健康与遗传心理学研究室知识库

单基因遗传病是指受一对等位基因控制的遗传病，遗传方式符合孟德尔遗传定律，所以又称为孟德尔疾病，呈现出明显的家族传递现象。在单基因疾病的大范畴中，累及神经系统的单基因疾病称为神经系统单基因遗传病。神经系统单基因疾病临床需求和科研地位很重要，迄今为止己经确定的6000多种单基因遗传病，与神经、精神系统相关的遗传病占一半以上。神经系统单基因疾病种类繁多，表型多样，潜在的生物学致病机制均较为复杂，临床上各种疾病之间症状常重叠，具有家族性和终身性的特点，目前为止仍有不少神经系统单基因病的病因和发病机制尚未阐明，给临床诊断和治疗带来很大的挑战。在这一背景下，本研究试图对一名临床高度疑似神经系统单基因疾病的病例，使用全外显子测序技术鉴定其致病突变，并基于功能获得策略通过细胞模型的方法探讨突变影响的生物学功能，最终总结了一套可以应用于神经系统单基因遗传疾病的研究策略。

本研究包括两部分内容:1)利用全外显子测序鉴定神经系统单基因疾病致病基因。本研究通过样本收集，全外显子测序，测序下机数据的分析，致病性候选突变的筛选，遗传学验证等过程得到疑似的致病位点。2)致病突变的生物学功能验证。本研究构建致病位点错义突变的细胞模型，通过瞬时转染的方式转染到人胚胎肾细胞293细胞系，使用转录组测序技术对转录组数据进行分析，然后对基因表达量进行加权基因共表达网络分析得到不同组别的基因共表达模块，即表达变化类似、功能相关的一些基因，最后对感兴趣的基因模块进行生物学功能注释。

本研究主要结果如下。1)本研究通过在一个具有常染色体隐性遗传模式的家系中，利用全外显子测序的方法鉴定到了先证者的疑似的致病位点ARSANM_ 000487.S:c.925G>Ap.E309K，该位点位于与异染性脑白质营养不良疾病相关的ARSA基因上，经过文献回顾发现，在以前的研究中，ARSA c.925G(c.925G>A, c.925G>T, c.925G>C)都被报道过可导致晚婴期异染性脑白质营养不良。2)基于该致病位点缺乏足够的生物学功能研究的现状，本研究通过构建突变细胞模型，使用系统生物学的方法如关键基因和蛋白一蛋白互作网络分析了四种ARSA过表达细胞模型(c.925G,c.925G> A, c.925G> T, c.925G> C)的转录组，结果表明:c.925G处的突变引起与能量代谢，离子结合，囊泡转运和转运相关的分子变化。最后，整合研究一与研究二的研究方法，我们提出了一套鉴定神经系统单基因病致病位点及分析其功能的策略，该策略结合了细胞生物学与生物信息学的方法，能够更加全面地揭示突变影响的生物学功能以及疾病的发病机制。

对神经系统单基因遗传病致病位点的鉴定与功能研究，可以有效鉴定出单基因遗传病致病位点并揭示其影响的生物学功能，为疾病的诊治及其分子机制的研究提供线索。

Monogenetic disease refers to a genetic disease controlled by a pair of alleles. The mode of inheritance conforms to Mendel's law of inheritance, so it is also known as Mendel's disease, showing obvious familial transmission. In the big category of single gene disease, the single gene disease involving the nervous system is called the single gene disease of the nervous system. The clinical needs and scientific research status of single-gene diseases of the nervous system are very important. So far, more than 6,000 single-gene genetic diseases have been identified, of which more than half are related to the nervous and mental systems. Single gene of nervous system diseases and more rare, highly genetic heterogeneity and clinical heterogeneity, the etiology and pathogenesis of many diseases has not been clarified, and has the characteristics of familial and permanent, death, disability, birth defects and to fool rates are very high, treatment difficulty, harm, posed great challenges to the clinical diagnosis and treatment. In this background, this study attempts to identify the pathogenic sites of a clinical highly suspected case of single gene of nervous system diseases by using whole exon sequencing technology, and to explore the biological functions of the sites, and a set of research strategies that can be applied to single gene genetic diseases of nervous system are summarized.

This study consists of two parts: 1) identify the pathogenic genes of single-gene diseases of the nervous system by whole exon sequencing. In this study, suspected pathogenic sites were obtained through sample collection, whole exon sequencing, analysis of sequencing data, screening of pathogenic candidate mutations, genetic verification and other processes.2) biological function verification of pathogenic mutations. In this stu街，one gene overexpression vector and three mutated vectors were constructed and transfected into the HEK293 cell line by transient transfection. The transcriptome data were analyzed by RNA-seq technology. According to the gene expression amount, WGCNA was used to carry out the weighted gene co-expression network analysis to obtain the gene co-expression modules of different groups, that is,some genes with similar expression changes and related functions. Finally, the gene modules of interest were annotated with biological functions.

The results of this study are as follows (1) in this study through in A family with autosomal recessive inheritance patterns, the method of using the fully sequenced exons risk loci identified in the proband of suspected ARSA NM_ 000487. 5: c. 925 g>A p. E 309 k, the site is located in disease and metachromatic leukodystrophy (MLD) ARSA genetically related, through literature review, found that in the previous study, ARSA c. 925 g (c. 925 g>A, c. 925 g>T,c.925G> c) has been reported to cause late infancy MLD. 2) based on the current lack of sufficient biological function studies on this pathogenic site, this study constructed cell models by mutation, and analyzed the transcriptomes of four ARSA-overexpressed cell models (c.925G,c.925G> A,c.925G> T,c.925G> c) through RNA-seq, WGCNA, Hub genes and PPI networks. c. 925 g mutation and energy metabolism, ion, vesicle transport: and related molecular changes. Finally, by integrating the research methods of study 1 and Study 2, we propose a complete set of strategies for identifying pathogenic mutations and functional analysis, which combine cell biology experiments with bioinformatics methods to reveal more comprehensively the biological function of mutations' effects and the pathogenesis of diseases.

The identification and functional study of the pathogenic loci of single-gene hereditary diseases in the nervous system can effectively identify the pathogenic loci of single-gene hereditary diseases and reveal their biological functions, providing clues for the study of the molecular mechanism of diseases.