Mechanisms of Sex Determination in Zebrafish

描述（由申请人提供）人类正常性腺发育在子宫内可能受到未知遗传和环境因素的不利影响，导致睾丸发育不良综合征和多囊卵巢综合征。在哺乳动物中，Y染色体基因SRY启动了一个不完全理解的遗传网络，指导性腺成为睾丸，而竞争基因指导卵巢发育。非哺乳类脊椎动物使用类似的下游网络，但初级性别决定（SD）基因不同。此外，在某些物种中，环境因素如温度或营养偏差SD。我们的广泛目标是了解遗传因素如何与环境相互作用，以打破SD平衡。研究的重点是斑马鱼，这是一种遗传学上易于驯服的脊椎动物，环境或遗传因素可以将性腺推向卵巢或睾丸的命运。斑马鱼的性腺是由双能器官发育而来的，其中一些原始生殖细胞（PGCs）在所有的幼鱼中都成为卵母细胞。有些个体卵母细胞死亡，性腺变成睾丸;有些个体卵母细胞存活，性腺变成卵巢。一个关键的问题是导致卵母细胞在一些青少年中死亡而在另一些青少年中存活的遗传级联的性质，以及环境因素如何改变这种级联。结果提示了这样的假设，即天然SD基因座（我们在天然菌株中发现，但从驯化的实验室菌株中消失）与环境因素（可能是可变的营养和生长速率）相互作用，以改变幼卵母细胞的存活率，这反过来又改变了作用于性腺索马的卵母细胞源信号的强度，以维持雌激素的产生，从而保留或不保留卵母细胞和性腺的命运。目的1是了解斑马鱼SD基因座的分子遗传基础。目的2是理解SD期间的转录组级联，使用：a）针对遗传性别进行基因分型的天然菌株，B）缺乏天然SD基因的环境敏感实验室菌株，和c）缺乏PGC的鱼，以鉴定PGC衍生因子及其下游靶标。目的3是了解性腺索马/生殖细胞相互信号传导的机制，通过了解相关的TGF β信号转导因子Gdf 9，Amh和Gsdf（性腺索马衍生因子）的作用，并发现女性特异性信号Wnt 4和Foxl 2如何在性腺发育中发挥作用，通过使用诱导突变，其中许多我们已经在手。拟议工作的创新包括识别和独特的使用斑马鱼拥有和基因型的本地SD位点;我们普遍适用的RAD性别方法，在不同的人口确定性别位点;独特的突变体和转基因资源，我们提供;和我们专注于性腺索马/生殖细胞的相互信号在斑马鱼性别决定。这项工作的成果和影响是更充分地了解脊椎动物性别平衡机制的分子遗传性质的潜力，这将增加我们对发达国家人类生殖疾病增加的潜在机制的认识。

DESCRIPTION (provided by applicant) Normal human gonad development can be adversely affected in utero by unknown genetic and environmental factors leading to testis dysgeneration syndrome and polycystic ovarian syndrome. In mammals, the Y-chromosome gene SRY initiates an incompletely understood genetic network that directs the gonad to become a testis, while competing genes direct ovary development. Non-mammalian vertebrates use similar down-stream networks, but primary sex determination (SD) genes differ. Additionally, in some species, environmental factors such as temperature or nutrition bias SD. Our broad objective is to learn how genetic factors interact with the environment to tip SD balance. Work focuses on zebrafish, a genetically tractable vertebrate in which environmental or genetic factors can nudge gonads to an ovary or testis fate. Zebrafish gonads develop from bipotential organs in which a few primordial germ cells (PGCs) become oocytes in all juveniles. In some individuals oocytes die and gonads become testes; in others oocytes survive and gonads become ovaries. A key question is the nature of the genetic cascade that causes oocytes to die in some juveniles and survive in others, and how environmental factors alter this cascade. Results suggest the hypothesis that the native SD locus (which we discovered in native strains but is gone from domesticated lab strains) interacts with environmental factors (likely variable nutrition and growth rates) to alter juvenile oocyte survival, which in turn changes the strength of an oocyte-derived signal acting on thegonadal soma to maintain estrogen production, thereby preserving or not preserving oocytes and gonad fate. The aims are: Aim 1 is to learn the molecular genetic basis of the native zebrafish SD locus. Aim 2 is to understand the transcriptomic cascade during SD using: a) a native strain genotyped for genetic sex, b) an environmentally sensitive lab strain that lacks the native SD gene, and c) fish lacking PGCs to identify PGC- derived factors and their downstream targets. Aim 3 is to understand mechanisms of gonadal soma/germ cell reciprocal signaling by learning the roles of the related TGFss signalers Gdf9, Amh, and Gsdf (gonadal soma derived factor), and to discover how the female-specifying signals Wnt4 and Foxl2 act in gonadal development by using induced mutations, many of which we have in hand. Innovation of proposed work includes the identification and unique use of zebrafish possessing and genotyped for the native SD locus; our generally applicable RAD-sex method for identifying sex loci in different populations; the unique mutant and transgenic resources that we make available; and our focus on gonadal soma/germ cell reciprocal signaling in zebrafish sex determination. Outcomes and impact of this work are the potential for a fuller understanding of the molecular genetic nature of the vertebrate sex-balance mechanism, which will increase our appreciation of possible mechanisms underlying increases in human reproductive diseases in developed countries.