Role of G9a methyltransferase in mediating ATM and DNA-PK signaling after DNA repair

DNA damage induces an orchestrated sequence of local changes in histone and protein modifications that support the efficient recruitment of DNA repair factors to sites of DNA breaks. The regulation of these modifications at DNA break sites is imperative in maintaining genome integrity. Recent studies in our lab have identified a role for the G9a methyltransferase in regulating DNA repair and signaling. The overall aim of this thesis was to elucidate G9a’s activity in regulating proteins involved within the DNA Damage Response (DDR), and investigate the effects of its inhibition in the repair process. Here, we report that the loss of G9a significantly impairs DNA damage repair, and is important for maintenance of the genome after repair has concluded. Using genetic and biochemical assays, we found that in response to DNA damage, cells deficient of G9a genetic expression and activity exhibited increased ?H2AX signaling catalyzed by ATM and DNA-PKcs, but not the ATR kinase. Additionally, the autophosphorylation and downstream signaling activities of ATM and DNA-PKcs were significantly increased at sites of double strand breaks (DSBs) in G9a deficient cells, as demonstrated through increased accumulation of ATM substrates such as MDC1 and CHK2 at sites of DSBs. Biochemical data showed that G9a interacts and regulates ATM activity during the DNA damage response. G9a inhibition led to decreased ATM methylation. These data suggest that G9a activity regulates the extent of ATM activation induced by DNA breaks and is required for the efficient recruitment of downstream DNA repair factors. Together, our findings reveal a previously unidentified role of G9a in regulating function and activity during DDR. Our improved understanding of G9a molecular functions within DDR provide further validation for the use of G9a inhibitors in the clinic as cancer therapeutics.