Dissecting ribosome pausing during embryogenesis: from global and single molecule studies to whole embryo phenotypes

剖析胚胎发生过程中的核糖体暂停：从整体和单分子研究到整个胚胎表型

• 批准号: BB/X007294/1
• 负责人: Hilary Ashe
• 金额: $ 106.12万
• 依托单位: University of Manchester
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2024
• 资助国家: 英国
• 起止时间: 2024 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FX007294%2F1
• 关键词: Dissecting; ribosome; pausing; during; embryogenesis

An organism's genes are the blueprint from which proteins are produced, yet in any given single cell, only a subset of all genes is ultimately decoded to give proteins. As such, the identity and abundance of proteins made in a cell determines whether it will become, say a nerve cell in the brain or an epithelial cell in the intestinal lining. This is particularly striking during the development of an organism, where distinct sets of proteins are made at various stages in growth so that different cell types form. Proteins are chains of amino acids which form the building blocks of life. The order and identity of amino acids in a protein is decoded from an intermediate copy of genes called a mRNA in a process termed 'translation'. Ribosomes are the machines inside cells responsible for translation. They bind to the mRNA and sequentially insert amino acids onto a growing protein chain according to the code read from the mRNA. However, not all amino acid insertion events occur with the same efficiency and for many reasons ribosomes pause at sticky patches on a mRNA. In various biological contexts these ribosome pauses are important. However, only a handful of ribosome pausing events have been studied in great detail owing to the technical challenges of following ribosomes on single molecules of mRNA.Our research studies ribosome pausing using the classic model, the fruitfly Drosophila, since the development of an adult fruitfly from a fertilised egg only takes ten days and represents a wonderful system to study how changes in protein production affect the development of a complex multicellular animal. Even at the earliest stages in the fruitfly embryo, the basic segmented pattern of the adult body is becoming apparent. Therefore, the precisely timed production of key proteins at specific sites within the embryo dictates which tissues will develop, and when/ where they will do so. In this proposal, we aim to determine how ribosome pausing controls the timing of protein production so that the correct cell types form during embryonic development. To achieve this goal, we will use state-of-the-art microscopy and sequencing approaches, which will allow us to answer three key questions. Firstly, which mRNAs have paused ribosomes in the fruitfly embryo? Secondly, what is it about these mRNAs that makes the ribosomes pause? Thirdly, how does disrupting ribosome pausing affect development of the fruitfly embryo?Overall, our data will provide important new information about the signals that direct ribosome pausing and the range of processes affected during embryonic development. As all known mechanisms of translational control are conserved across animal and plant cells, results from this study will be directly relevant to human development. Therefore, our findings will be important for understanding the many human diseases that are associated with misregulation of translation elongation. Finally, our data will also benefit stem cell research, where the ability to manipulate gene expression to efficiently differentiate stem cells into particular cell types is a major therapeutic goal.

生物体的基因是产生蛋白质的蓝图，但在任何给定的单细胞中，只有所有基因的一个子集最终被解码以给出蛋白质。因此，细胞中蛋白质的身份和丰度决定了它是否会变成，例如大脑中的神经细胞或肠壁上的上皮细胞。这在生物体的开发过程中尤其引人注目，在生物体的开发中，在生长的各个阶段制造了不同的蛋白质，从而形成了不同的细胞类型。蛋白质是氨基酸的链，构成生命的基础。蛋白质中氨基酸的顺序和身份是从称为“翻译”的过程中从称为mRNA的中间副本解码的。核糖体是负责翻译的电池内部的机器。它们与mRNA结合，并根据从mRNA中读取的代码顺序将氨基酸插入生长的蛋白质链中。但是，并非所有氨基酸插入事件都以相同的效率发生，并且由于许多原因，核糖体在mRNA上的粘性斑块上暂停。在各种生物学环境中，这些核糖体的暂停很重要。然而，由于跟随核糖体在mRNA的单分子上遇到的核糖体的技术挑战，仅研究了少量核糖体暂停事件。由于使用经典模型，果蝇果蝇的发展核糖体暂停，因为果蝇果蝇的发展起来，因此从肥料的蛋白质中发展成成人果蝇，只需十天就可以花费十天，并且可以花费十天，并且代表了一种精彩的系统，可以研究构成动物的生产。即使在果蝇胚胎的最早阶段，成人身体的基本分段模式也变得显而易见。因此，胚胎中特定部位的关键蛋白质精确产生决定了哪些组织将发展出来，以及何时/何时会发展。在此提案中，我们旨在确定核糖体暂停如何控制蛋白质产生的时间，从而在胚胎发育过程中形成正确的细胞类型。为了实现这一目标，我们将使用最先进的显微镜和测序方法，这将使我们能够回答三个关键问题。首先，哪些mRNA在果蝇胚胎中暂停了核糖体？其次，这些使核糖体暂停的mRNA是什么？第三，破坏核糖体暂停如何影响果蝇胚胎的发展？总的来说，我们的数据将提供有关直接核糖体暂停以及在胚胎开发过程中受影响的过程范围的信号的重要新信息。由于在动物和植物细胞中均保留了所有已知的翻译控制机制，因此这项研究的结果将与人类发展直接相关。因此，我们的发现对于理解与翻译伸长延伸的许多人类疾病至关重要。最后，我们的数据还将受益于干细胞研究，其中操纵基因表达以有效区分干细胞为特定细胞类型的能力是一个主要的治疗目标。