Deciphering the role of XACT lncRNA in human development

解读 XACT lncRNA 在人类发育中的作用

• 批准号: 10460910
• 负责人: Anna Afasizheva
• 金额: $ 5.18万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10460910
• 关键词: 3-Dimensional; Address; Affect; Cells; Chromatin; Chromosome Territory; DNA sequencing; Development; Dosage Compensation (Genetics); Epigenetic Process; Female; Gene Deletion; Gene Dosage; Gene Expression; Gene Expression Regulation; Gene Silencing; Genes; Genetic Transcription; Genomic Segment; Goals; Human; Human Development; Image; Interference Microscopy; Link; Mediating; Microscopy; Modeling; Molecular Mechanisms of Action; Mus; Nuclear; Pattern; Phenotype; Pluripotent Stem Cells; Primates; Process; RNA; Regulation; Resolution; Role; System; Testing; Transcript; Untranslated RNA; X Chromosome; X Inactivation; base; blastocyst; density; experimental study; genome-wide; genomic locus; human embryonic stem cell; human female; implantation; knockout gene; male; natural Blastocyst Implantation; novel; pluripotency; preimplantation; premature; prevent; programs; transcriptome; transcriptome sequencing

Project Summary / Abstract The goal of this proposal is to define the role of the long non-coding RNA (lncRNA) XACT (X-active coating transcript). XACT is a primate-specific and X-chromosome-linked lncRNA expressed exclusively in pluripotent stem cells. In naïve human embryonic stem cells (hESCs), which model the pluripotent state of the pre-implantation embryo, the XACT RNA forms two clouds in female and one cloud in male cells over the respective X-chromosome territories. At this stage of development, female and male cells express X-linked genes at comparable levels despite both female X-chromosomes being transcriptionally active. This gene dosage compensation mechanism, known as X-chromosome dampening (XCD), is unique to human development. Upon implantation, female pluripotent cells switch to complete silencing of one X-chromosome via X-chromosome inactivation (XCI), a process mediated by the well-studied chromatin-associated lncRNA XIST. In primed hESCs, which capture the pluripotent state of the post-implantation embryo, XACT expression is silenced on the inactive X-chromosome (Xi) of female cells, but is retained on the active X-chromosome in female and male cells. Considering that many lncRNAs function as transcriptional regulators, we hypothesize that XACT modulates the naïve and primed pluripotent states in early human development, potentially by regulating X- chromosome dosage compensation. Intriguingly, preliminary findings in our lab have shown that XIST also controls X-chromosome dampening in naïve pluripotent cells, but does not induce complete silencing in this developmental state. One possible function of XACT may be to counteract the silencing capacity of XIST in naïve pluripotent cells, in which XCD is characterized by concurrent expression of XACT and XIST from the X- chromosome. We posit that XACT antagonizes the activity of XIST during XCD through competitive chromatin association, thus preventing premature XCI prior to implantation. To unveil the role of XACT in pluripotent states and in the regulation of X-chromosome dosage compensation, we will: 1) delete XACT in naïve and primed hESCs and analyze the effect on cell states and transcriptomes; and 2) define its molecular mechanism of action as a chromatin-associated lncRNA. Taken together, these approaches will elucidate the contribution of XACT to the control of pluripotent states and its role in X-chromosome gene dosage regulation. By exploring the link between XACT function and the modulation of XIST in early human development, we may uncover a novel, primate-specific strategy for gene dosage compensation and define fundamental features of human pluripotency.

项目摘要 /摘要 该提案的目的是定义长非编码RNA（lncRNA）XACT的作用（X-Active 涂料成绩单）。 XACT是一种独家表达的私人特异性和X-chromosoms-LncRNA 多能干细胞。在幼稚的人类胚胎干细胞（hESC）中，该干细胞对多能状态进行了模拟 植入前的胚胎，XACT RNA在女性中形成两个云，在雄性细胞中形成一个云 主要的X染色体领土。在发育阶段，雌性和男性细胞表达X连锁 在可比水平的基因目的地，两个雌性X染色体都具有转录活性。这个基因 剂量补偿机制，称为X染色体减弱（XCD），是人类独有的 发展。植入后，雌性多能细胞切换到一个X染色体的沉默 X染色体灭活（XCI），这是由研究良好的染色质相关的lncRNA xist介导的过程。 在捕获植入后胚胎的多能状态的液压hESC中，xact表达是 在雌性细胞的无活性X染色体（XI）上沉默，但保留在女性的活性X-chromosom上 和雄性细胞。考虑到许多LNCRNA充当转录调节剂，我们假设XACT 在早期人类发展中调节幼稚和启动的多能状态，可能通过调节x- 染色体剂量补偿。有趣的是，我们实验室中的初步发现也表明了xist 控制幼稚的多能细胞中X染色体抑制作用，但不会引起完全沉默 发展状态。 XACT的一种可能功能可能是抵消Xist的沉默能力 多能细胞，其中XCD的特征是X-cACT和XIST的同时表达。 染色体。我们指出，XACT通过竞争性染色质在XCD过程中拮抗XIST的活性 关联，从而防止植入之前过早XCI。揭示XACT在多能状态中的作用 在X染色体剂量补偿的调节中，我们将：1）在幼稚中删除XACT hESC并分析对细胞状态和转录组的影响； 2）定义其分子作用机理 作为染色质相关的lncRNA。综上所述，这些方法将阐明XACT对 多能状态的控制及其在X染色体基因剂量调节中的作用。通过探索链接 在XACT功能和XIST在早期人类发展中的调制之间，我们可能会发现一个小说， 基因剂量补偿的灵长类动物特异性策略并定义了人类多能的基本特征。