Understanding biological functions of repeat-containing noncoding RNAs

传统上认为DNA通过编码具有不同酶、调节和结构功能的大量蛋白质来存储有关我们身体发育和功能的信息。然而，只有约2%的人类基因组实际上用于产生随后翻译成蛋白质的RNA信使。其余98%的大部分——通常被称为基因组的“暗物质”——最近被证明产生了相对较长的非蛋白编码rna (lncrna)，其功能尚不清楚。值得注意的是，我们基因组的这一部分也富含重复DNA序列，包括所谓的短串联重复序列（STRs）。由于基因突变而扩增的一些异常长的STR阵列的表达，通过将重要的rna结合蛋白（rbp）的多个拷贝招募到重复序列单元，已知会导致毁灭性的退行性疾病。另一方面，健康人类基因组中编码的大多数STRs的表达状况和可能的生物学功能尚未得到系统的研究。我们提出通过与相应的rbp形成多重接触并控制其活性和细胞定位，来验证含有str的lncRNAs作为细胞RNA代谢的普遍调节剂的假设。我们将通过追求两个相互关联的目标来发展我们的计划。在第一个目标中，我们将估计基因组中编码的STR-lncRNAs的数量，并询问这些分子如何参与RBP调控。为此，我们将通过使用计算和实验工具的创新组合，鉴定在生物医学上重要样本（包括癌细胞和发育中的神经元）中表达的str - lncrna。新发现的str - lncrna中最有趣的例子将被检查其RBP相互作用特性，定位以及在塑造细胞基因表达中的可能作用。在第二个目标中，我们将重点关注我们在初步工作中发现的称为PNCTR的特定STR-lncRNA。我们已经知道，PNCTR在癌细胞中过量产生，它聚焦一个关键的RBP，多嘧啶束结合蛋白（PTBP1），以高级别和转移性肿瘤的点状模式为特征。此外，PNCTR可能是细胞存活和增殖所必需的。我们将通过阐明控制PNCTR表达的机制及其在癌症生物学中的潜在功能贡献来继续这条研究路线。总而言之，这些研究将大大提高我们对人类基因组“暗物质”的理解，为RNA代谢的调控提供基本见解，并最终为疾病诊断和治疗开辟新的途径。

DNA has been traditionally thought to store information about development and function of our body by encoding numerous proteins with distinct enzymatic, regulatory and structural functions. Yet, only ~2% of the human genome is actually used for this purpose by producing RNA messengers subsequently translated into proteins. A large fraction of the remaining 98% - often referred to as genome's "dark matter" - has been recently shown to give rise to relatively long non-protein-coding RNAs (lncRNAs) with poorly understood functions. Notably, this part of our genome is also enriched in repeated DNA sequences including so-called short tandem repeats (STRs). Expression of a few aberrantly long STR arrays expanded as a result of genetic mutations is known to lead to devastating degenerative diseases by recruiting multiple copies of important RNA-binding proteins (RBPs) to the repeated sequence units. On the other hand, the expression status and possible biological functions of most STRs encoded in healthy human genomes have not been investigated systematically.We propose to test the hypothesis that STR-containing lncRNAs function as pervasive regulators of cellular RNA metabolism by forming multiple contacts with corresponding RBPs and controlling their activity and cellular localization. We will develop our program by pursuing two interrelated objectives.In the first objective, we will estimate the number of STR-lncRNAs encoded in the genome and ask how these molecules can contribute to RBP regulation. For this purpose, we will identify STR-lncRNAs expressed in biomedically important samples - including cancer cells and developing neurons - by using an innovative combination of computational and experimental tools. The most interesting examples of the newly discovered STR-lncRNAs will be examined for their RBP interaction properties, localization, and possible role in shaping cellular gene expression.In the second objective, we will focus on a specific STR-lncRNA called PNCTR that we uncovered in our preliminary work. We already know that PNCTR is over-produced in cancer cells where it focuses a critical RBP, polypyrimidine tract-binding protein (PTBP1), in a dot-like pattern characteristic of high-grade and metastatic tumors. Moreover, PNCTR might be required for cell survival and proliferation. We will continue this line of research by elucidating the mechanisms controlling PNCTR expression and underlying its functional contributions to cancer biology.All in all, these studies should substantially improve our understanding of the "dark matter" of the human genome, provide fundamental insights into regulation of RNA metabolism, and ultimately open up new avenues in disease diagnosis and treatment.