Successful embryonic development is dependent on the female gamete progressing correctly through its meiotic divisions, with the first division occurring during oocyte maturation and the second completing upon fertilization. Defects in these processes during meiosis I or II can compromise egg quality and competence to form a healthy embryo, thus negatively impacting female fertility. The fundamental biological question addressed in this proposal is how does the oocyte undergo the necessary asymmetric cell divisions during meiotic cytokinesis to create the egg and the polar bodies? The answer to this fundamental question is not as simple as might be assumed. From a cellular mechanics standpoint, it is remarkable that this cell division occurs at all, as fluid dynamics would predict that the polar body would simply collapse into the oocyte. Our work has identified a novel and previously unappreciated contributor to successful mammalian female meiosis - cortical tension in the oocyte, including demonstration that abnormal cortical tension is linked with aberrant spindle function and cytokinesis during completion of meiosis (Mol. Biol. Cell 21, 3182- 3192). We also identified some of the molecular basis of oocyte mechanics, as we demonstrated that oocyte tension is altered upon perturbation of actin, the actin-associated motor protein nonmuscle myosin-II, and the actin-to-membrane crosslinking proteins known as ERMs (for the family members ezrin, radixin, and moesin). To expand on this published work on the structural components involved in mammalian oocyte mechanics, this project seeks to analyze the functions of key proteins in cellular mechanics and cell shape regulation during progression of the mammalian oocyte through meiosis and cytokinesis. Specifically, we seek to characterize the system that regulates and fine-tunes actomyosin-mediated contractility in the oocyte cortical cytoskeleton.
Aim 1 is a brief series of studies that will be used to refine methodologies for this work.
Aim 2 is the substantial work of this proposal, testing the specific hypothesis that a signaling module composed of the small GTPase Rac1, 14-3-3, p21-activated kinase, and IQ-motif and GTPase-containing protein regulates the oocyte's structural cytoskeletal elements to modulate the oocyte's mechanical properties. We have preliminary data that provide the foundation for this hypothesis, and now seek to discover the contribution of these molecules and this signaling network to cortical tension in mouse oocytes. The overarching goal of this R03 project is to identify the molecules and signaling pathways that modulate mammalian oocyte mechanics, to build a conceptual framework that will be the foundation for future more detailed studies. This project will extend understanding of the female gamete in a completely new direction - bringing insights from cellular mechanics into our view of the oocyte-to-egg and egg-to-embryo transitions.
Successful embryo development is dependent on the female gamete progressing correctly through meiosis and its two cell divisions, one of which occurs with ovulation and then a second with fertilization. Defects in these processes can compromise egg quality and egg competence to form a healthy embryo, and have been associated with female infertility or subfertility. The research proposed here will discover regulatory signaling mechanisms that control how the oocyte undergoes cell division during meiosis.