FUNCTION AND REGULATION OF MATERNAL PROTEINS THAT ARE PHOSPHOREGULATED DURING EGG ACTIVATION
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Egg activation is the essential transition through which a mature oocyte becomes a developmentally competent egg. During this transition, the oocyte completes meiosis and remodels its transcriptome and proteome through post-transcriptional and post-translational regulations to prepare for embryogenesis. Phosphoregulation is a particularly important type of post-translational modification at the time of egg activation. In addition to the prevalence of protein phosphorylation state changes during this transition, conserved phosphoregulators like calcineurin and CaMKII are essential for egg activation in different species. Since the phosphorylation state of a protein is tightly associated with its activities, the proteins that are subject to phosphoregulation during egg activation are likely involved in the transition from oocyte to embryo. In this dissertation, I present my studies on the function and regulation of proteins that go through phosphorylation state changes during egg activation in Drosophila. With germline-specific RNAi, I tested the function of 189 such proteins in female fertility, and identified 53 proteins whose germline depletion led to defective oogenesis, as well as 51 proteins whose germline depletion led to significant impairment or abolishment of the eggs' ability to hatch. By carefully examining the knockdown phenotypes, I identified a set of regulators that are essential for processes in both early oogenesis and early embryogenesis, and revealed 15 proteins with new roles in egg activation and embryogenesis. To explore the mechanisms that mediate the phosphorylation state changes of proteins during egg activation, I investigated the activity of phosphoregulator calcineurin in this transition using proteomic and phosphoproteomic analysis. I quantified the phosphorylation state changes and protein abundance changes that occur during egg activation, and examined how these changes are affected when calcineurin function is perturbed in female germ cells. I showed that calcineurin is involved in the regulation of hundreds of phosphosites upon egg activation, and is also required for the abundance changes of numerous proteins during this transition. My results indicate that eggs with perturbed calcineurin activity fail to exit metaphase I, likely due to unsuccessful activation of APC/C. I also showed that calcineurin activity is required for Pan Gu kinase activation, and affects the phosphoregulation of several other regulators of protein translation upon egg activation. In summary, my studies showed the functional importance of proteins that are phosphoregulated upon egg activation, and shed lights on the mechanism of phosphoregulation during this transition.
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Schimenti, John C.