Mechanisms of mRNA localization and translational control in Drosophila development

项目摘要 我们的长期目标是了解时空调控基因表达的转录后机制， 动物发育过程中的活动。我们专注于细胞内mRNA的定位，翻译控制， 降解，其在调节来自母体提供的转录物的蛋白质产生中起关键作用。 这些转录本是几乎所有动物最初发育程序的基础， 蛋因此，控制在何处以及何时使用它们来生产蛋白质必须在生产后进行。 转录。我们的研究利用了高度可操作的果蝇模型，该模型在很大程度上依赖于 母体转录本，研究产生不对称母体mRNA和蛋白质的机制， 分布所需的胚胎体图案和种系发育。我们之前的工作表明， 许多mRNA通过掺入到大的， 非膜结合的核糖核蛋白（RNP）组件称为胚芽颗粒。在胚胎发育过程中， 颗粒将这组mRNA递送到原始生殖细胞，在那里它们支持生殖细胞发育。 生殖颗粒是动物界原始生殖细胞的标志。它们含有保守的 在RNA调节中具有促进分化、增殖和分化的共同功能的组分， 生殖细胞系的维持。细菌颗粒只是众多细胞RNP组件中的一种， 被描述为相变行为。mRNA调控如何在RNA背景下发生 然而，对更普遍的颗粒及其生物学意义仍然知之甚少。这样复杂而 生物学相关的RNP组件，胚芽颗粒是研究RNA定位，翻译， 控制和降解，特别是它们的生物学意义。我们的研究导致了一个细菌模型 颗粒“客户”RNA积累的自我募集和组织成同型集群。该模型 提供了一个框架，以调查如何mRNA自组织在颗粒和这种组织如何赋予 选择性控制共享颗粒内不同mRNA的翻译和稳定性。我们将联合收割机 定量高分辨率成像与强大的生化和遗传/基因组策略，以及新的 方法，打破颗粒，调查的基本原则，管理组织的颗粒客户端 RNA。这些研究将使我们能够破译之间的关系颗粒协会，翻译 活性和胚芽颗粒功能。鉴于转录后机制在基因工程中的广泛应用， 发展，以及颗粒中RNA区室化的流行，这项工作将产生广泛的影响。 从我们的研究中获得的机制见解也将为理解RNA缺陷提供基础 代谢与许多疾病相关，包括不孕症、癌症和神经变性。

PROJECT SUMMARY Our long-term goal is to understand post-transcriptional mechanisms that spatially and temporally control gene activity during animal development. We focus on intracellular mRNA localization, translational control, and degradation, which play crucial roles in regulating production of proteins from maternally supplied transcripts. These transcripts, which underpin the initial developmental program of nearly all animals, are pre-loaded in the egg. Thus, control over where and when they are deployed to produce protein must be exerted post- transcriptionally. Our research capitalizes on the highly manipulable Drosophila model, which relies heavily on maternal transcripts, to investigate mechanisms that generate asymmetric maternal mRNA and protein distributions needed for embryonic body patterning and germline development. Our previous work revealed that numerous mRNAs accumulate at the posterior of the Drosophila oocyte through their incorporation into large, non-membrane-bound ribonucleoprotein (RNP) assemblies called germ granules. During embryogenesis, germ granules deliver this cohort of mRNAs to the primordial germ cells, where they support germline development. Germ granules are a hallmark of primordial germ cells throughout the animal kingdom. They contain conserved components that indicate a common function in RNA regulation to promote the differentiation, proliferation, and maintenance of the germline. Germ granules are just one of a plethora of cellular RNP assemblies that have been characterized for their phase-transitioned behavior. How mRNA regulation occurs in the context of RNA granules more generally and its biological significance remain, however, poorly understood. As complex and biologically relevant RNP assemblies, germ granules are ideal for studying RNA localization, translational control, and degradation and, in particular, their biological significance. Our studies lead to a model for germ granule "client" RNA accumulation by self-recruitment and organization into homotypic clusters. This model provides a framework to investigate how mRNAs self-organize in granules and how this organization imparts selective control over the translation and stability of different mRNAs within shared granules. We will combine quantitative high resolution imaging with powerful biochemical and genetic/genomic strategies, as well as novel methods that disrupt granules, to investigate fundamental principles governing the organization of granule client RNAs. These studies will enable us to decipher the relationship between granule association, translational activity, and germ granule function. Given the widespread use of post-transcriptional mechanisms during development, and the prevalence of RNA compartmentalization in granules, this work will have broad impact. Mechanistic insights gained from our studies will also provide a foundation for understanding defects in RNA metabolism associated with numerous disorders including infertility, cancer, and neurodegeneration.