To initiate development, a newly fertilized egg must generate pronuclei that can combine to create a zygotic genome capable of initiating mitosis. The fertilized egg must also stimulate translation of previously quiescent, maternally deposited mRNAs. The goal of this proposal is to increase our understanding of this critical, but incompletely understood, developmental stage of """"""""egg activation"""""""" in the model organism Drosophila. The first specific aim focuses on the function of an important player in the fertilized egg: the """"""""Young Arrest"""""""" (Ya) gene. YA is a maternally encoded nuclear lamina protein that is essential for male and female pronuclei to initiate their first mitotic division. Our phenotypic analysis of Ya mutants suggests either that YA mediates changes in chromatin condensation needed for passage into S phase of the first mitosis, or alternatively that it replaces a meiotic protein that is inhibitory to subsequent mitotic cycles. The experiments described in this aim will test these hypotheses for YA function, and may further identify other molecules that interact with YA in fulfilling its roles. The results of these experiments will elucidate molecular changes necessary to transition frommeiosis to mitosis. We also expect these results to be of relevance to the mechanism of human diseases, such as Emery-Dreifuss Muscular Dystrophy, that are caused by mutations in nuclear lamina proteins.
The second aim will investigate the implications of our finding that the subcellular location of YA changes during development. YA is excluded from nuclei during oogenesis, but is able to enter nuclei after egg activation. During the same transition, there are also changes in YA?s phosphorylation state. We will test whether the change in YA?s ability to enter nuclei is a direct consequence of changes in its phosphorylation state, and whether MAP kinase mediates these events. We will also identify additional components of a macromolecular complex we have identified that retains YA in the cytoplasm prior to egg activation.
The third aim of the proposal is designed to achieve a broader view of the molecular events during egg activation. We will determine whether ovulation, which we have shown to trigger egg activation in Drosophila, causes a rise in calcium, parallel to findings in other systems. We will also test whether modulation of phosphorylation state, such as we see with YA, happens to many proteins during egg activation. Such modulation could mediate the rapid, concerted changes in egg physiology that occur at that time.
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