Regulation of X-chromosome inactivation and imprinting by non-coding elements

项目摘要 现在越来越多的人认识到长链非编码RNA和表观遗传复合体之间的相互作用， 发展决策的重要监管者。一个典型的例子是X染色体失活（XCI）， 一个X染色体在早期胚胎中失活的表观遗传过程。XCI可以 随机地或以印记的方式放置在父方X上。在小鼠中，随机XCI观察到在 胚胎固有体（索马），并在胎盘中印迹XCI。两者都需要“X-失活中心”（Xic）， 包含几个必需的非编码基因的X连锁区域。一个关键的基因是Xist，它产生一个 非编码RNA，它“描绘”X染色体并顺式募集沉默因子。我们的工作表明 在随机XCI过程中，Xist受到Xic因子的正向和负向调节，Jpx在Xic因子中。 阳性通路和阴性通路中的Tsix/Xite。我们还证明了同源X- 染色体配对坐标，X染色体将被灭活。在这个融资周期中， 进一步深入了解Xic在X染色体计数、等位基因选择和印记中的作用。 对于计数机制，Jpx RNA --一种Xist激活剂--通过滴定 常染色体“分母”CTCF。这种滴定考虑了X与常染色体（X：A）的比率，并产生了一个 Xist诱导的许可状态。我们还了解到，X-X配对需要一个新的长非编码RNA 称为“PAR-TERRA”，由X连锁的假常染色体区域（PAR）产生。有趣的是， 相互作用发生在两个Xic和两个PAR的“四分体”内。X-X配对然后通过mRNA 脱帽酶DCP 1A。对于印迹XCI，我们已经产生了一个离体细胞模型来研究XP 印记我们已经证明，200 kb的Xic区域足以指导Xist印迹， 印记需要雄性种系中的未配对状态，类似于减数分裂性染色体 灭活（MSCI）。我们的研究产生了新的机会和许多额外的机制 问题.在下一个资金周期，我们的目标是（i）研究X-X配对的机制， X染色体对称性的破坏，（ii）确定Jpx-CTCF滴定如何调节XCI的起始， 阐明印迹XCI的作用机理。拟议的工作将需要五年时间才能完成 预计将为表观遗传学和RNA领域的学术事业培养三名博士后研究员 生物学

PROJECT SUMMARY Interactions between long noncoding RNAs and epigenetic complexes are now increasingly recognized as important regulators of developmental decisions. A classic example is X-chromosome inactivation (XCI), an epigenetic process by which one X-chromosome becomes inactivated in the early embryo. XCI can take place randomly or in an imprinted manner on the paternal X. In the mouse, random XCI is observed in the embryo proper (soma), and imprinted XCI in the placenta. Both require the ‘X-inactivation center’ (Xic), an X-linked region that harbors several essential noncoding genes. A key gene is Xist, which produces a noncoding RNA that ‘paints’ the X-chromosome and recruits silencing factors in cis. Our work has shown that, during random XCI, Xist is regulated both positively and negatively by Xic factors, with Jpx in the positive pathway and Tsix/Xite in the negative pathway. We have also shown that homologous X- chromosome pairing coordinates which X chromosome will be inactivated. In this funding cycle, we have further advanced understanding of the Xic’s role in X-chromosome counting, allelic choice, and imprinting. For the counting mechanism, Jpx RNA — an Xist activator — serves as an X-linked “numerator” by titrating the autosomal “denominator”, CTCF. This titration accounts for the X-to-autosome (X:A) ratio and creates a permissive state for Xist induction. We also learned that X-X pairing requires a new long noncoding RNA called “PAR-TERRA” produced from the X-linked pseudoautosomal region (PAR). Interestingly, all pairing interactions take place within a “tetrad” of two Xic’s and two PAR’s. X-X pairing is then resolved by mRNA decapping enzyme, DCP1A. For imprinted XCI, we have generated an ex vivo cellular model to study XP imprinting. We have demonstrated that a 200-kb Xic region is sufficient to direct Xist imprinting and that Xist imprinting requires an unpaired state in the male germ line that is akin to meiotic sex chromosome inactivation (MSCI). Our research has generated new opportunities and many additional mechanistic questions. In the next funding cycle, we aim to (i) investigate mechanisms underlying X-X pairing and breaking of X-chromosome symmetry, (ii) determine how Jpx-CTCF titration regulates the initiation of XCI, and (iii) elucidate the mechanisms of 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 epigenetics and RNA biology.