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: 10133457
• 负责人: Anna Afasizheva
• 金额: $ 3.78万
• 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/10133457
• 关键词: 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; three dimensional structure; 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.

项目总结/摘要 该提案的目标是定义长非编码RNA（lncRNA）XACT（X-active）的作用。 包被转录物）。XACT是一种灵长类特异性和X染色体连锁的lncRNA，仅在 多能干细胞在幼稚的人类胚胎干细胞（hESC）中，它模拟了胚胎干细胞的多能状态， 在植入前胚胎中，XACT RNA在雌性细胞中形成两个云，在雄性细胞中形成一个云。 各自的X染色体区域。在发育的这个阶段，雌性和雄性细胞表达X连锁的 尽管两个女性X染色体都是转录活跃的，但基因水平相当。该基因 剂量补偿机制，称为X染色体阻尼（XCD），是人类特有的 发展在植入后，雌性多能细胞通过以下方式切换到一条X染色体的完全沉默： X染色体失活（XCI），一个由研究充分的染色质相关lncRNA XIST介导的过程。 在捕获植入后胚胎的多能状态的致敏hESC中，XACT表达被激活。 在雌性细胞的非活性X染色体（Xi）上沉默，但在雌性细胞中保留在活性X染色体上 男性细胞考虑到许多lncRNA作为转录调节因子发挥作用，我们假设XACT 调节人类早期发育中的幼稚和启动多能状态，可能是通过调节X- 染色体剂量补偿有趣的是，我们实验室的初步发现表明，XIST还 在幼稚多能细胞中控制X染色体阻尼，但在这种细胞中不诱导完全沉默。 发展国家。XACT的一个可能的功能可能是抵消XIST在幼稚细胞中的沉默能力。 在多能细胞中，XCD的特征在于同时表达来自X-细胞的XACT和XIST。 染色体我们证实XACT通过竞争染色质来拮抗XCD过程中XIST的活性 因此，在植入之前防止过早的XCI。揭示XACT在多能状态中的作用 在X染色体剂量补偿的调控中，我们将：1）在幼稚和致敏中删除XACT hESC并分析对细胞状态和转录组的影响;和2）确定其作用的分子机制 作为染色质相关的lncRNA。总之，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 在人类早期发育中XACT功能和XIST调节之间，我们可能会发现一种新的， 灵长类动物特异性的基因剂量补偿策略，并定义了人类多能性的基本特征。