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

描述（由申请方提供）：在雌性哺乳动物中，两条X染色体中的一条失活，以确保雄性和雌性具有相等的X染色体剂量。X染色体失活（XCI）可以随机发生，也可以在父体X（XP）上以印记方式发生。在小鼠中，在胚胎本体（索马）中观察到随机XCI，并在胎盘中印迹XCI。体细胞和胎盘XCI都需要“X失活中心”（Xic），这是一个已知的X连锁区域，用于制造非编码RNA（ncRNA）的基因。Xic产生Xist RNA，这是一种大的非翻译RNA，它“描绘”X并顺式招募沉默因子到X。我们的实验室以前确定的反义Tsix基因和第二个非编码元件，Xite，作为Xist的基本阻遏物。在上一个资助周期中，我们重点关注了Tsix和Xite调节XCI不同步骤的分子手段。这项研究导致了三个主要发现：（i）在女性中，XCI发作时，X短暂地彼此“配对”。我们认为配对介导了X之间的“串扰”，以确保相互排斥的染色体命运。Tsix和Xite介导配对相互作用。(ii)Tsix通过其反义转录驱动的染色质结构的抢先变化来控制Xist表达。Tsix RNA还结合Dnmt3a并指导Xist启动子的从头DNA甲基化。(iii)剂量补偿在发育过程中比以前认为的要早得多。我们相信印记XCI在受孕时就已经发生，并在此基础上提出XP是作为来自父系种系的“预失活”染色体遗传的。这些发现提出了许多新的问题，我们现在打算在三个不同的目标中进行研究。在目标1中，我们将定义X染色体配对所需的分子因子。在目标2中，我们打算研究Tsix转录如何指导染色质修饰以及Tsix RNA招募了哪些染色质修饰剂。在目标3中，我们将进行遗传和生化测试，以区分预灭活和从头模型的印记XCI。从研究项目中获得的信息将对人类遗传学，癌症和表观遗传重编程产生广泛的影响。它还将增强我们对基本生物学过程的理解，特别是那些涉及染色体动力学，ncRNA调控和跨代遗传的过程。

DESCRIPTION (provided by applicant): In the female mammal, one of two X-chromosomes is inactivated to ensure that males and females have equal X-chromosome dosage. X-chromosome inactivation (XCI) can take place randomly or in an imprinted manner on the paternal X (XP). In the mouse, random XCI is observed in the embryo proper (soma) and imprinted XCI in the placenta. Both somatic and placental XCI require the 'X-inactivation center' (Xic), an X-linked region known for genes making noncoding RNA (ncRNA). The Xic produces Xist RNA, a large untranslated RNA that 'paints' the X and recruits silencing factors to the X in cis. Our laboratory previously identified the antisense Tsix gene and a second noncoding element, Xite, as essential repressors of Xist. In the previous funding cycle, we focused on the molecular means by which Tsix and Xite regulate different steps of XCI. This investigation has resulted in three major discoveries: (i) In females, the Xs 'pair' transiently with each other at the onset of XCI. We believe that pairing mediates 'cross-talk' between the Xs to ensure mutually exclusive chromosome fates. Tsix and Xite mediate the pairing interaction. (ii) Tsix controls Xist expression through preemptive changes in chromatin structure driven by its antisense transcription. Tsix RNA also binds Dnmt3a and directs de novo DNA methylation to the Xist promoter. (iii) Dosage compensation occurs much earlier in development than previously thought. We believe that imprinted XCI has already taken place by the time of conception and, on this basis, propose that the XP is inherited as a 'pre-inactivated' chromosome from the paternal germline. These discoveries have raised many new questions, which we now intend to pursue in three distinct Aims. In Aim 1, we will define molecular factors required for X-chromosome pairing. In Aim 2, we intend to investigate how chromatin modifications are directed by Tsix transcription and what chromatin modifiers are recruited by Tsix RNA. In Aim 3, we will carry out genetic and biochemical tests to distinguish between the pre-inactivation and de novo models for imprinted XCI. Information gained from the research program will have broad implications for human genetics, cancer, and epigenetic reprogramming. It will also enhance our understanding of basic biological processes, especially those involving chromosome dynamics, ncRNA regulation, and transgenerational inheritance.