Regulation of X-chromosome Inactivation and Imprinting by Non-coding Elements

项目总结（见说明）： 过去几年的研究一再强调长链非编码RNA（lncRNA）在表观遗传调控、发育和疾病中的重要性。然而，RNA控制这些过程的机制仍然不清楚。提高对基于RNA的机制的理解是至关重要的，特别是考虑到哺乳动物基因组的70-80%是转录的，并且大部分转录是非编码的。在雌性哺乳动物中，lncRNA的丰度最明显的是X-失活中心（Xic），这是一个控制X染色体失活（XCI）的X连锁区域。XCI是研究lncRNA调控的一个很好的模型，因为这个过程是由一系列基于RNA的开关控制的。沉默是由17 kb Xist RNA启动的，因为它将Polycomb蛋白募集到X染色体上。Xist RNA反过来由反义转录物Tsix控制，Tsix通过排斥含有Polycomb蛋白的RNA-蛋白质复合物的募集来拮抗Xist。我的实验室还发现，Xist上调需要另外两种lncRNA的作用，RepA将Polycomb蛋白招募到Xic，而Jpx则需要激活Xist转录。在过去的16年里，GM 58839基金为这项lncRNA研究提供了至关重要的支持。在此，我们建议通过研究Xic的RNA如何控制XCI的不同步骤和形式来提高我们对RNA调控的理解。具体而言，我们将：（i）研究lncRNA如何在同源X染色体配对中发挥作用，这是一个调节X染色体计数和选择失活的过程;（ii）确定Jpx RNA激活Xist表达的机制;（iii）阐明随机和印迹XCI之间的根本差异。拟议的工作将需要五年时间才能完成，预计将培养三名博士后研究员，从事表观基因组学，RNA调控和干细胞生物学领域的学术事业。

PROJECT SUMMARY (See instructions): Studies from the last several years have repeatedly highlighted the importance of long noncoding RNAs (lncRNA) in epigenetic regulation, development, and disease. However, the mechanisms by which RNAs control these processes remain poorly defined. Improved understanding of RNA-based mechanisms is crucial, especially given that 70-80% of the mammalian genome is transcribed and that the vast bulk of transcription is noncoding. Nowhere is the abundance of lncRNAs more evident than at the X-inactivation center (Xic), an X-linked region that controls the X-chromosome inactivation (XCI) in the female mammal. XCI serves as an excellent model to study lncRNA regulation because this process is controlled by a series of RNA-based switches. Silencing is initiated by the 17-kb Xist RNA as it recruits Polycomb proteins to the X-chromosome. Xist RNA is in turn controlled by an antisense transcript, Tsix, which antagonizes Xist by repelling the recruitment of an RNA-protein complex containing Polycomb proteins. My laboratory has also found that Xist upregulation requires the action of two additional lncRNAs, RepA which recruits Polycomb proteins to the Xic, and Jpx which is required to activate Xist transcription. The GM58839 grant has, for the past 16 years, provided crucial support for this lncRNA research. Herein, we propose to advance our understanding of RNA regulation by studying how RNAs of the Xic control the different steps and forms of XCI. Specifically, we will: (i) Investigate how lncRNAs play a role in homologous X-chromosome pairing, a process proposed to regulate counting and choosing of X-chromosomes for inactivation; (ii) Determine the mechanism by which Jpx RNA activates Xist expression; and (iii) Elucidate fundamental differences between random and imprinted XCI. The proposed work will require five years to complete and is expected to train three postdoctoral fellows for academic careers in the fields of epigenomics, RNA regulation, and stem cell biology.