Exploring the Molecular Mechanisms by which AID Recombinase Interacts with DNA Secondary Structures involved in Cancer
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PublisherThe University of Arizona.
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EmbargoRelease after 8-Nov-2018
AbstractGenomic complexity in non-Hodgkin’s Diffuse Large B-cell Lymphoma (DLBCL) leads to a treatment failure in ~40% of patients. Activation-Induced Cytosine Deaminase (AID), one of the enzymes involved in generating antibody diversity via class switching recombination (CSR) and somatic hypermutation (SHM) of immunoglobulin (Ig) genes in activated B-cells is one mechanism for the introduction of genomic lesions. In previous studies, AID was shown to preferentially bind to super-enhancer (SE) regions within the genome, but 26% of AID targets were not within the SE regions. The mechanism by which AID interacts with SE elements and its off-target interactions still remains a mystery. Recent evidence suggests that AID may cause genomic lesions in DLBCL via interaction with oncogenes such as MYC and BCL2 resulting in mutations and translocations. Sequences within the MYC promoter contain the four-nucleotide AID target sequence (WRCY) and highly G-rich sequences known to form G-quadruplex DNA secondary structures. We hypothesize that key DNA secondary structures act as recruiting elements for aberrant AID activity at promoters and SEs of key genes involved in the development of DLBCL. Here, we first sought to determine whether known AID DNA targets have the potential to form G-quadruplex DNA secondary structures. The data collected from activated mouse B-cells showed 90% of the AID targets contained sequences that could potentially form G-quadruplexes and the data collected from the human Ramos cell line showed 100% of the sequences had the potential to form G-quadruplexes. To further study our hypothesis we used the techniques circular dichroism (CD) and the electrophoresis motility shift assay (EMSA) to explore the potential interaction between AID and the BCL2 and MYC G-quadruplexes. We observed no significant interactions between AID and these two G-quadruplexes, however further experimentation with different conditions and molecular techniques may show interaction. Additional studies will not only provide key insight into the genomic instability within DLBCL, but will also provide a potential mechanism by which AID is recruited to its DNA targets.
Degree ProgramGraduate College
Cellular and Molecular Medicine