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

描述（由申请人提供）：过去几年的研究反复强调了长链非编码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rna机制的理解是至关重要的，特别是考虑到70-80%的哺乳动物基因组是转录的，而大部分转录是非编码的。在雌性哺乳动物中，lncrna的丰度在x -失活中心（Xic）最为明显，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17kb的Xist RNA发起的，它将Polycomb蛋白招募到x染色体上。Xist RNA反过来由反义转录物t6控制，t6通过排斥含有Polycomb蛋白的RNA-蛋白复合物的招募来对抗Xist。我的实验室还发现，Xist的上调需要另外两个lncrna的作用，RepA将Polycomb蛋白招募到Xic， Jpx则需要激活Xist转录。在过去的13年里，这项RO1拨款为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进一步加深我们对RNA调控的理解。具体来说，我们将：(i)研究rna是否以及如何在同源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如何激活Xist RNA；（iii）阐明随机XCI和印迹XCI之间的根本差异，我们的研究表明，XCI在沉默方面依赖于Xist RNA的差异。这项工作将需要五年时间才能完成，预计将培养五名博士后，从事表观遗传学、RNA调控和干细胞生物学等领域的学术工作。

DESCRIPTION (provided by applicant): 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 turned 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. This RO1 grant has, for the past 13 years, provided crucial support for this lncRNA research. Herein, we propose to further 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 whether and how RNAs play a role in homologous X-chromosome pairing, a process proposed to regulate counting and choosing of X-chromosomes for inactivation; (ii) Determine how Xist RNA is turned on by Jpx RNA; and (iii) Elucidate fundamental differences between random and imprinted XCI, which our research has shown to be differentially dependent on Xist RNA for silencing. That the proposed work will require five years to complete and is expected to train five postdoctoral fellows for academic careers in the fields of epigenetics, RNA regulation, and stem cell biology.