RNA binding proteins and mRNA localisation in Drosophila

mRNA在特定亚细胞区域的定位是极化细胞中普遍存在的现象，在发育（如图案）和分化细胞类型（如长期记忆）中都是至关重要的。亚细胞定位的mrna必须与特定的rna结合蛋白相互作用，以确保其正常定位/锚定。通常它们也处于翻译控制之下，以确保它们仅在目标位置被翻译。这些现象已经在几个系统中得到了研究，尤其是在发育模式、细胞迁移和神经元中。然而，通过高通量研究发现了许多亚细胞定位的mrna，但尚未研究其定位和翻译的机制。我们已经发现了一组局部的mrna，它们特别存在于果蝇精子的生长端，形状类似于彗星或杯子。我们还发现了一组已知的rna结合蛋白也定位在这个区域。在这个项目中，你将研究rna结合蛋白在定位这些特定mrna中的潜在作用。RNA定位和翻译将在RNA结合蛋白突变体中进行研究。蛋白质- rna相互作用将在体外用纯化组分和提取物（允许三元络合物形成）进行测定。您还将确定定位mrna和rna结合蛋白的突变是否以及如何影响发育中的精细胞尾尖的复杂结构。这将为进一步分析这一新颖且几乎未表征的一组局部mrna提供基础。例如，一旦确定了定位的基本参数，使用超分辨率方法，以及生化和功能水平上的相互作用，系统生物学和数学建模方法就可以应用。目的：描述和比较彗星和杯状mrna以及rna结合蛋白在精子生长末端的定位。-确定已知的rna结合蛋白对任何彗星和杯状mrna的定位是否重要。-识别和表征定位mrna与rna结合蛋白之间的直接（或间接）蛋白质- rna结合相互作用。-研究彗星和杯状基因以及rna结合蛋白的突变是否会导致细长精子生长端细胞结构的缺陷。-揭示“彗星”和“杯状”转录本定位模式之间的关系。-确定已知rna结合蛋白是否调控comet和cup mRNA的翻译。

mRNA localisation to specific sub-cellular regions is a widespread phenomenon in polarised cells and is critical both during development (eg for patterning) and in differentiated cell types (eg for long term memory). Sub-cellularly localised mRNAs must interact with specific RNA-binding proteins to ensure their normal localisation/ anchoring. Typically they are also under translational controls, to ensure they are only translated when at the target location. These phenomena have been studied in several systems, most notably in developmental patterning, cell migration and neurons. However many sub-cellularly localised mRNAs have been discovered through high-throughput studies, and the mechanisms underlying their localisation and translation have not been investigated. We have discovered a set of localised mRNAs, which are found specifically at the growing ends of Drosophila spermatids, in patterns resembling comets or cups. We also discovered a set of known RNA-binding proteins that also localise to this region. In this project you will investigate potential roles of the RNA-binding proteins in localising these specific mRNAs. RNA localisation and translation will be investigated in RNA-binding protein mutants. Protein-RNA interactions will be assayed in vitro with both purified components and extracts (to allow ternary complex formation). You will also determine whether, and how, mutations in the localised mRNAs and RNA-binding proteins affect the intricate structure of developing spermatid tail tips. This will provide the basis for a further analysis of this novel and virtually uncharacterised set of localised mRNAs. For example, systems biology and mathematical modelling approaches can be applied once the basic parameters of localisations, using super-resolution methods, and interactions at the biochemical and functional levels have been determined. Objectives - To describe and compare the comet and cup mRNAs', and RNA-binding proteins', localisations at the growing ends of spermatids. - To determine whether the known RNA-binding proteins are important for localisation of any comet and cup mRNAs. - To identify and characterise direct (or indirect) protein-RNA binding interactions between the localised mRNAs and the RNA-binding proteins. - To investigate whether mutations in comet and cup genes, and the RNA-binding proteins, cause defects in the cellular structure at the growing ends of elongating spermatids. - To uncover the relationship between "comet" and "cup" transcript localisation patterns. - To determine whether the known RNA-binding proteins regulate comet and cup mRNA translation.