Oral Presentation The Joint Annual Scientific Meetings of the Endocrine Society of Australia and the Society for Reproductive Biology 2017

Double strand break DNA Repair occurs via non homologues end joining in mouse MII oocytes (#15)

Jacinta H Martin 1 2 , Elizabeth G Bromfield 1 2 , R. John Aitken 1 2 , Brett Nixon 1 2
  1. Priority Research Centre in Reproductive Science (PRC), University of Newcastle, Callaghan, Newcastle, NSW, Australia
  2. University of Newcastle, CALLAGHAN, NSW, Australia

DNA damage detection and repair are well characterized facets of somatic cell biology. However, the distinct biology of the oocyte means that the accepted paradigm for DNA repair and protection does not apply to this highly specialised cell. Instead protection of the genomic integrity of the oocyte must depend on stores of pre-synthesised proteins and/or mRNA that are accumulated during oogenesis. This study aimed to determine whether these stored proteins have the capacity to contribute to DNA damage detection and repair in the MII oocyte. For this purpose, DNA double strand breaks (DSB) were elicited using etoposide a chemotherapeutic agent that inhibits the action of topoisomerase II. Using this strategy, we confirmed that etoposide lead to an increase in DSBs (P < 0.003), and a consequential increase in the incidence of metaphase plate abnormalities (P< 0.0002). Despite this, treated MII oocytes retained their ability to participate in in vitro fertilisation though only 3% of these embryos displayed the developmental competence to progress beyond the 2-cell stage (P <0.002). To determine if the MII oocyte possesses additional protective mechanisms to ameliorate damage prior to fertilisation, we analysed the resolution of DNA DSB over a 6-hour time-course. This study revealed that a functional DSB DNA repair response is mounted in the MII oocyte (P <0.008). Furthermore, pharmacological inhibition of DNA-PKcs and DNA ligase IV established that this response is mediated by the canonical Non-Homologous End Joining (NHEJ) pathway. In keeping with this hypothesis, we have confirmed that several key repair proteins (ATM, XRCC5, DNA-PKcs) critical for the fidelity of the NHEJ pathway reside in the MII oocyte. Taken together, these data provide a unique insight into the innate biochemical defences within the oocyte and identifies a potential target pathway for manipulation in future studies to enhance the protection of the maternal genome.