Control of Oocyte Maturation in C. elegans

描述（由申请人提供）：我们的目标是确定线虫细胞间信号协调减数分裂和受精的机制。在有性生殖的动物中，卵母细胞在二倍体或分裂时停止，并在激素的作用下恢复减数分裂（减数分裂成熟）。染色体分离错误在女性减数分裂I是人类出生缺陷的主要原因。与年龄相关的卵巢激素环境变化是一个可能的原因。我们的研究确立了秀丽隐杆线虫作为研究激素控制减数分裂成熟的遗传范例。秀丽隐杆线虫的精子输出主要精子蛋白（MSP）激素来触发减数分裂成熟。在之前的研究中，我们发现卵泡样鞘细胞中的gs -腺苷酸环化酶信号是所有已知的生殖系中msp依赖性减数分裂成熟反应所必需的。相反，当MSP缺失时，VAB-1 MSP/Eph受体在卵母细胞抑制减数分裂成熟中起非必需作用。本应用的具体目的是测试一个体细胞控制减数分裂成熟的模型，其中鞘细胞控制卵母细胞的反应。AIM 1定义鞘细胞如何接收MSP信号。我们将验证MSP结合多个G蛋白偶联受体（gpcr）促进减数分裂成熟的假设。FRET测量将询问MSP是否激活鞘细胞中的gs -腺苷酸环化酶途径。一系列的功能测试将检验鞘细胞表达的gpcr在减数分裂成熟中的作用。保守的msp结合gpcr可能介导哺乳动物中msp相关配体的信号转导。AIM 2阐明鞘细胞如何将MSP信号传递给卵母细胞。我们将测试体细胞camp依赖性蛋白激酶A通过拮抗两种抑制途径促进减数分裂成熟的假设。基因测试将确定PKA是否在鞘细胞中是减数分裂成熟所必需的，以及单个acy-4抑制位点是否在PKA下游起作用。我们将从分子上识别关键的抑制基因座。这些研究将确定鞘细胞如何将MSP的存在传递给卵母细胞。AIM 3剖析了减数分裂成熟所需的翻译控制机制。保守的锌指蛋白OMA-1和OMA-2 （OMA蛋白）在减数分裂成熟过程中是冗余必需的。蛋白质组学分析表明，OMA蛋白具有转录后基因调控功能。我们将验证OMA蛋白在MSP缺失时抑制减数分裂成熟因子mrna翻译的假设，但在MSP存在时激活翻译。我们将首先使用rna结合蛋白免疫沉淀微阵列分析来鉴定OMA-核糖核蛋白中的mrna。然后，我们将确定OMA蛋白、MSP信号和OMA-核糖核蛋白组分对关键靶mrna表达的影响。这些研究将为减数分裂成熟的翻译调控提供见解。

DESCRIPTION (provided by applicant): Our objective is to define the mechanisms by which intercellular signaling coordinates meiosis and fertilization in the nematode C. elegans. In sexually reproducing animals, oocytes arrest at diplotene or diakinesis and resume meiosis (meiotic maturation) in response to hormones. Chromosome segregation errors in female meiosis I are the leading cause of human birth defects. And age-related changes in the hormonal environment of the ovary are a suggested cause. Our research established C. elegans as a genetic paradigm for studying hormonal control of meiotic maturation. C. elegans sperm export the major sperm protein (MSP) hormone to trigger meiotic maturation. A major advance in the prior award period was our finding that G1s-adenylate cyclase signaling in the follicle-like sheath cells is required for all known MSP-dependent meiotic maturation responses in the germline. By contrast, the VAB-1 MSP/Eph receptor plays a non-essential role in the oocyte to inhibit meiotic maturation when MSP is absent. The Specific Aims of this application test a model for the somatic control of meiotic maturation in which the sheath cells control the oocyte response. AIM 1 defines how the sheath cells receive the MSP signal. We will test the hypothesis that MSP binds multiple G protein-coupled receptors (GPCRs) to promote meiotic maturation. FRET measurements will ask whether MSP activates the G1s-adenylate cyclase pathway in sheath cells. A series of functional tests will examine the role of sheath cell-expressed GPCRs in meiotic maturation. Conserved MSP-binding GPCRs may mediate the signaling of MSP-related ligands in mammals. AIM 2 elucidates how the sheath cells transduce the MSP signal to the oocyte. We will test the hypothesis that somatic cAMP-dependent protein kinase A promotes meiotic maturation by antagonizing two inhibitory pathways. Genetic tests will address whether PKA is required in the sheath cells for meiotic maturation and whether individual acy-4 suppressor loci function downstream of PKA. We will molecularly identify critical suppressor loci. These lines of investigation will define how the sheath cells communicate the presence of MSP to the oocyte. AIM 3 dissects translational control mechanisms required for meiotic maturation. The conserved zinc finger proteins OMA-1 and OMA-2 (OMA proteins) are redundantly required for meiotic maturation. Proteomic analyses suggest that OMA proteins function as post-transcriptional gene regulators. We will test the hypothesis that OMA proteins function to repress translation of meiotic maturation factor mRNAs when MSP is absent, but activate translation when MSP is present. We will first identify mRNAs in OMA- ribonucleoproteins using RNA-binding protein immunoprecipitation-microarray profiling. We will then determine the effects of OMA proteins, MSP signaling, and OMA-ribonucleoprotein components on expression of key target mRNAs. These studies will provide insights into translational regulation of meiotic maturation. PUBLIC HEALTH RELEVANCE: Chromosome segregation errors in female meiosis I are the leading cause of human birth defects and age- related changes in the hormonal environment of the ovary are a suggested cause. The small roundworm C. elegans is an established model for studying the hormonal control of meiosis. These studies will define the signaling mechanisms controlling female meiosis and provide insights into the origin of meiotic errors.