To characterize nongenomic progestin receptors via knockouts in zebrafish

描述（由申请人提供）：类固醇的许多重要反应，如减数分裂恢复、精子激活、GnRH释放、抗细胞凋亡和性行为改变，通过非基因组机制在几秒或几分钟内迅速发生。这种途径不需要细胞核或基因转录的存在。甾体激素的非基因组信号通路尚不清楚，主要是由于围绕负责受体的身份存在争议。类固醇激素黄体酮的快速作用是这些非基因组类固醇作用的模型。近年来，一些候选蛋白，膜黄体酮受体（mPRs），核黄体酮受体和黄体酮受体膜组分被认为是脊椎动物，包括人类的非基因组黄体酮受体。然而，需要进一步的研究来确定它们的生理功能、信号传导和潜在的相互作用。目前还不清楚哪一个是

DESCRIPTION (provided by applicant): Many important responses of steroids, such as meiosis resumption, sperm activation, GnRH releasing, antiapoptosis, and sexual behavior changes, occur rapidly within seconds or minutes via a nongenomic mechanism. This pathway does not require the presence of a nucleus or gene transcription. The nongenomic signaling pathways of steroids are poorly understood, mainly due to the controversies surrounding the identity of the responsible receptors. The rapid action of the steroid hormone progestin is a model for these nongenomic steroid actions. Recently, several candidate proteins, membrane progestin receptors (mPRs), nuclear progestin receptor, and progestin receptor membrane components have been suggested to be the nongenomic progestin receptor in vertebrates, including humans. However, further studies are required to determine their physiological functions, their signaling, and potential interactions. Currently, it is not clear which one is the primary receptor or, alternatively, whether a set of complementary proteins are needed for nongenomic signaling. We will use a loss-of-functional approach to study the roles and nongenomic progestin signaling of these purported nongenomic progestin receptors in zebrafish knockout models. We will generate knockouts using TALENs (transcriptional activator-like effector nucleases), a simple, cost- effective and highly efficient knockout method. Thereafter, we will examine effects of knockouts in animal fertility, maturation competency, and changes in oocyte maturation, sperm activation and nongenomic progestin signaling in the germ cells. This proposed study will be the first to generate and characterize knockout models for purported nongenomic progestin receptors. Our long- term goals are to determine the functions and signaling of nongenomic steroid receptors in vertebrates and to develop specific antagonists and agonists for fertility control or treatments of infertility. Our short-term goal is to generate genetically modified models to solve the most hotly contested issues, i.e. if mPRs are the receptors responsible for the nongenomic progestin signaling, and whether a single or multiple classes of receptors is/are involved in the nongenomic progestin signaling.