and human Burkitt's lymphomas. This indicates that the two genes or their double-strand break ends come into close proximity at a biologically relevant frequency. However, the proximity of c-Myc and Igh has never been measured in germinal center B cells, where many such translocations are thought to occur. We hypothesized that in germinal center B cells, not only is c-Myc near Igh , but other mutating non-Ig genes are deaminated by AID because they are near Ig genes, the primary targets of AID. We tested this “collateral damage” model using 3D-fluorescence in situ hybridization (3D-FISH) to measure the distance from non-Ig genes to Ig genes in germinal center B cells. We also made mice transgenic for human MYC and measured expression and mutation of the transgenes. We found that there is no correlation between proximity to Ig genes and levels of AID targeting or gene mutation, and that c-Myc was not closer to Igh than were other non-Ig genes. In addition, the human MYC transgenes did not accumulate mutations and were not deaminated by AID. We conclude that proximity to Ig loci is unlikely to be a major determinant of AID targeting or mutation of non-Ig genes, and that the MYC transgenes are either missing important regulatory elements that allow mutation or are unable to mutate because their new nuclear position is not conducive to AID deamination.

Absence of the Uracil DNA Glycosylase of Murine Gammaherpesvirus 68 Impairs Replication and Delays the Establishment of Latency In Vivo

“…hi B cells with high levels of host UNG2 (40). Our data clearly demonstrate an important role for the vUNG in virus replication in the lung that is largely devoid of host UNG, and yet the mechanisms by which the host and viral UNGs contribute to gammaherpesvirus replication are not well characterized.…”

Regulation of AID, the B-cell genome mutator

“…The myc oncogene was not found to be any closer to IgH than other known AID mutated genes in these studies. Based on these observations, the investigators concluded that the proximity of the Ig locus to other genes is not a major determinant in causing AID mutations at non-Ig genes (Gramlich et al 2012).…”

The Origin of B Cell Recurrent Chromosomal Translocations: Proximity versus DNA Damage

“…Unlike AID-independent events, the frequency of translocations at hotspots does not correlate with target loci proximity but with the amount of damage inflicted by AID and measured by RPA or Rad51 accumulation during DNA-end resection by homologous recombination (Figure 5A in (Hakim et al, 2012) and Figure 3C in (Yamane et al, 2013), Spearman’s ρ > 0.6). These ideas were independently confirmed in germinal center cells by 3D FISH (Gramlich et al, 2012). In contrast to these studies, Rocha et al reanalyzed our published TC-Seq only from AID +/+ samples and concluded that the frequency of translocations correlates with nuclear proximity (Rocha et al, 2012).…”