Role of Target DNA sequences in mutation and DNA Break Generation

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DESCRIPTION (provided by applicant): In response to antigens, activated B cells undergo somatic hypermutation (SHM) and class switch recombination (CSR) to enhance antibody diversity. SHM and CSR each require the Activation Induced Deaminase (AID), which catalyses DNA-specific cytidine deaminase activity. AID targets the highly repetitive switch (S) regions of immunoglobulin heavy chain (Igh) loci to induce DNA double strand breaks (DSBs), which are intermediates in CSR. When targeting to variable (V) region exons of IgH and IgL loci, AID predominantly induces point mutations, termed SHM, resulting in the increased affinity of antibody for antigen. Intensive efforts have identified potential AID co-factors, which could bind S region DNA or RNA transcripts of Igh loci. This suggests a link between AID targeting and transcriptional pausing or RNA processing of S regions. In contrast to the relatively well studied targeting mechanisms for CSR, the mechanism of AID targeting to V region locus is much less well understood. Although AID has the ability to deaminate any transcribed substrate in vitro and could potentially access the genome widely to induce genomic instability in B cells, its physiological targets during SHM are almost exclusively restricted to V regions of Ig loci. However, due to the lack of robust experimental systems, it remains largely unknown how AID is specifically targeted to V regions, which is one of the most interesting unresolved questions in the field of AID regulation. Thus, we have established a unique experimental system, and propose to address these fundamental questions. The completion of our proposal will provide novel insights into the regulation of AID-mediated SHM in B cell immune responses and elucidate molecular mechanism of genomic instability in B cell lymphomas.