mRNA Localization in Organelle Biogenesis

描述（由申请人提供）：了解调节内膜细胞器维持和生物发生的分子原理是细胞生物学中的一个原则问题。迄今为止的研究已经建立了支持内膜细胞器维持的蛋白质/膜运输途径，并且通过这些研究，已经确定了ER作为常驻内膜蛋白质合成位点的核心重要性。然而，内膜细胞器生物发生的分子基础仍然是神秘的。 这项研究的目的是确定ER定向mRNA定位在内膜细胞器生物发生中的作用。已经确定的是，真核细胞的蛋白质运输途径还指导mRNA转录组在细胞中的两个主要蛋白质合成区室（细胞质和内质网（ER））之间的分配。因此，编码分泌/整合膜蛋白的mRNA通过信号识别颗粒（SRP）途径定位于ER，而编码胞质/核质蛋白的mRNA（其缺乏编码的信号序列）保留胞质中的默认定位。虽然这种正选择模型已经建立，但我们最近报道了i）编码内膜细胞器的驻留蛋白（mRNAendo）的mRNA可以通过SRP和非依赖性的途径定位于ER; ii）mRNAendo群组通过直接的、不依赖于核糖体的相互作用与ER结合; iii）mRNAendo群组的特征在于其异常高的ER富集。这些研究结果表明，mRNA定位服务的一个组成部分，但未探索，在细胞器的生物发生的作用。我们假设，自主定位内膜常驻蛋白编码mRNA的ER，和他们的直接，核糖体独立协会与ER膜，代表了一个自组织机制在细胞器的生物发生功能。这一假说将在以下具体目标中进行检验：1）在全基因组范围内建立内膜驻留蛋白编码mRNA的亚细胞分配模式; 2）鉴定将内膜驻留蛋白编码mRNA引导至ER的分子信号; 3）鉴定介导mRNA与ER膜的核糖体非依赖性缔合的RNA结合蛋白和ER驻留蛋白; 4）确定内膜蛋白编码mRNA与ER的核糖体非依赖性结合是否是细胞器生物发生所必需的。 真核细胞中mRNA分配的全基因组研究揭示了mRNA定位的亚细胞模式的意想不到的复杂性。在这个建议中，我们建议延长最近发现的mRNA集群特定模式的RNA定位到ER的细胞器生物发生的问题。如果成功的话，这些研究将为细胞器生物发生的分子原理提供基本的见解，并支持细胞器功能障碍疾病的治疗方法的发展。 公共卫生相关性：真核细胞的细胞器执行许多对细胞活力至关重要的功能。事实上，许多遗传性代谢疾病以及衰老疾病都是由于细胞器功能的破坏而引起的。这项研究的目标是确定细胞器如何创建和修复的基本原则，以便更好地理解基于细胞器的疾病，并确定基于细胞器的疾病的新疗法。

DESCRIPTION (provided by applicant): Understanding the molecular principles that regulate endomembrane organelle maintenance and biogenesis stands as a principle question in cell biology. Research to date has established the protein/membrane trafficking pathways that support endomembrane organelle maintenance and through these studies, the central importance of the ER as the site of resident endomembrane protein synthesis has been identified. The molecular basis for endomembrane organelle biogenesis remains, however, mysterious. The objective of the proposed research is to define the role of ER-directed mRNA localization in endomembrane organelle biogenesis. It is well established that the protein trafficking pathways of eukaryotic cells also direct the partitioning of the mRNA transcriptome between the two primary protein synthesis compartments in the cell, the cytoplasm and the endoplasmic reticulum (ER). Thus, mRNAs encoding secretory/integral membrane proteins are localized to the ER via the Signal Recognition Particle (SRP) pathway and mRNAs encoding cytosolic/nucleoplasmic proteins, which lack encoded signal sequences, retain default localization in the cytosol. Although this positive selection model is well established, we have recently reported that i) mRNAs encoding resident proteins of the endomembrane organelles (mRNAendo) can be localized to the ER via a SRP- and translation-independent pathway; ii) the mRNAendo cohort is bound to the ER via direct, ribosome-independent interactions; and iii) the mRNAendo cohort is distinguished by its exceptionally high ER enrichment. These findings suggest that mRNA localization serves an integral, though unexplored, role in organelle biogenesis. We hypothesize that the autonomous localization of endomembrane resident protein-encoding mRNAs to the ER, and their direct, ribosome-independent association with the ER membrane, represents a self-organization mechanism functioning in organelle biogenesis. This hypothesis will be tested in the following specific aims: 1) Establish, at a genome-wide scale, the subcellular partitioning patterns of endomembrane resident protein-encoding mRNAs; 2) Identify the molecular signals that direct endomembrane resident protein-encoding mRNAs to the ER; 3) Identify the RNA binding proteins and ER resident proteins that mediate ribosome-independent association of mRNAs with the ER membrane; 4) Determine if ribosome-independent binding of endomembrane protein encoding mRNAs to the ER is essential for organelle biogenesis. Genome-wide studies of mRNA partitioning in eukaryotic cells have revealed an unexpected complexity in the subcellular patterns of mRNA localization. In this proposal, we propose to extend recent discoveries of mRNA cohort-specific patterns of RNA localization to the ER to the question of organelle biogenesis. If successful, these studies will provide fundamental insights into the molecular principles of organelle biogenesis and support the development of therapeutic approaches to diseases of organelle dysfunction. PUBLIC HEALTH RELEVANCE: The organelles of eukaryotic cells perform numerous functions that are essential for cell viability. Indeed, numerous inherited metabolic diseases as well as diseases of ageing arise through disruptions in organelle function. The goal of this research is to identify the basic principles of how organelles are created and repaired, so that organelle-based diseases can be better understood and new therapies for organelle- based disease identified.