Preventing Aneuploidy in Aging Oocytes: Investigating the effects and mechanisms of cohesion enrichment in Drosophila melanogaster.

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Abstract The overall goal for meiosis I is to successfully segregate homologous chromosomes into two diploid cells. Failure to do so results in aneuploidy, which is the leading cause of genetic disorders, such as Trisomy 21, and miscarriages due to spontaneous abortions. Incidences of chromosomal nondisjunction (NDJ) increase exponentially as an oocyte ages, a phenomenon termed the maternal-age effect. Age-related chromosomal NDJ is hypothesized to be caused by the inability of an oocyte to maintain a chiasma, the physical manifestation of a crossover (CO) between homologous chromosomes, until ovulation. Sister chromatid cohesion (SCC) is vital for maintenance of chiasmata; thus, SCC deterioration is thought to contribute to age- related NDJ. Studies have shown that an overall reduction of SCC causes age-related NDJ, but it is currently unknown whether the loss of SCC around the centromere has a greater impact on age-related NDJ than loss of SCC at the chromosome arms. Because arm SCC and centromere SCC have distinct roles in meiosis, understanding the role of each may provide insight into the mechanism underlying age-related NDJ. In Drosophila, loading of arm cohesion proteins is facilitated by Nipped-B, but the process governing centromere cohesin loading remains largely unknown. Recently, I characterized a separation-of-function allele, mcm5A7, and demonstrated that the mcm5A7 mutation does not affect the replication role of MCM5; instead, this mutation increases meiotic NDJ presumably by reducing loading of the cohesion protein SMC1 at the centromere. This result suggests that MCM5 is involved in recruiting or establishing SMC1 at the centromere in meiotic cells, a role for MCM5 that has not been previously described. As described in this proposal, I will test the importance of centromeric SCC versus arm SCC in age-related NDJ in Drosophila by utilizing mcm5A7 to disrupt centromeric SCC and a mutation in Nipped-B to disrupt arm SCC. I will then employ an oocyte aging protocol to measure the incidence of age-related NDJ in both mutants. Further, through proteomic studies and immunofluorescence, I propose to investigate the mechanism by which MCM5 functions to facilitate SMC1 at the centromere in meiotic cells. Overall, the lack of knowledge in the field regarding the importance of centromeric SCC and arm SCC in age-related NDJ, as well as the role of MCM5 in establishing centromeric cohesion, prevents progression of therapeutics and basic understanding in how to prevent age-related aneuploidy. The proposed research will help fill this fundamental gap and advance the development of prevention strategies.