| Somatic hypermutation (SHM) diversifies immunoglobulin loci prodominately in the context of an antigen dependent germinal center reaction. SHM is a gene-specific reaction, primarily mutating the V regions of immunoglobulin (Ig) genes. The mechanism of this gene-specific targeting is not understood, although Ig enhancers and chromatin modifications have been suggested to be involved. In contrast to previous studies, I find that the addition of Ig kappa (Igkappa) enhancers to a non-Ig substrate does not reproducibly target SHM to the substrate, suggesting that Igkappa enhancers are not sufficient to recruit the hypermutation machinery in the context of these constructs. Furthermore, we find that the endogenous 3' Igkappa enhancer is not required for targeting SHM to the Igkappa locus, but instead promoters a level of Igkappa transcription required for robust SHM. A number of modified histones, including acetylated H3 and H4 and phosphorylated H2AX (gammaH2AX), are associated with V(D)J recombination and class switch recombination (CSR). In contrast, little is known concerning the chromatin modifications associated with SHM in vivo. Data presented here shows that several modifications---including histone acetylation and H3-lysine 4 methylation---fail to demarcate an actively hypermutating Ig locus or to correlate spatially with SHM within Ig loci. Furthermore, no obvious association between SHM and gammaH2AX could be detected. Instead, we find that the phosphorylated form of histone H2B (H2BSer14P) correlates tightly with SHM and CSR. Phosphorylation of H2B within Ig variable and switch regions requires AID and may be mediated by the histone kinase Mst1. These findings indicate that SHM and CSR trigger distinct DNA damage responses and identify a novel histone modification signature for SHM consisting of H2BSer14P in the absence of gammaH2AX. |
