Nuclear-retained long noncoding RNAs

核保留的长非编码RNA

• 批准号: 8579244
• 负责人: Gordon G Carmichael
• 金额: $ 32.63万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-16 至 2018-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8579244
• 关键词: Address; Affinity; Binding; Boxing; Cell Cycle; Cell Nucleus; Cells; Chromatin; Data; Exonuclease; Family; Gene Expression Profile; Genes; Genomics; Hela Cells; Hereditary Disease; Human Genetics; In Vitro; Introns; Knowledge; Lead; Learning; Literature; Molecular; Names; Nuclear; Pathology; Pattern; Poly(A) Tail; Porifera; Prader-Willi Syndrome; Process; Production; Proteins; RNA Binding; RNA Processing; RNA Splicing; Role; Route; Small Nucleolar RNA; Small Nucleolar Ribonucleoproteins; Structure; Tissues; Untranslated RNA; cell type; genome-wide; human disease; human embryonic stem cell; in vivo; insight; interest; novel; prevent; public health relevance; research study; vector

DESCRIPTION (provided by applicant): We have recently discovered novel intron-derived long noncoding RNAs (lncRNAs) that are processed on both ends by the snoRNA machinery (we named these sno-lncRNAs). Most snoRNAs are processed from introns. After debranching, exonucleases degrade the introns but not the snoRNAs because these associate with specific factors that prevent degradation and lead to snoRNP assembly. Sno-lncRNAs are processed from introns that contain two snoRNAs. During exonucleolytic trimming, the sequences between the snoRNAs are not degraded, leading to the accumulation of lncRNAs lacking 5' cap structures or 3' poly(A) tails. Some of these RNAs can accumulate to very high levels in cells but were missed before because they are long, they derive from introns, and they lack poly(A) tails. These constitute an entirely new class of nuclear lncRNAs and this proposal is to study them in greater detail. Results so far suggest that whenever introns contain not one but two snoRNA genes, sno-lncRNAs can be produced. Such RNAs can be found in every cell examined so far, and appear to be widely expressed in tissues. Further, the genomic region encoding some of the most abundant of these RNAs (15q11-q13) is specifically deleted in an important human genetic disease, Prader-Willi Syndrome. We will characterize how sno-lncRNAs are made, what proteins associate with them and how many of them there are. In addition, we will ask what they do in cells and begin to address their possible role in Prader-Willi Syndrome. In the first aim, we will characterize the structure and expression of sno-lncRNAs. We will also examine their sub-nuclear localization, and efforts will be made to follow their synthesis and localization dynamically and throughout the cell cycle. Finally in this aim, we will carry out genomewide studies in several different cell types and using several different approaches in order to identify more sno-lncRNAs, some of which will be studied in greater detail. The second aim is to investigate the mechanism of action of sno-lncRNAs. Of particular interest is the connection of the chr15 sno-lncRNAs to Fox1 family splicing regulators. Compelling evidence suggests that these lncRNAs may bind Fox2 and perhaps Fox1 with high affinity. Thus, they may act as molecular "sponges" to titrate splicing factors since in preliminary experiments we have found that they alter splicing patterns. This angle will be examined in detail and could lead directly to new insights into Prader-Willi pathology. We will also ask whether some sno-lncRNAs can alter chromatin organization like a number of other lncRNAs do. Finally, since these new RNAs appear to be relatively stable and strictly retained in the nucleus, we will begin to use our knowledge of their expression to generate a new class of vectors for nuclear expression of a variety of sequences.

描述（由申请人提供）：我们最近发现了新型内含子源性的长期非编码RNA（LNCRNA），它们在SNORNA机械上都在两端处理（我们命名为这些SNO-LNCRNA）。大多数snornas都是通过内含子处理的。脱离分支后，外核酸酶会降低内含子，而不会降解snornas，因为这些与防止降解并导致SNORNP组装的特定因素相关。 SNO-LNCRNA是从包含两个snornas的内含子中处理的。在外核解整齐的修剪过程中，snornas之间的序列不会降解，从而导致缺乏5'帽结构或3'poly（a）尾巴的lncrnas积累。这些RNA中的一些可以在细胞中积累到很高的水平，但以前由于长期而丢失，它们源于内含子，并且缺乏poly（a）尾巴。这些构成了全新的核LNCRNA类别，该建议是更详细地研究它们。迄今为止的结果表明，每当内含子不包含一个而是两个snorna基因时，就可以产生sno-lncrnas。到目前为止，可以在检查的每个细胞中找到此类RNA，并且似乎在组织中广泛表达。此外，编码这些RNA中一些最丰富的基因组区域（15q11-Q13）被特异性地删除在重要的人类遗传疾病Prader-Willi综合征中。我们将表征如何制作SNO-LNCRNA，与它们相关的蛋白质以及其中有多少个蛋白质。此外，我们将询问它们在细胞中的作用，并开始解决它们在Prader-Willi综合征中的可能作用。在第一个目标中，我们 将表征sno-lncrnas的结构和表达。我们还将检查它们的亚核定位，并将努力在整个细胞周期中动态遵循它们的合成和定位。最后，在这个目标中，我们将以几种不同的细胞类型进行全基因组研究，并使用几种不同的方法来识别 更多的SNO-LNCRNA，其中一些将进行更详细的研究。第二个目的是研究SNO-LNCRNA的作用机理。特别有趣的是CHR15 SNO-LNCRNA与FOX1家族剪接调节器的联系。令人信服的证据表明，这些LNCRNA可能会以高亲和力结合FOX2，也许可以将FOX1结合。因此，它们可以充当分子“海绵”，以滴定剪接因子，因为在初步实验中，我们发现它们改变了剪接模式。该角度将进行详细检查，并可能直接导致 对Prader-Willi病理学的新见解。我们还将询问某些SNO-LNCRNA是否可以像其他许多LNCRNA一样改变染色质组织。最后，由于这些新的RNA似乎相对稳定，并且严格保留在核中，因此我们将开始利用他们对它们的表达的知识来生成一类新的向量来用于多种序列的核表达。