Elucidating the effects of extra chromosome elimination in mosaic aneuploidy syndromes: Pallister-Killian syndrome as a model

项目摘要/摘要 额外染色体(三体或标记染色体)的存在是先天性的常见原因。 畸形和发育障碍。大约1%的染色体异常是镶嵌的，并且 这些患者有两个细胞群体，分别有和没有染色体异常。帕利斯特-基利安 综合征(PKS)是一种常见的嵌合体染色体疾病，这种综合征是由于存在 一种额外的额外标记染色体，等染色体12p(i(12p)，由两条12p臂组成，这两条臂是 镜像)。我们先前证明，PKS中的I(12P)是在减数分裂过程中形成的。所有PKS受精卵 含有I(12P)，但在有丝分裂过程中，一些细胞通过一个称为非整倍体拯救的过程失去I(12P)。 因此，所有的PKS患者都表现出嵌合体。在马赛克染色体疾病中，改变的基因 剂量被认为是临床表型的主要驱动因素，而被拯救的细胞没有 染色体异常被认为是“正常的”。基于这一前提，有人提出， 马赛克率(出现染色体异常的细胞比例)与 任何给定的组织)和患者临床症状的严重程度，但我们先前证明缺乏 在PKS中存在这样的相关性。缺乏相关性的一个可能的解释是，丢失了 额外的染色体继续显示出功能改变。通过利用马赛克的性质， 在PKS中发现的染色体异常，我们评估了非整倍体挽救的PKS细胞是否保留了基因表达 和没有I(12P)的表观遗传学异常，我们发现了类似的转录组和组蛋白修饰 有无额外12P的PKS细胞的分布。因此，我们的数据指向一个持续的“记忆” I(12P)对消除I(12P)后不能恢复的转录组和表观基因组的影响 通过非整倍体拯救。该项目的长期目标是确定改善临床特征的治疗方法。 与儿童的PKS和其他数字染色体疾病有关。我们的初步数据显示 为了完全恢复PKS的细胞功能，不仅消除了额外的染色体，而且还 表观遗传重置是必要的。本研究的总体目标是探讨I(12P)在大脑皮层中的作用。 在PKS中观察到转录和表观遗传学的变化。为实现我们的总体目标，我们将 通过两个具体目标进行进一步的基因组分析。在目标1中，我们将研究基因表达的变化 在单细胞水平采用单细胞RNA测序和马赛克PKS皮肤成纤维细胞系。在目标2中，我们 将研究诱导多能干细胞重编程是否可以减轻转录和 在无I(12P)的PKS细胞中可见表观基因组改变。该项目有可能推动 我们对马赛克染色体疾病的基本病理生物学的理解。这项工作将具有重要的意义 与为广泛的染色体疾病开发新疗法的持续努力具有相关性。

Project Summary/Abstract The presence of an extra chromosome (trisomy or marker chromosome) is a common cause of congenital anomalies and developmental disabilities. Approximately 1% of chromosomal abnormalities are mosaic, and these patients have two cellular populations with and without chromosomal abnormalities. Pallister-Killian syndrome (PKS) is a common mosaic chromosomal disorder, and this syndrome is caused by the presence of an extra supernumerary marker chromosome, isochromosome 12p (i(12p), composed of two 12p arms that are mirror-images). We previously demonstrated that the i(12p) in PKS is formed during meiosis. All PKS zygotes contain i(12p), but during mitosis, some cells lose the i(12p) through a process known as aneuploid rescue. Consequently, all patients with PKS demonstrate mosaicism. In mosaic chromosomal disorders, altered gene dosage is believed to be the primary driver of the clinical phenotype, and the rescued cells without the chromosome abnormality are thought to be “normal.” Based on this premise, it has been proposed that there exists a correlation between the mosaic ratio (proportion of the cells possessing chromosomal abnormality in any given tissue) and the severity of patient’s clinical symptoms, but we previously demonstrated the lack of such a correlation in PKS. One possible explanation for the lack of correlation is that cells that have lost the extra chromosome continue to demonstrate functional alterations. By taking advantage of the mosaic nature of chromosomal abnormality seen in PKS, we assessed if aneuploid-rescued PKS cells retain gene expression and epigenetic abnormalities without i(12p), and we discovered similar transcriptome and histone modification profiles between PKS cells with and without the extra 12p. Hence, our data points to an ongoing “memory” effect of the i(12p) on both the transcriptome and epigenome that are not restored after the elimination of i(12p) by aneuploid rescue. The long-term goal of this project is to identify treatments to improve the clinical features associated with PKS and other numerical chromosomal disorders in children. Our preliminary data suggest that, for the full restoration of cellular functions in PKS, not only elimination of the extra chromosome but also epigenetic resetting is needed. The overall objective of this study is to investigate the role of i(12p) in the transcriptional and epigenetic alterations observed in PKS. To achieve our overall objective, we will perform further genomic analyses through two Specific Aims. In Aim 1, we will investigate gene expression alterations at the single-cell level using single-cell RNA-sequencing and mosaic PKS skin fibroblast cell lines. In Aim 2, we will investigate whether induced pluripotent stem cell reprogramming can alleviate transcriptome and epigenome alterations seen in PKS cells without i(12p). This project has a potential to drive a paradigm shift in our understanding of the basic pathobiology of mosaic chromosomal disorders. This work will have significant relevance for ongoing efforts to develop novel therapies for a wide range of chromosomal disorders.