Molecular mechanisms of activation induced deaminase specificity

Resumo

B lymphocytes are key effectors of the humoral immune response through the secretion of antibodies. The most distinctive event in mature B lymphocytes biology is the secondary diversification of their immunoglobulin genes during the germinal center (GC) reaction, which is fundamental to generate a repertoire of antibodies with virtually unlimited specificities. Activation Induced Deaminase (AID) initiates secondary antibody diversification in GC B cells through the deamination of cytosines on immunoglobulin genes. Remarkably, AID can also target other regions in the genome, triggering mutations or chromosome translocations, with major implications for oncogenic transformation. However, understanding the specificity of AID has proved extremely challenging, mostly because of the difficulty to detect AID-induced mutations, which occur at very low frequencies. In this work we have developed a novel capture-based approach to explore AID mutagenesis in a representation of the B cell genome. We have sequenced at very high depth 1588 genomic regions from GC B cells and identified 275 genes targeted by AID, including 30 of the previously known 35 AID targets. We have also identified the most highly mutated hotspot for AID activity described to date. Further, integrative analysis of the molecular features of mutated genes coupled to machine learning has produced a powerful predictive tool for AID targets, which has been experimentally validated. We have also found that Ease Excision Repair and Mismatch Repair pathways back-up each