Dissecting the mechanisms underlying RNA processing and function of non-coding RNAs derived by SINE elements.

剖析 SINE 元件衍生的非编码 RNA 的 RNA 加工机制和功能。

• 批准号: RGPIN-2018-05955
• 负责人: Zovoilis, Athanasios
• 金额: $ 2.26万
• 依托单位: University of Lethbridge
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2022
• 资助国家: 加拿大
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=748500
• 关键词: Dissecting; mechanisms; underlying; RNA; processing

Background: More than 97% of human and mouse genomes do not encode for proteins (non-coding genome) and until recently, a large part of this non-coding genome was referred as "junk DNA" that was believed to have no functional importance. More than 50% of the non-coding genome consists of repetitive elements, with small interspersed nuclear elements (SINEs) making the largest group of these elements. Interestingly, a large number of these SINE elements are transcribed into non-coding RNAs (SINE RNAs). However, the function of these SINE RNAs remains largely unknown. Specific classes of SINE RNAs, such as the B2 and Alu RNAs, have been recently shown to inhibit transcription of specific target genes by binding RNA Polymerase II. In a previous study, we revealed that after application of environmental stimuli, these RNAs are extensively processed and destabilized, which results in their inability to bind and inhibit RNA Polymerase II and thus, it adds an additional layer of regulation of expression of target genes. However, the mechanism of processing of SINE RNAs and the factors regulating this process remain unclear.Objectives: My proposed research program aims to dissect the mechanisms underlying RNA processing of SINE RNAs. It will focus on 1) identifying the factors that trigger this processing and regulate RNA processing rate, 2) identifying and studying the function of the RNA products of the SINE RNA processing in regulation of cellular stress response, including both linear and circular RNA fragments, and 3) investigating how changes in the sequence of SINE RNAs affect their processing, to understand better the sequence requirements for this process.Research Approach: To achieve these aims, my trainees and I will employ and customize a series of RNA genomics and bioinformatics approaches, such as library construction approaches for RNA sequencing and bioinformatics pipelines adapted to the study of repetitive, short and circular RNAs. These approaches include our previously developed short-RNA sequencing for estimation of the SINE RNA processing as well as RNA-sequencing and circRNA-sequencing for identification of linear and circular RNAs, respectively. The proposed research program will also employ RNA biochemistry and cell culture techniques to test in vitro and in cells the interaction between SINE RNAs and a number of proteins that control SINE RNA processing.Impact: This research program will shed light to the function of a part of the non-coding genome previously believed to be "junk DNA". It will allow for a better understanding of the under-estimated until now role of SINE RNAs in response to cellular stress and regulation of gene expression that are critical for cell function.

背景：超过97％的人和小鼠基因组没有编码蛋白质（非编码基因组），直到最近，该非编码基因组的很大一部分被称为“垃圾DNA”，据信被认为没有功能重要性。超过50％的非编码基因组由重复的元素组成，散布的核元素（SIN）是这些元素中最大的群体。有趣的是，这些正弦元素中有很大一部分被转录为非编码RNA（正弦RNA）。但是，这些正弦RNA的功能在很大程度上还不清楚。最近已证明特定类别的正弦RNA（例如B2和Alu RNA）通过结合RNA聚合酶II抑制特定靶基因的转录。在先前的研究中，我们透露，在应用环境刺激后，这些RNA被广泛处理和不稳定，这导致它们无法结合和抑制RNA聚合酶II，因此增加了靶基因表达的附加调节层。但是，正弦RNA的处理机制和调节此过程的因素仍然不清楚：我提出的研究计划旨在剖析正弦RNA的RNA处理的基础机制。它将重点关注1）确定触发该处理并调节RNA处理率的因素，2）识别和研究正弦RNA处理在调节细胞压力响应调控中的RNA产物的功能，包括线性和圆形RNA片段，以及3）研究对我的过程的变化，以使我的过程变得更好，以使我能够实现该过程，以使其能够实现该过程，以使其能够实现该过程，以使其能够实现以下过程，以实现这一方法，以实现以下过程。并定制了一系列RNA基因组学和生物信息学方法，例如用于RNA测序和生物信息学管道的图书馆构建方法，适合于重复，短和圆形RNA的研究。这些方法包括我们先前开发的短RNA测序，用于估计正弦RNA处理，以及分别用于鉴定线性和圆形RNA的RNA测序和circRNA序列。拟议的研究计划还将采用RNA生物化学和细胞培养技术在体外测试和细胞中正弦RNA和许多控制正弦RNA处理的蛋白质之间的相互作用。影响力：该研究计划将阐明以前认为是“ junk DNA”的非编码基因组的一部分功能。到目前为止，它将更好地理解正弦RNA对细胞应激的作用和对细胞功能至关重要的基因表达的调节的作用。