Grant: Discovery Grants Program - Individual

Egg activation is the process by which a mature egg is triggered to complete meiosis and at the same time undergo many developmental changes, including microtubule reorganization and the destabilization of specific maternal mRNAs, that prepare it for subsequent embryonic development. Cort, a meiosis-specific activator of the Anaphase Promoting Complex (APC/C), is involved in almost all aspects of egg activation. We are interested in understanding how APC/C is regulated in meiosis and and how it promotes the transition from egg to embryo. Aim 1. Regulation of the APC/C in Drosophila meiosis. We propose a model for stepwise activation of APC/C in meiosis. We found that the APC/C is active prior to anaphase, in the metaphase I arrested oocyte, where it specifically targets CycA. Surprisingly, this is independent of Cort or other known meiotic APC/C activators. We will determine how this occurs and its signficance. We found that CycB3-Cdk1 phosphorylation of APC/C subunit, APC3 is necessary for anaphase. We will determine how this event leads to APC/C activation. We will also determine how CycB3-Cdk1 itself is activated and how the CaN phosphatase relates to this and to activation of APC/C. Aim 2. APC/C targets in meiosis and egg activation. We found that chromosome segregation in Drosophila meiosis requires APC/C-mediated degradation of Securin to release the Separase protease. We also found that the known Separase targets, the kleisins, are not the targets in meiosis. We will identify the meiotic Separase target. How Cort orchestrates the many events of egg activation is unknown. We will identify the critical APC/C-Cort targets that mediate transcript destabilization and microtubule re-organization; and we will determine the significance of these processes for the transition from egg to embryo. Aim 3. Role of APC/C-Cort in sex determination. We found that misexpression of Cort outside of oogenesis transforms females to pseudomales, by targeting Tra, the key determinant of female fate downstream of Sxl. We found that loss of Cort in the female also also affects sexual fate, but in the next generation. Surprisingly, maternal Cort does not act on Tra. We will follow up on preliminary evidence that Cort acts on one of the X-chromosome signal elements (XSEs) that establish sexual identity in the early embryo by regulating Sxl. We will also study the evolution of Cort's role in sex determination, focusing on insect species where Tra, not Sxl is at the top of the sex determination pathway. This research program will train 2 PhD and 10 undergraduate students. All will receive high level training in a broad range of molecular, genetics, biochemistry, and imaging techniques. All students will be trained in scientific method, they will develop the ability to design and interpret experiments and they will learn to write and present their work. Students will be highly qualified for jobs in research, teaching and industry.