Regulation and functional effects of localized RNAs

大量的rna不是弥漫性地分布在细胞质中，而是被主动地转运到各个亚细胞部位。到达最终目的地后，局部rna被翻译，从而指导局部蛋白质的生产。虽然越来越多的定位rna被发现，但这些事件的功能重要性尚未得到很好的理解。我们正在专注于我们已经确定的定位途径，它将许多rna靶向到细胞突起的尖端。我们发现该途径的一个重要组成部分是肿瘤抑制蛋白腺瘤性息肉病大肠（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（我们称之为APC- rnps）中细胞突起处的rna相关。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硬度促进APC-RNPs的定位，我们已经描绘了潜在的机制。具体来说，我们发现细胞外环境刚度的增加促进了肌动球蛋白的收缩性，这反过来又增强了一类特定的去酪氨酸微管的形成。这些稳定的微管是定位和/或在突起处锚定APC-RNPs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，我们目前的重点是测试我们发现的机制是否参与促进转移细胞的迁移。APC-RNPs的另一个组成部分是rna结合蛋白FUS/TLS。FUS属于一组含有朊病毒结构域的蛋白质，其表现出凝聚形成液体-分离相的特性，这似乎是细胞核和细胞质中各种无膜室形成的基础。在FUS的病例中，与肌萎缩侧索硬化症（ALS）相关的突变促进其形成这种类型的细胞质包涵体的倾向。我们一直致力于了解这些细胞质包涵体的形成如何影响APC-RNPs的定位和功能。我们之前已经证明，als相关的FUS突变体错定位APC-RNPs并误导其翻译。在我们最近的工作中，我们已经表明FUS内含物影响RNA定位的机制是间接的，并且涉及运动蛋白1的新功能。具体来说，我们发现激酶1通过靶向微管蛋白羧肽酶活性，在促进去酪化微管的形成中具有新的作用。在FUS包涵体存在的情况下，激酶1 mRNA和激酶1蛋白都被隔离在FUS包涵体中。这阻止了去脱氧微管的有效形成，从而导致APC-RNPs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神经退行性变的一个因素。我们还对研究这种新途径在含有FUS基因易位的癌症中的作用感兴趣。

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.