Signal Transduction at Fertilization

受精时的信号转导

基本信息

批准号：
10381516
负责人：
LAURINDA A. JAFFE
金额：
$ 49.2万
依托单位：
UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
依托单位国家：
美国
项目类别：
财政年份：
1981
资助国家：
美国
起止时间：
1981-03-01 至 2024-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10381516
关键词：
Address Adenylate Cyclase Affinity Agonist Antral Biochemistry Biological Assay C-Type Natriuretic Peptide CDC2 gene Calcium Cancer Patient Catalytic Domain Cell Cycle Cellular biology Clinical Complex Confocal Microscopy Cyclic AMP Cyclic AMP-Dependent Protein Kinases Cyclic GMP Data Development Diffuse Enzyme Inhibitor Drugs Enzymes Epidermal Growth Factor Receptor Event Fertilization Fertilization in Vitro Future GTP-Binding Proteins Gap Junctions Glutamates Guanylate Cyclase Hour Human Immunoprecipitation In Vitro Knowledge LH Receptors Luteinizing Hormone Maintenance Measurement Measures Mediating Meiosis Methods Modification Molecular Mus Natriuretic Peptides Oocytes Ovarian Follicle Ovarian tissue cryopreservation Ovary Patients Peptide Receptor Permeability Phosphoric Monoester Hydrolases Phosphorylation Phosphorylation Site Phosphotransferases Physiological Processes Pituitary Hormones Polycystic Ovary Syndrome Process Prophase Protein Dephosphorylation Proteins Rattus Receptor Activation Receptor Signaling Regulation Reproductive Medicine Research Signal Transduction Signaling Protein Site Testing Tissues Transgenic Mice Western Blotting Woman Work base cGMP production clinical development egg enzyme activity experimental study granulosa cell hormonal signals in vitro activity infertility treatment inhibitor interest mutant oocyte maturation optical sensor phosphoric diester hydrolase public health relevance response

项目摘要

DESCRIPTION (provided by applicant): The objective of this project is to understand the signaling mechanisms by which eggs are activated to begin development. The current renewal application, for years 30-34, addresses the question of how luteinizing hormone (LH) from the pituitary acts on the ovary to cause oocytes to progress to the stage at which they can be fertilized. Mammalian oocytes are stored in the ovary, arrested at meiotic prophase, for decades in women. Then in response to luteinizing hormone signaling in the surrounding follicle, meiosis resumes, and the mature egg is ovulated. Studies in mice have shown that meiotic arrest in antral follicles is maintained by cyclic GMP that is produced by the granulosa cells and diffuses into the oocyte through gap junctions; LH signaling decreases gap junction permeability and cGMP production by the granulosa cells, thus lowering cGMP in the oocyte. The cGMP decrease leads to the release of inhibition of the meiotic cell cycle. Despite knowledge of this cascade, many important questions still remain. This proposal focuses on the key event of the reduction in cGMP by investigating how LH signaling reduces the guanylyl cyclase activity of natriuretic peptide receptor 2 (NPR2) (aims 1 and 2), and how LH signaling lowers cGMP through activity of cGMP phosphodiesterases (aim 3). Aim 1 will test whether LH signaling decreases NPR2 guanylyl cyclase activity by dephosphorylating NPR2 regulatory sites. Aim 2 will investigate how LH activation of G-proteins and the EGF receptor initiates the decrease in the guanylyl cyclase activity of NPR2. Aim 3 will investigate which cGMP phosphodiesterases function to lower cGMP in the follicle, and whether they are stimulated by LH. The methods to be used include isolation and culture of ovarian follicles and granulosa cells from mice and rats, confocal microscopy, immunoprecipitation, Western blotting, use of transgenic mice and specific enzyme inhibitors, use of optical sensors to measure cGMP and calcium in live ovarian follicles, and assays of enzyme activities in vitro. The conclusions reached from these studies will also be applicable to understanding the regulation of meiosis in women. In vitro oocyte maturation, in which LH receptor stimulation is performed in vitro, is an emerging component of methods for human in vitro fertilization, particularly for patients with polycystic ovary syndrome and in combination with cryopreservation of ovarian tissue from cancer patients. The proposed studies will contribute to understanding of the biochemistry and cell biology underlying clinical advances in in vitro maturation.

描述（由申请人提供）：该项目的目的是了解激活卵以开始发育的信号传导机制。目前的续订应用程序为30 - 34年，它解决了垂体对卵巢的黄体激素（LH）如何导致卵母细胞发展到可以受精的阶段的问题。哺乳动物的卵母细胞存储在卵巢中，在减数分裂的预言中被捕数十年。然后，为了响应于周围卵泡中的激素信号传导，减数分裂恢复并排卵成熟的卵。在小鼠中的研究表明，由颗粒细胞产生的环状GMP维持了肛门卵泡中的减数分裂阻滞，并通过间隙连接扩散到卵母细胞中。 LH信号传导可降低颗粒细胞的间隙连接性渗透性和CGMP的产生，从而降低卵母细胞中的CGMP。 CGMP减少导致减数分裂细胞周期的抑制作用。尽管知道这一级联，但仍然存在许多重要的问题。该提案通过研究LH信号传导如何降低Natriuretic肽受体2（NPR2）（AIMS 1和2）的关键事件（目标1和2），以及LH信号传导如何通过CGMP降低CGMP磷酸二酯酶的活性（AIM AIM 3）。 AIM 1将测试LH信号是否通过去磷酸化NPR2调节位点来降低NPR2 Guanylyl环化酶活性。 AIM 2将研究G蛋白的LH激活和EGF受体如何启动NPR2的Guanylyl环化酶活性的降低。 AIM 3将研究哪种CGMP磷酸二酯酶在卵泡中的CGMP降低，以及它们是否受到LH的刺激。要使用的方法包括来自小鼠和大鼠的卵巢卵泡和颗粒细胞的隔离和培养，共焦显微镜，免疫沉淀，蛋白质印迹，使用转基因小鼠以及特定的酶抑制剂，使用光学传感器在活卵巢卵泡中测量CGMP和钙化的eNzyme和sissyzeme noceme of vitro of vitro。这些研究得出的结论也将适用于了解女性减数分裂的调节。体外卵母细胞在体外进行LH受体刺激是人类体外受精方法的新兴成分，特别是对于多囊卵巢综合征的患者，以及与癌症患者的卵巢组织的冷冻保存结合在一起。拟议的研究将有助于理解体外成熟的临床进步的生物化学和细胞生物学。