Somatic Hyperrepair: A Novel Tumor Suppression Mechanism for Germinal Center B Cells.

All somatic cell types, including all B lineage cells, constitutively express a number of DNA repair proteins to maintain genomic stability and thwart tumorigenesis in the face of ongoing constitutive levels of DNA damage. Compromises in expression of these essential DNA repair factors have been shown to readily induce the development of various cancers including B cell lymphomas, suggesting expression levels are tightly regulated and limited in quantity. In germinal center (GC) B cells, however, additional DNA repair capacity is likely required to counterbalance the heightened level of mutagenic activity owing to the induced expression of activation-induced cytidine deaminase (AID) during somatic hypermutation (SHM). AID is a DNA editing enzyme which introduces somatic mutations into immunoglobulin (Ig) V regions at an estimated rate which is almost a million-fold higher than the spontaneous rate in somatic cells.

We hypothesize that to maintain the genomic wellness of GC B cells which are undergoing SHM, there is a need for induction of an accompanying robust DNA repair system. We further hypothesize that inefficient induction of these repair genes may predispose to malignant transformation.

Using tonsillar tissue sections and purified tonsillar B cell subpopulations, we compared expression levels of various DNA repair genes and related these levels to AID expression across the subsets using immunohistochemistry, real-time RT-PCR, and Western blot analysis. To characterize the nature of signals capable of inducing expression levels of AID and/or DNA repair proteins, peripheral blood B cells were activated in vitro using a panel of stimuli, including coculture with activated CD4 T cells. As a surrogate measure of mismatch repair (MMR) activity in the relative absence of T cell help, we quantitated the number of somatic hypermutations at A/T sites in the Ig heavy chain variable (IGHV) region genes in a collection of IGHV sequences obtained from normal B cells and HIV-related lymphoma cells.

Using immunohistochemistry, we observed that, similar to the expression of AID, DNA MMR genes are significantly induced in tonsillar GC B cells. These results were further validated using a more sensitive real-time RT-PCR assay and analysis by Western blotting. By expanding our DNA repair gene panel, we observed that proteins of homologous recombination, base excision repair and DNA single strand break signalling pathway are also similarly induced in GC B cells at RNA, protein, and functional levels compared to their expression in naïve and memory B cells. By contrast, expression of non-homologous end joining and DNA double strand break signalling molecules are unchanged. We have termed this selective induction of repair mechanisms in GC B cells as somatic hyperrepair (SHR). To identify pathways that lead to the activation of AID and SHR, we used an in vitro system and a variety of stimuli and we discovered that multiple B cell stimuli including CpG, CD40L, and anti-BCR could each independently induce the expression of AID while SHR induction strictly required the engagement of CD4+ T cells. This provocative observation suggests a novel role for CD4+ T cells in mitigating tumorigenesis of post-GC B cells through their ability to induce the SHR pathway in cells that have been induced to undergo SHM. To demonstrate the possible role of SHR in lymphomagenesis, we analysed the mutation pattern of IGHV genes from a panel of B cell lymphomas obtained from HIV infected (CD4+ T cell suppressed) patients. We found that HIV-related lymphoma cells displayed a significantly lower frequency of SHM at A/T positions relative to normal memory B cells, indicative of compromised MMR of their precursor cells during GC transit. Our findings resolve the lingering paradox that B cell malignancies are overwhelmingly prevalent under T cell suppression conditions such as HIV infection, post-organ transplant, and aging. Finally, our results also suggest for the first time that mounting efficient tumor suppression for some cells may depend on signals transmitted by neighboring cells and the specific microenvironment.

No relevant conflicts of interest to declare.