Dissecting genetic determinants of epigenetic instability in pluripotent stem cells

剖析多能干细胞表观遗传不稳定性的遗传决定因素

• 批准号: 10609911
• 负责人: Matthias Stadtfeld
• 金额: $ 39.91万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-15 至 2026-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10609911
• 关键词: Aberrant DNA Methylation; Affect; Alleles; Biological Assay; Biological Markers; Cell Line; Cells; Chromatin; Chromosome 13; Chromosome Mapping; Complex; DNA; DNA Methylation; Degenerative Disorder; Development; Developmental Gene; Diagnostic; Disease; Epigenetic Process; Essential Genes; Event; Exhibits; Future; Gene Cluster; Gene Expression; Generations; Genes; Genetic; Genetic Determinism; Genetic Heterogeneity; Genetic Models; Genetic Variation; Genome; Genome engineering; Genomic Imprinting; Genomics; Goals; Health; Human; Hybrids; Hypermethylation; Impairment; Inbred Mouse; Maintenance; Mammals; Maps; Mediating; Methylation; Modeling; Molecular; Mouse Strains; Mus; Patients; Pluripotent Stem Cells; Population; Predisposition; Process; Protocols documentation; Quantitative Trait Loci; Regulation; Regulator Genes; Reporter; Reporter Genes; Signal Pathway; Somatic Cell; Susceptibility Gene; Syndrome; System; Testing; Therapeutic; Trans-Activators; Transcription Repressor; Transgenic Organisms; Variant; Work; analysis pipeline; causal variant; cell type; epigenetic marker; epigenetic regulation; epigenome; gene product; genetic approach; genetic regulatory protein; genetic variant; genome-wide; genomic locus; human disease; human pluripotent stem cell; imprint; improved; insight; mouse genetics; novel; novel strategies; preservation; prevent; protective allele; rational design; risk prediction; stem cells

Project Summary The remarkable ability of pluripotent stem cells (PSCs) to give rise to a plethora of fully functional somatic cell types becomes progressively impaired upon ex vivo culture by aberrant gain or loss of DNA methylation marks at essential developmental gene loci. This pervasive epigenetic instability represents a significant hurdle for biomedical applications using PSCs, but the underlying molecular reasons remain almost entirely unknown. An important example for epigenetic instability in PSCs is loss of genomic imprinting, an essential gene regulatory mechanism in mammals. By combining transgenic reporter alleles for imprint stability with mouse genetics, we found that imprint dysregulation in PSCs is caused by identifiable genetic variants that affect trans acting chromatin regulators. The goal of this proposal is to develop a comprehensive molecular understanding on how specific genetic variants affect the stability of vital epigenetic marks in PSCs. To work towards this goal, we will first functionally dissect the causal variants within a genomic susceptibility region that we discovered and which drives locus-specific DNA hypermethylation and loss of developmental potential. This will allow us to pinpoint gene regulatory features that predispose imprinted and other genes to aberrant gain of DNA methylation (Aim 1). We will then use genetic mapping in hybrid PSCs to discover quantitative trait loci that can protect against detrimental DNA hypomethylation and validate the function of associated gene products in both mouse and human pluripotent cells (Aim 2). Finally, we will model the genetics of imprint stability in a panel of PSCs with highly diverse yet well-characterized genomes. This will enable us to dissect the molecular regulation of epigenetic stability in complex genetic backgrounds resembling the human population and help us to establish new biomarkers for the identification of culture conditions that preserve developmental potential in PSCs of unknown genetic background (Aim 3). Our work will establish novel conceptual and mechanistic frameworks for the detrimental epigenetic vulnerability of pluripotent cells, which will aid the safe and reliable use of these cells to study developmental and disease processes. In addition, our findings will have relevance for a wide range of human diseases and developmental syndromes characterized by epigenetic instability.

项目摘要 多能干细胞（PSC）具有产生大量功能齐全的体细胞的显著能力， 在离体培养时，由于DNA甲基化标记的异常获得或丧失， 在重要的发育基因位点。这种普遍的表观遗传不稳定性是一个重大障碍， 生物医学应用中使用PSC，但潜在的分子原因仍然几乎完全未知。一个 PSC中表观遗传不稳定性的一个重要例子是基因组印记的丢失，这是一种必需的基因调控因子， 哺乳动物的机制。通过将转基因报告等位基因与小鼠遗传学相结合， 发现PSC中的印记失调是由可识别的遗传变异引起的， 染色质调节因子该提案的目标是发展一个全面的分子理解， 特定的遗传变异如何影响PSC中重要表观遗传标记的稳定性。为了实现这一目标， 我们将首先从功能上剖析我们发现的基因组易感区域内的致病变异， 并导致基因座特异性DNA超甲基化和发育潜能丧失。这将使我们能够 精确定位使印记基因和其他基因易于获得异常DNA的基因调控特征 甲基化（目的1）。然后，我们将在杂交PSC中使用遗传作图来发现数量性状基因座， 可以防止有害的DNA低甲基化，并验证相关基因产物的功能， 小鼠和人多能细胞（Aim 2）。最后，我们将在一个 一组具有高度多样性但特征良好的基因组的PSC。这将使我们能够解剖分子 在类似于人类群体的复杂遗传背景中调节表观遗传稳定性，并帮助我们 建立新的生物标志物，用于鉴定保存发育潜力的培养条件， 遗传背景未知的PSC（目标3）。我们的工作将建立新的概念和机制 多能细胞有害的表观遗传脆弱性的框架，这将有助于安全可靠的 利用这些细胞来研究发育和疾病过程。此外，我们的发现将与 用于以表观遗传不稳定性为特征的广泛的人类疾病和发育综合征。