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Regulation of the inositol 1,4,5-trisphosphate receptor degradation during fertilization and its impact on [calcium(2+)](i) signaling in mammalian eggs
Fertilization initiates a series of intracellular calcium [Ca 2+]i oscillations in the oocyte that promote its exit from and completion of meiosis. In mouse oocytes, the [Ca2+] i oscillations arrest at the pronuclear stage and this is accompanied by a down-regulation in the inositol 1,4,5-trisphosphate receptor type 1 protein (IP3R-1). The goal of this dissertation was first, to investigate the signaling pathway responsible for IP3R-1 degradation during fertilization. Eggs were either incubated in or injected with several compounds that induce egg activation, and the IP3R-1 protein content evaluated using Western blot analysis. Exposure to ethanol or ionomycin, which induce a single [Ca 2+]i rise, failed to induce down-regulation of the IP3R-1. However, injection of porcine sperm factor (pSF), which presumably stimulates IP3 production, or adenophostin A, an IP3R-1 agonist, both induced degradation of the receptor. Exposure to thimerosal, that modulates the IP3R without stimulating IP3 production, also initiated down-regulation. [Ca 2+]i oscillations induced by SrCl2, which does not trigger IP3 production, failed to evoke down-regulation. Degradation of the IP3R-1 appears to be mediated by the proteasome pathway because it was inhibited by lactacystin, a specific proteasome inhibitor. It was therefore propose that persistent stimulation of the phosphoinositide pathway by sperm during fertilization leads to down-regulation of the IP3R-1. The second goal of this research was to investigate the factor(s) involved in the arrest of [Ca2+]i oscillations seen at PN formation. This was achieved by generating parthenotes with similar IP3R-1 numbers and similar kinase activity at three different stages of the cell cycle, metaphase II (MII), pronuclear (PN), and the post pronuclear (PPN) stages. These models allowed us to study the effect of different parameters on PN [Ca2+]i arrest: (1) Cell cycle, (2) IP3R-1 number and sensitivity, and (3) content of intracellular Ca2+ stores. Results from these experiments demonstrate that the cell cycle determines the ability of the egg to exhibit [Ca2+] i oscillations by controlling the decrease in sensitivity of the IP3R-1 at the PN stage. This desensitization appears to be due to the inability of the IP3R-1 to undergo a conformational change and this phenomenon seems to be regulated by presently unknown factors.
Anatomy & physiology|Animals|Animal diseases
Jellerette, Teru Jacqueline, "Regulation of the inositol 1,4,5-trisphosphate receptor degradation during fertilization and its impact on [calcium(2+)](i) signaling in mammalian eggs" (2002). Doctoral Dissertations Available from Proquest. AAI3068567.