Regulation of mRNA transport by multi-domain mRNA binding proteins

使用的技术(不变)：生物层干涉(BLI)、等温滴定热法(ITC)、质谱仪(MS)、核磁共振(NMR)光谱、非支架分析(SIA)、小角X射线散射(SAXS)、X射线结晶学。RNA结合蛋白(RBPs)通过招募特定的mRNAs到参与RNA加工、运输、降解和翻译的细胞机，以空间和时间的方式调节基因表达。RBP Syncrip是胚胎神经元发育的重要调节因子，也是了解局部mRNAs转运的重要系统。已知Syncrip以一种靶标依赖的方式结合一系列不同的RNA靶标，并发挥多种功能。Syncrip功能障碍会导致许多神经系统疾病。该蛋白质由一个N-末端结构域组成，我们最近证明它可以与RNA结合，我们被命名为N-末端RNA识别单元(NURR)和三个C-末端RRM结构域。基于mRNA相互作用组捕获数据，所有四个结构域都参与了mRNA的识别-尽管每个靶点的程度不同。这也适用于非mRNA靶标，例如，已被证明是HEXO基序与加载到外体上的miRNAs相互作用的关键结构域，以及对于外体加载。然而，这三个RRM结构域也参与了结合。我们建议研究Syncrip如何利用这些结构域来识别特定的mRNA靶标，这些靶标将被运输到特定的胚胎隔室。在这种情况下，我们将研究RRM结构域对特定RNA序列的识别，蛋白质在RNA分子包装中的作用，以及蛋白质通过磷酸化的调节--磷酸化将RNA结合蛋白质的RNA调节连接到信号网络。我们将同时研究信号和RNA识别之间的联系是如何在其他RNA结合蛋白中介导的。

Techniques used (unchanged):Bio-layer interferometry (BLI), isothermal titration calorimetry (ITC), mass spectrometry (MS), nuclear magnetic resonance (NMR) spectroscopy, scaffold-independent analysis (SIA), small-angle x-ray scattering (SAXS), x-ray crystallography.RNA-binding proteins (RBPs) regulate gene expression in a spatial and temporal manner, by recruiting specific mRNAs to cell machinery involved in RNA processing, transport, degradation and translation. The RBP Syncrip, is an important regulator of embryonic neuronal development, and is an important system to understand the transport of localised mRNAs. Syncrip is known to bind a range of different RNA targets, and exert multiple functions, in a target-dependent manner. Dysfunction of Syncrip results in a number of neurological diseases. The protein consists of an N-terminal domain, which we have recently demonstrated to bind RNA and that we have named N-terminal Unit for RNA Recognition (NURR) and of three C-terminal RRM domains. Based on mRNA interactome capture data, all four domains are involved in mRNA recognition - albeit to different extents with each target. This is observed also for non-mRNA targets, for example, it has been shown that the NURR domain is the key domain for the interaction of hEXO motif on miRNAs that are loaded onto exosomes, and for exosomal loading. However, the three RRM domains also participate in the binding. We propose to examine how Syncrip utilises these domain to recognise specific mRNA targets to be transported to specific embryonic compartments. In this context, we will be looking at the recognition of specific RNA sequences by the RRM domains, at the role of the protein in packaging of the RNA molecule and at the protein's regulation by the phosphorylation - which connects RNA regulation by RNA-binding proteins to signalling networks. We will in parallel look at how the link between signalling and RNA recognition is mediated in other RNA-binding proteins.