Understanding the complexity of gene dosage imbalance in Down syndrome

摘要 唐氏综合征（DS，21三体，T21）患者表现出一系列的临床表型， 21号染色体额外拷贝（HSA 21）。临床范围包括智力残疾，早发性 老年痴呆症、先天性心脏病和血液异常，包括儿童白血病。DS 表型可能与HSA 21额外拷贝引起的基因表达改变有关， 了解导致不同表型的基因组决定因素是DS的一个主要目标 research.我们的努力将解决几个现有的挑战，其中包括：小鼠模型不重复 所有人DS表型，ii.小鼠和人DS细胞中HSA 21基因剂量的操纵 模型依赖于以非生理剂量改变表达的方法，和iii.转录组学研究 正常和T21组织之间的差异受到来自固有基因表达的相当大的“噪音”的挑战， 个体之间的差异。在这个应用程序中，我们将通过使用创新的 创造、遗传操作和分析患者来源的诱导多能干细胞的技术 （iPSC）。我们将使用一组确定的T21 iPSC系和CRISPR-CAS9基因编辑来产生等基因细胞， 仅在HSA 21或特定候选基因的拷贝数上不同的品系。使用同基因T21和整倍体 我们将测试T21是否会破坏整个染色体结构，导致全基因组基因突变。 表达失调。我们将在基因表达，染色体结构，DNA 甲基化和染色质签名，以提供全基因组转录组的比较视图， 在同基因T21和整倍体细胞中染色质接触。进一步的研究将检查异常是否 由于一种或多种特异性HSA 21基因的过表达，或由于杂合性 在患有白血病的DS患者中鉴定出CTCF或内聚成分的突变。我们的目标是生产 新的，医学相关的知识，促进我们对DS基因剂量失衡的理解， 对DS相关白血病和潜在的新治疗方法特别感兴趣。我们的目标不仅是提供洞察力 但可能对与非整倍体相关的其他疾病有更广泛的影响。

ABSTRACT Patients with Down syndrome (DS, trisomy 21, T21) demonstrate a spectrum of clinical phenotypes due to an extra copy of chromosome 21 (HSA21). The clinical spectrum encompasses intellectual disability, early onset Alzheimer’s disease, congenital heart defects, and blood abnormalities including childhood leukemia. DS phenotypes are likely related to alteration of gene expression due to the extra copy of HSA21, and understanding the genomic determinants that contribute to the different phenotypes is a major objective in DS research. Our efforts will address several existing challenges that include: i. murine models do not recapitulate all of the human DS phenotypes, ii. manipulation of HSA21 gene dosage in murine and human DS cellular models have relied on methods that alter expression in non-physiologic doses, and iii. transcriptomic studies between normal and T21 tissues are challenged by considerable “noise” from inherent gene expression variation among individuals. In this application, we will address these challenges by using innovative technologies to create, genetically manipulate, and analyze patient-derived induced pluripotent stem cells (iPSCs). We will use a defined set of T21 iPSC lines and CRISPR-CAS9 gene editing to produce isogenic cell lines that differ only by copy number of HSA21 or specific candidate genes. Using isogenic T21 and euploid iPSCs, we will test whether T21 disrupts overall chromosomal architecture that results in genome-wide gene expression dysregulation. We will perform studies in gene expression, chromosome architecture, DNA methylation, and chromatin signatures to provide a comparative view of genome-wide transcriptome and chromatin contacts in isogenic T21 and euploid cells. Further studies will examine whether abnormalities are due to the overexpression of one or more specific HSA21 genes, or as a consequence of heterozygous mutations in CTCF or cohesion components identified in DS patients with leukemia. Our goal is to produce novel, medically relevant knowledge that advances our understanding of gene dosage imbalance in DS, with a particular interest in DS-associated leukemia and potential new treatments. We aim to not only provide insight into DS abnormalities, but may have broader implications for other diseases associated with aneuploidy.