The Role of Rbpms2 in establishing oocyte polarity

Rbpms2 在建立卵母细胞极性中的作用

• 批准号: 9129301
• 负责人: Odelya Hartung Kaufman
• 金额: $ 0.85万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE, INC
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-01-15 至 2016-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9129301
• 关键词: 3' Untranslated Regions; Address; Affect; Animals; Antibodies; Assisted Reproductive Techniques; Balbiani Body; Binding; Biochemistry; Biological; Cell division; Chimeric Proteins; Clinic; Complementary DNA; Complex; Congenital Abnormality; Cytoplasm; Data; Defect; Deposition; Detection; Development; Diagnostic; Elements; Embryo; Embryonic Development; Feedback; Fertility; Fertilization; Fertilization in Vitro; Fishes; Future; Gene Structure; Genes; Genetic; Genetic Models; Genome; Genomics; Genotype; Germ Lines; Human; Image; Imagery; Implant; Introns; Invertebrates; Lead; Mammals; Maps; Mediating; Modeling; Molecular; Multiple Birth Offspring; Mutation; N-terminal; Nature; Oocytes; Oogenesis; Organelles; Organism; Ovary; Pattern; Phenotype; Point Mutation; Post-Transcriptional Regulation; Pregnancy; Premature Birth; Prevalence; Process; Proteins; RNA; RNA Splicing; RNA-Binding Proteins; Rana; Recruitment Activity; Regulation; Regulator Genes; Regulatory Element; Ribonucleoproteins; Role; Side; Spontaneous abortion; Staging; Structure; System; Techniques; Testing; Tissues; Transcript; Transgenes; Transgenic Organisms; Translations; Vertebrates; Work; Yeasts; Zebrafish; advanced maternal age; base; cell fate specification; egg; follow-up; gain of function; genetic approach; implantation; improved; in vivo; loss of function; mutant; overexpression; public health relevance; reproductive; tool; trend; yeast two hybrid system; zygote

 DESCRIPTION (provided by applicant): With the societal trend toward delaying pregnancy until advanced maternal ages, the reliance on assisted reproductive techniques is growing rapidly. A fundamental question for the field of In Vitro Fertilization (IVF) has been that of distinguishing healthy eggs and subsequent embryos from unhealthy ones. The inability to differentiate healthy embryos from unhealthy ones has resulted in the propensity for Fertility Clinics to transfer multiple embryos in the hopes that one will implant and result in a successful pregnancy. This practice has resulted in increased prevalence of multiple births that are inherently riskier and often result in premature delivery. In order to improve our understanding of embryos suitable for implantation, it is imperative that we understand the molecular mechanisms that make a healthy egg develop into a healthy embryo. Formation and maturation of a developmentally competent egg requires that RNAs and proteins within the egg cytoplasm be poised to direct early zygotic development after fertilization. This is especially crucial when considering that the first cell divisions and cell fate specifications occur before the zygotic genome is active; thus, maternally deposited gene products are the sole drivers of early embryonic development. We use the zebrafish to study this process because zebrafish eggs are fertilized externally, and the embryos develop outside of the body and are optically transparent. This allows for clear visualization of early phenotypes that often result in embryonic lethality. Additionally, zebrafish have excellent genetic tools and are exceptionally fertile, layig hundreds of eggs in a week, and are thereby a very powerful model for the genetic basis of egg and embryonic defects. In zebrafish and frogs, maternally deposited gene products include asymmetrically distributed RNAs and proteins responsible for establishing the germ line and the first embryonic axis. In mammals, it has not yet been determined if asymmetries in the egg are required for patterning of the embryonic axes. However, the oocytes of all animals (including humans) contain an asymmetrical structure, the Balbiani body (Bb), which is a transient subcellular aggregation of organelles and gene products. In zebrafish, the formation of the Bb is essential to deliver patterning molecules to one side of the egg. To date, only one gene has been demonstrated as necessary for Bb formation, the zebrafish bucky ball gene. We hypothesize that Bucky ball mediates assembly of the Balbiani body by interacting with RNA- binding proteins that, in turn, recruit patterning RNAs to this structure. To address this hypothesis, we will use classical genetics gain-of-function and loss-of-function approaches to assess the role of a conserved RNA- binding protein (RNAbp) in oocyte and embryonic patterning. Further, we will use biochemistry techniques to determine the nature of the interaction between Bucky ball and this RNAbp, and follow up this analysis with in vivo characterization on proteins lacking the interaction domains. Through imaging of transgenic and mutant zebrafish oocytes and embryos, this project will advance our understanding of oocyte patterning in vertebrates.

 描述（由申请人提供）：随着推迟怀孕的社会趋势，直到高龄产妇，对辅助生殖技术的依赖正在迅速增长。体外受精（IVF）领域的一个基本问题是区分健康的卵子和不健康的胚胎。无法区分健康胚胎与不健康胚胎导致生育诊所倾向于转移多个胚胎，希望其中一个胚胎能够植入并成功怀孕。这种做法导致多胎分娩的流行率增加，多胎分娩本身就具有更大的风险，往往导致早产。为了提高我们对 为了确保胚胎适合植入，我们必须了解使健康卵子发育成健康胚胎的分子机制。一个有发育能力的卵子的形成和成熟需要卵子细胞质中的RNA和蛋白质准备好指导受精后的早期合子发育。考虑到第一次细胞分裂和细胞命运规范发生在合子基因组活跃之前，这一点尤其重要;因此，母系沉积的基因产物是早期胚胎发育的唯一驱动力。 我们使用斑马鱼来研究这个过程，因为斑马鱼的卵是在体外受精的，胚胎在体外发育，并且是光学透明的。这允许清楚地观察早期表型，这些表型通常导致胚胎发育不良。 杀伤力此外，斑马鱼具有出色的遗传工具，并且异常肥沃，一周内产下数百个卵，因此是卵和胚胎缺陷遗传基础的非常强大的模型。在斑马鱼和青蛙中，母体沉积的基因产物包括不对称分布的RNA和蛋白质，负责建立生殖系和第一个胚胎轴。在哺乳动物中，尚未确定卵中的不对称是否是胚胎轴图案化所必需的。然而，所有动物（包括人类）的卵母细胞都含有不对称的结构，Balbiani体（Bb），这是细胞器和基因产物的瞬时亚细胞聚集体。在斑马鱼中，Bb的形成对于将图案化分子传递到卵的一侧至关重要。迄今为止，只有一个基因已被证明是必要的Bb形成，斑马鱼巴基球基因。 我们假设巴基球通过与RNA结合蛋白相互作用来介导巴尔比亚尼体的组装，RNA结合蛋白反过来又将图案化RNA招募到该结构中。为了解决这一假设，我们将使用经典的遗传学功能获得和功能丧失方法来评估保守的RNA结合蛋白（RNAbp）在卵母细胞和胚胎模式中的作用。此外，我们将使用生物化学技术来确定Bucky球和这种RNAbp之间的相互作用的性质，并在缺乏相互作用结构域的蛋白质的体内表征后进行这种分析。通过转基因和突变斑马鱼卵母细胞和胚胎的成像，该项目将促进我们对脊椎动物卵母细胞模式的理解。