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Functional analysis of a non-coding RNA in the mammalian circadian clock system

哺乳动物生物钟系统中非编码RNA的功能分析

基本信息

批准号：
10488206
负责人：
Shihoko Kojima
金额：
$ 32.47万
依托单位：
VIRGINIA POLYTECHNIC INST AND ST UNIV
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-09-01 至 2024-08-31
项目状态：
已结题

项目摘要

Circadian rhythmicity is a fundamental aspect of the temporal organization of cells in the body, and it modulates much of biochemistry, physiology, behavior, and pathological state. Essentially every cell in the body is capable of generating circadian rhythmicity in mammals, and within each cell, a set of clock genes (which are highly conserved among animals) form transcription–translation feedback loops that drive circadian oscillation. Even though the molecular basis of circadian rhythmicity is thought to be well-characterized, recent transcriptome analyses identified a novel transcript that appears to play an interesting role in these core clock loops. This transcript, Per2AS, is a natural antisense transcript, a class of non-coding RNAs (ncRNAs), of Period2 (Per2), one of the core clock genes. Per2AS is transcribed from the strand opposite to Per2 and its expression is rhythmic and antiphasic to Per2. Given that Per2AS expression is rhythmic and antiphasic to its sense partner and that rhythmic transcription is more energy-consuming than non-rhythmic transcription, it is hypothesized that Per2AS is a functional molecule and plays an important role in the mammalian circadian clock system. Even though ncRNAs were originally considered to be mere transcriptional noise and lack defined functions, a few dozen examples have expanded the scope of ncRNAs from mere “junk” to functional molecules having a wide spectrum of regulatory roles. In fact, antisense transcripts of a core clock gene have been reported in Neurospora crassa and Antheraea pernyi and been shown to confer robust and sustained rhythmicity, implying that sense-antisense interactions of a core clock gene constitute a common mechanism for circadian clock regulation across kingdoms. Three specific aims have been designed to test our central hypothesis and define the biological function of Per2AS in the mammalian circadian clock system. The first aim addresses whether the presence, rhythmicity, and phase of Per2AS expression relative to Per2 are biologically significant. The second aim takes advantage of traditional strategies and directly asks whether perturbations of Per2AS expression result in changes in the circadian clock machinery. The third aim, based on our preliminary data, focuses on specific molecules that Per2AS may regulate in order to shed light on the molecular function(s) of Per2AS. Successful completion of this study not only advances our knowledge of circadian biology but also of regulatory ncRNAs. Only in relatively few cases have interactions between sense and antisense RNA pairs been explored and the physiological importance and mode-of-action of these pairs remain poorly understood. Outcomes from the proposed project will have significant impact in understanding the role of antisense transcripts and shed light on the molecular mechanisms by which antisense transcripts elicit physiological functions without producing a protein. Deepening the understanding of the clock oscillatory mechanism and how a non-coding gene (i.e., Per2AS) contributes to the clock output will help us to, ultimately, develop therapeutics for people who suffer from disrupted internal clocks.

昼夜节律是体内细胞时间组织的基本方面，它调节许多生物化学、生理学、行为和病理状态。基本上，在哺乳动物中，身体能够产生昼夜节律，在每个细胞中，一组时钟基因（在动物中高度保守）形成转录-翻译反馈环，振荡尽管昼夜节律的分子基础被认为是很好的特征，但最近的研究表明，转录组分析鉴定了一种新的转录本，它似乎在这些核心时钟中起着有趣的作用循环这种转录本Per 2AS是一种天然反义转录本，是一类非编码RNA（ncRNA）， Period 2（Per 2）是一个核心时钟基因。Per 2AS从与Per 2相反的链转录，其表达与Per 2有节律且反相。鉴于Per 2AS表达是有节奏的，并且与其表达相反。有节奏的转录比无节奏的转录更消耗能量，假设Per 2AS是一种功能性分子，在哺乳动物的昼夜节律中起着重要作用时钟系统尽管ncRNA最初被认为仅仅是转录噪音，虽然已经定义了功能，但几十个例子已经将ncRNA的范围从仅仅是“垃圾”扩展到了功能性的。具有广谱调节作用的分子。事实上，核心时钟基因的反义转录本在粗糙脉孢菌和柞蚕中有报道，节律性，这意味着核心时钟基因的正反义相互作用构成了一个共同的机制生物钟调节的重要性设计了三个具体目标来测试我们的核心目标假设和定义Per 2AS在哺乳动物生物钟系统中的生物学功能。第一个目标解决了Per 2AS表达相对于Per 2的存在、节律性和时相是否在生物学上是显著第二个目标利用传统的战略，直接问是否扰动的 Per 2AS表达导致生物钟机制的变化。第三个目标，根据我们初步的数据，侧重于Per 2AS可能调节的特定分子，以揭示分子 Per 2AS的功能。这项研究的成功完成不仅推进了我们对昼夜节律的了解，生物学和调控性ncRNA的研究。只有在相对较少的情况下，反义RNA对已被探索，这些对的生理重要性和作用模式仍然存在不太了解。拟议项目的成果将对理解这一作用产生重大影响的反义转录本和阐明的分子机制，反义转录本引起不产生蛋白质的生理功能。深化对时钟振荡的认识机制以及非编码基因（即，Per 2AS）对时钟输出的贡献将帮助我们，最终，为生物钟紊乱的人开发治疗方法。