Characterizing the Sexually Dimorphic Role of Topoisomerase II During the Sister Chromatid Cohesion Release Pathway.

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PROJECT SUMMARY People in every socioeconomic class, both males and females, and every nationality and ethnicity are affected by infertility. Aneuploidy is the leading cause of infertility, as well as birth defects, and miscarriages. Aneuploidy occurs when there are errors in the specialized cell division of meiosis. Meiotic success is dependent on proper segregation of homologous chromosomes at meiosis I, and then subsequent sister chromatid segregation at meiosis II. There is still much that is unknown about the molecular mechanisms that contribute to infertility, and even less is known about the sexually dimorphic mechanisms during gametogenesis that can be contributing factors. The overall goal of the lab is to determine how these sexually dimorphic molecular mechanisms contribute to infertility. The goal of this proposal is to characterize the sex-specific roles of an enzyme crucial for relieving topological abnormalities in DNA during male and female meiosis. This enzyme, Topoisomerase II (Topo II), has been shown to have sex-specific phenotypes in both spermatogenesis and oogenesis in a variety of model organisms. Topo II also has roles in chromosome structure, condensation, and is a chromosome axis component in both mitosis and meiosis. Preliminary data indicates that Topo II also plays a role in the release of sister chromatid cohesion, an important step in ensuring the success of meiosis. The goals of this proposal will take a combined genetic, molecular biology, and biochemical approach to determine the sex-specific role of Topo II in meiosis. To achieve this, the applicant will be advised by a mentoring team that includes a sponsor with expertise in the fields of Meiosis and Genetics, as well as a co-sponsor who is an expert in Biochemistry, including protein expression, purification, and function. The collaboration between the candidate and her sponsors provides her with mentorship that is unique, interdisciplinary, and allows for the opportunity to develop a skillset that will be utilized throughout the entirety of her academic career, from post- doc to independent researcher. The resources, facilities, equipment, and faculty, available at the University of Delaware far exceed the needs of the proposal, ensuring a successful training environment to complete the aims proposed. Aim 1 uses single chromosome fluorescence labeling and nano-resolution microscopy to characterize the sexually dimorphic role of TOP-2 on sperm chromosome structure in late meiotic prophase l. I will also use a combination of genetic and confocal microscopy to determine the role of TOP-2 on oocyte chromosomes. Aim 2 will determine the differential protein interactions of TOP-2 in spermatogenesis and oogenesis using a biochemical approach in vivo and in vitro. These aims are innovative in the field of meiosis because the mechanisms known to govern the sex-specific differences in spermatogenesis and oogenesis are still largely unknown, exploring the role of Topo II will give us insight into one of the many functional and regulatory differences between male and female gametogenesis.