Role of long non-coding RNAs in sarcoma pathogenesis

项目总结 尤文肉瘤是一种罕见的肿瘤，发生在儿童和年轻人中。其中的驱动力致癌事件 疾病是最常见的易位EWS/FLI。涉及ETS转录的相似易位 包括前列腺癌在内的其他恶性肿瘤中也存在这些因素，这扩大了我们的潜在相关性 学习。先前的工作已经确定，EWS/FLI可以通过抑制一些 增强剂，同时通过直接结合激活其他分子。我们自己的数据表明，EWS/FLI也 上调lncRNAs的表达。这项提议的主要目的是阐明分子功能。 尤因肉瘤中的IncRNA。我们的初步数据表明，两个LncRNA，EWSAT1和EWSAT2，可能是 参与尤文肉瘤的致癌作用。这两个RNA中的任何一个的敲击率都大幅下降 尤文肿瘤在体内外的增殖情况。此外，RNAseq分析表明，这两个 LncRNAs抑制了大量已知也被EWS/FLI抑制的基因。我们的主要目标是 这项建议是为了阐明这些lncRNA促进尤因的分子功能。 肉瘤。我们将把研究重点放在确定这些lncRNA是如何导致转录抑制的。 在目标1中，我们将阐明EWSAT1和EWSA2的表达如何改变关键的蛋白质-蛋白质相互作用 尤因细胞。具体地说，我们将测试EWSAT1或2表达改变EWS/FLI的PPI的假设， EWS或hnRNPK。在目标2中，我们将检验EWSAT1或EWSA2通过调节 EWS/FLI绑定到特定的增强剂。ChIPseq和ChIRPseq将用于确定这些lncRNAs 与染色质相互作用，以及它们如何改变组蛋白和其他染色质调节剂的结合。在《目标3》中，我们将 利用尤因患者来源阐明EWSAT1/2缺失或过表达的功能后果 异种移植和细胞系。

PROJECT SUMMARY Ewing sarcoma is a rare tumor that occurs in children and young adults. The driving oncogenic event in this disease is most commonly the translocation EWS/FLI. Similar translocations involving ETS transcription factors occur in other malignancies, including prostate cancer, broadening the potential relevance of our studies. Prior work has determined that EWS/FLI can lead to chromatin reprogramming by repressing some enhancers while activating others through direct binding. Our own data indicates that EWS/FLI also upregulates expression of lncRNAs. The primary goal of this proposal is to elucidate the molecular function of lncRNAs in Ewing sarcoma. Our preliminary data indicates that two lncRNAs, EWSAT1 and EWSAT2, may be involved in the oncogenic effects of Ewing sarcoma. Knock-down of either of these RNAs strongly decreases proliferation of Ewing tumors in vitro and in vivo. Furthermore, RNAseq analysis indicates that both of these lncRNAs repress a significant number of genes also known to be repressed by EWS/FLI. Our primary goal in this proposal is to elucidate the molecular function through which these lncRNAs act to promote Ewing sarcoma. We will focus our studies on determining how these lncRNAs lead to transcriptional repression. In Aim 1, we will elucidate define how expression of EWSAT1 and 2 alters key protein-protein interactions in Ewing cells. Specifically, we will test the hypothesis that EWSAT1 or 2 expression alters PPIs for EWS/FLI, EWS or hnRNPK. In Aim 2, we will test the hypothesis that EWSAT1 or 2 function by modulating the ability of EWS/FLI to bind to specific enhancers. ChIPseq and ChIRPseq will be used to determine how these lncRNAs interact with chromatin and how they alter binding of histones and other chromatin regulators. In Aim 3, we will elucidate the functional consequences of EWSAT1/2 loss or overexpression using Ewing patient derived xenografts and cell lines.