Regulation and functional effects of localized RNAs

局部RNA的调控和功能效应

• 批准号: 10014707
• 负责人: Stavroula Mili
• 金额: $ 128.4万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10014707
• 关键词: 3-Dimensional; Active Biological Transport; Adenomatous Polyposis Coli; Amyotrophic Lateral Sclerosis; Animal Model; Area; Biological Models; Cell physiology; Cells; Colorectal Neoplasms; Coupled; Cytoplasm; Defect; Destinations; Disease; Disease Progression; Environment; Epithelial; Epithelial Cells; Event; Goals; Individual; Investigation; Mammalian Cell; Microtubules; Modeling; Molecular; Mutate; Neoplasm Metastasis; Nerve Degeneration; Pathologic; Pathway interactions; Peripheral; Physiological; Positioning Attribute; Process; Production; Proteins; RNA; RNA-Binding Protein FUS; Regulation; Role; Site; Surface; System; Testing; Transcript; Translating; Translations; Tumor Suppressor Proteins; Tumor stage; Work; cancer cell; cancer type; cell motility; combat; extracellular; functional outcomes; in vivo; interest; intestinal epithelium; mechanical properties; migration; novel; physical state; therapeutic target; tool; tumor; tumorigenesis

A large number of RNAs are not diffusely distributed in the cytoplasm, but are actively transported to various subcellular sites. After reaching their final destinations, localized RNAs are translated, thus directing local protein production. While increasing numbers of localized RNAs are being identified, the functional importance of these events, as well as the overall applicability of the above model, is not well understood. We are focusing on a localization pathway that we have identified, which targets a number of RNAs to cellular protrusions. This pathway is of interest because it targets RNAs encoding factors that have been implicated in progression of various types of cancers. Furthermore, as we have found, it is regulated by disease-associated proteins, such as APC (Adenomatous Polyposis Coli), a tumor-suppressor protein whose loss underlies the initiation of most colorectal tumors, as well as by the RNA-binding protein FUS, a protein mutated in cases of Amyotrophic Lateral Sclerosis. Our overall goals have been to: a) understand the basic mechanisms underlying localization of RNAs at cell protrusions, b) understand the functional outcomes of their localization, either as a group or at the level of individual transcripts and c) set up appropriate model systems and tools to study RNA localization events in physiologically and pathologically relevant settings. With regards to the underlying mechanisms, we have revealed that localization of RNAs at protrusions requires a specific subset of modified microtubules in a process that is disrupted by pathologic inclusions associated with neurodegeneration. We have further shown that RNA localization at protrusions is modulated by the mechanical properties of the extracellular environment and have probed into the molecular basis of this phenomenon. Finally, we have made the intriguing discovery that, contrary to the prevailing model, translation of protrusion-localized RNAs is not affected by their position in the cytoplasm, but is rather coordinated with specific peripheral cellular processes, being activated at extending protrusions and silenced upon protrusion retraction. This coordination is dynamic and is coupled to a change in the physical state of the RNA. With regards to the functional roles of protrusion-localized RNAs, we have shown that their localization is important for efficient cell migration. We have devised ways to disrupt localization of specific, individual RNAs and have shown that disrupting the localization of just one RNA is sufficient to cause significant migration defects in 2D and 3D systems, as well as to reduce collective invasion of cancer cells. We are optimizing our current approaches for in vivo delivery to further test the roles of protrusion-localized RNAs in animal models of tumor metastasis. Apart from migrating cells, our recent studies have revealed that APC-dependent RNAs are also localized in non-invasive, epithelial cells, where they are concentrated at the basal surface. A separate line of investigation is thus exploring the functional roles of basally localized APC-dependent RNAs in 2D or 3D epithelial systems. Given the role of APC as an initiator of tumorigenesis in the intestinal epithelium, we believe these studies might reveal roles of localized RNAs relevant to the initial stages of tumor formation.

大量RNA并未扩散地分布在细胞质中，而是积极运输到各种亚细胞位点。到达最终目的地后，将局部RNA翻译，从而指导局部蛋白质的产生。尽管正在识别出越来越多的局部RNA数量，但这些事件的功能重要性以及上述模型的整体适用性尚不清楚。我们专注于已确定的定位途径，该途径将许多RNA靶向细胞突起。该途径引起了人们的关注，因为它针对RNA的编码因子，这些因素与各种类型的癌症的进展有关。此外，正如我们发现的那样，它受疾病相关蛋白的调节，例如APC（腺瘤息肉大肠杆菌），一种肿瘤 - 抑制蛋白，其损失构成了大多数结直肠肿瘤的启动，以及RNA结合蛋白融合的蛋白质，一种蛋白质，一种在Amyotrophic cllopopic sclersis的蛋白质中突变的蛋白质。我们的总体目标是：a）了解RNA在细胞突起处的基本机制，b）了解其本地化的功能结果，无论是小组还是在单个转录本的级别和C）建立适当的模型系统和工具，以研究在生理和病理上相关的环境中研究RNA定位事件。关于潜在机制，我们透露，在突起处的RNA定位需要在与神经变性相关的病理夹杂物破坏的过程中特定的修饰微管子集。我们进一步表明，在突起处的RNA定位是通过细胞外环境的机械性能调节的，并已探测为该现象的分子基础。最后，我们提出了一个有趣的发现，与普遍的模型相反，突出裂解的RNA的翻译不受它们在细胞质中的位置的影响，而是与特定的外围细胞过程相当协调，并在延伸突出时被激活并在缩短时被激活。这种协调是动态的，并耦合到RNA的物理状态的变化。关于突发定位的RNA的功能作用，我们表明它们的定位对于有效的细胞迁移很重要。我们已经设计了破坏特定单个RNA定位的方法，并表明破坏仅一个RNA的定位足以在2D和3D系统中引起明显的迁移缺陷，并减少癌细胞的集体入侵。我们正在优化当前的体内递送方法，以进一步测试突出 - 局部RNA在肿瘤转移动物模型中的作用。除了迁移的细胞外，我们最近的研究表明，依赖APC的RNA也位于非侵入性上皮细胞中，它们集中在基础表面。因此，单独的研究线正在探索2D或3D上皮系统中基本局部依赖APC的RNA的功能作用。鉴于APC是肿瘤发生在肠上皮中的作用，我们认为这些研究可能揭示了与肿瘤形成初始阶段有关的局部RNA的作用。