Regulation and functional effects of localized RNAs

局部RNA的调控和功能效应

基本信息

批准号：
9556607
负责人：
Stavroula Mili
金额：
$ 90.37万
依托单位：
DIVISION OF BASIC SCIENCES - NCI
依托单位国家：
美国
项目类别：
财政年份：
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/9556607
关键词：
Active Biological Transport Actomyosin Adenomatous Polyposis Coli Adenomatous Polyposis Coli Protein Affect Amyotrophic Lateral Sclerosis Area Behavior Biochemical Biological Assay Carboxypeptidase Cell Culture System Cell Nucleus Cells Colorectal Cancer Complex Cytoplasm Cytoplasmic Inclusion Cytoskeleton Destinations Disease Environment Event Exhibits Extracellular Matrix Genes Goals Kinesin Link Liquid substance Malignant Neoplasms Membrane Messenger RNA Microscopy Microtubules Motor Mutation Nerve Degeneration Neurodegenerative Disorders Pathway interactions Phase Prions Process Production Property Proteins RNA RNA-Binding Protein FUS Regulation Ribonucleoproteins Role Site Testing Translating Translations Tubulin Tumor Suppressor Proteins Work base cell motility extracellular interest migration mutant novel prevent tumor progression

项目摘要

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 is not well understood. We are focusing on a localization pathway that we have identified, which targets a number of RNAs to the tips of cellular protrusions. We have found that an important component of this pathway is the tumor-suppressor protein Adenomatous Polyposis Coli (APC) whose mutation is the initiating event in the progression of the majority of colorectal cancers. APC associates with RNAs at cellular protrusions in ribonucleoprotein complexes, which we term APC-RNPs. In our recent work we have found that localization of these RNAs is required for efficient cell migration and invasion. Furthermore, we have discovered that the stiffness of the extracellular matrix promotes localization of APC-RNPs and we have delineated the underlying mechanism. Specifically, we found that increased stiffness of the extracellular environment promotes actomyosin contractility, which in turn enhances formation of a specific class of detyrosinated microtubules. These stable microtubules are required for localization and/or anchoring of APC-RNPs at protrusions. Given that the stiffness of the extracellular environment promotes various aspects of metastatic behavior, we are currently focusing on testing whether the mechanism we have uncovered is involved in promoting migration of metastatic cells. An additional component of APC-RNPs is the RNA-binding protein FUS/TLS. FUS belongs to a group of prion-domain containing proteins which exhibit the property of condensing to form liquid-demixed phases, which appear to underlie the formation of various membrane-less compartments in the nucleus and cytoplasm. In the case of FUS, mutations linked to Amyotrophic Lateral Sclerosis (ALS) promote its propensity to form such types of cytoplasmic inclusions. We have been focusing on understanding how formation of these cytoplasmic inclusions impacts on localization and function of APC-RNPs. We have previously shown that ALS-associated mutants of FUS mislocalize APC-RNPs and misdirect their translation. In our more recent work we have shown that the mechanism through which FUS inclusions affect RNA localization is indirect and involves a novel function of the kinesin-1 motor protein. Specifically, we find that kinesin-1 has a novel role in promoting formation of detyrosinated microtubules through targeting the tubulin carboxypeptidase activity. In the presence of FUS inclusions, both the kinesin-1 mRNA and the kinesin-1 protein become sequestered within FUS inclusions. This prevents efficient formation of detyrosinated microtubules leading to mislocalization of APC-RNPs. These findings reveal a novel mechanism regulating the microtubule cytoskeleton which could be a contributing factor in ALS neurodegeneration. We are additionally interested in investigating any contribution of this novel pathway in cancers containing translocations of the FUS gene.

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。尽管正在识别出局部RNA数量的增加，但这些事件的功能重要性尚不清楚。我们专注于已确定的定位途径，该途径将许多RNA靶向细胞突起的尖端。我们发现，该途径的重要组成部分是肿瘤 - 抑制蛋白腺瘤性息肉大肠杆菌（APC），其突变是大多数结直肠癌进展的起始事件。 APC与核糖核蛋白复合物中细胞突起的RNA相关联，我们称其为APC-RNP。在我们最近的工作中，我们发现这些RNA的定位是有效的细胞迁移和侵袭所必需的。此外，我们发现细胞外基质的刚度促进了APC-RNP的定位，并且已经描述了基本机制。具体而言，我们发现细胞外环境的刚度增加会促进肌动菌素的收缩力，从而增强了特定类别的驱虫微管的形成。这些稳定的微管是在突起处定位和/或锚定APC-RNP所必需的。鉴于细胞外环境的刚度促进了转移行为的各个方面，我们目前正在专注于测试我们发现的机制是否参与促进转移细胞的迁移。 APC-RNP的另一个组成部分是RNA结合蛋白FUS/TLS。 FUS属于一组含有蛋白质的prion域，这些蛋白具有凝结的特性，形成了液体二个相的相，这些相似乎是核和细胞质中各种无膜室形成的基础。在FUS的情况下，与肌萎缩性外侧硬化症（ALS）相关的突变促进了其形成这种类型的细胞质内包含的倾向。我们一直专注于理解这些细胞质内含物的形成如何影响APC-RNP的定位和功能。我们先前已经表明，与ALS相关的FUS突变体错误地定位APC-RNP并误导其翻译。在我们最近的工作中，我们表明，FUS夹杂物影响RNA定位的机制是间接的，并且涉及驱动蛋白-1运动蛋白的新功能。具体而言，我们发现驱动蛋白-1通过靶向微管蛋白羧肽酶活性在促进驱虫微管的形成中具有新颖的作用。在存在FUS夹杂物的情况下，驱动蛋白-1 mRNA和驱动蛋白-1蛋白都被隔离在FUS夹杂物中。这可以防止有效地形成驱散微管，从而导致APC-RNP的错误定位。这些发现揭示了一种调节微管细胞骨架的新机制，这可能是ALS神经变性的一个因素。我们还有兴趣研究包含FUS基因易位的癌症中这种新途径的任何贡献。