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

项目总结/摘要 额外染色体（三体或标记染色体）的存在是先天性染色体异常的常见原因。 异常和发育障碍。大约1%的染色体异常是镶嵌性的，并且 这些患者具有两个细胞群，有和没有染色体异常。帕利斯特基利安 综合征（PKS）是一种常见的嵌合染色体疾病，这种综合征是由存在 一种额外的额外标记染色体，等染色体12 p（i（12 p），由两个12 p臂组成， 镜像）。我们以前证明了PKS中的i（12 p）是在减数分裂过程中形成的。所有PKS合子 含有i（12 p），但在有丝分裂过程中，一些细胞通过称为非整倍体拯救的过程失去i（12 p）。 因此，所有PKS患者都表现出镶嵌现象。在嵌合体染色体疾病中， 剂量被认为是临床表型的主要驱动因素，并且没有剂量的拯救细胞被认为是临床表型的主要驱动因素。 染色体异常被认为是“正常的”。基于这一前提，有人提出， 嵌合体比率（具有染色体异常的细胞的比例， 任何给定的组织）和患者临床症状的严重程度，但我们以前证明了缺乏 PKS中的这种相关性。缺乏相关性的一个可能解释是，失去了细胞功能的细胞 额外染色体继续显示功能改变。通过利用马赛克的性质， 在PKS中观察到染色体异常，我们评估非整倍体拯救的PKS细胞是否保留基因表达 和表观遗传异常没有i（12 p），我们发现了类似的转录组和组蛋白修饰 PKS细胞与无额外12 p之间的曲线。因此，我们的数据指向一个持续的“记忆”， i（12 p）对i（12 p）消除后未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治疗方法，以改善临床特征 与PKS和其他儿童染色体数目异常有关。我们的初步数据显示 为了完全恢复PKS的细胞功能，不仅要消除额外的染色体， 需要表观遗传重置。本研究的总体目标是研究i（12 p）在 在PKS中观察到转录和表观遗传改变。为了实现我们的总体目标，我们将 通过两个特定目的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改变 在单细胞水平上使用单细胞RNA测序和嵌合PKS皮肤成纤维细胞系。在目标2中， 将研究诱导多能干细胞重编程是否可以减轻转录组， 在没有i（12 p）的PKS细胞中观察到的表观基因组改变。这个项目有可能推动一个范式转变， 我们对镶嵌染色体疾病的基本病理学的理解。这项工作将具有重大意义 与正在进行的为广泛的染色体疾病开发新疗法的努力相关。

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.