Investigation of DUF1220 domains in human brain function and disease

DUF1220 结构域在人脑功能和疾病中的研究

基本信息

批准号：
9313332
负责人：
JAMES M SIKELA
金额：
$ 38.75万
依托单位：
UNIVERSITY OF COLORADO DENVER
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-08-01 至 2021-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9313332
关键词：
1q21 Address Affect Alleles Autistic Disorder Biological Assay Brain Brain Diseases CRISPR/Cas technology Cell Proliferation Chromosomes Cognition Disorders Collaborations Copy Number Polymorphism DNA DNA copy number Data Data Set Development Disease Dose Evolution Exhibits Family Family member Genetic Genome Genome Mappings Genomic DNA Genomics Growth Haploidy Human Human Genome Individual Investigation Label Link Macrocephaly Maps Mass Spectrum Analysis Measurement Measures Methods Microcephaly National Institute of Mental Health Neurons Normal Statistical Distribution Open Reading Frames Optics Pathologic Pathway interactions Population Process Reporting Resolution Resources Risk Risk Factors Role Sampling Schizophrenia Series Severities Severity of illness Source Symptoms Technology Tertiary Protein Structure Testing Time Transfection Variant brain abnormalities brain size disorder risk dosage improved insight nerve stem cell novel novel strategies protein expression transcriptome sequencing whole genome

项目摘要

SUMMARY DUF1220 protein domains have undergone the largest human lineage-specific increase in copy number of any protein coding region in the genome (~290 copies in human haploid genome), and map primarily to 1q21, a region where copy number variations containing many DUF1220 copies have been repeatedly linked to autism, schizophrenia, micro- and macrocephaly. DUF1220 copy number exhibits a broad Gaussian distribution in human populations, and is a rich source of unexamined functional allelic variation. Previously we have implicated DUF1220 copy number (dosage) in human brain expansion and normal and pathological changes in brain size. Over the past year we have demonstrated that DUF1220 (CON1 subtype) dosage is linearly associated with increasing symptom severity of autism (now confirmed by a replication study) and also inversely with schizophrenia severity and risk. These findings are remarkable in that they imply that dosage variations within the same protein domain family (DUF1220) may be involved in human brain evolution, autism and schizophrenia, and that these processes may be genetically and mechanistically interrelated. Building on these significant findings, we will expand our study of DUF1220 copy number in autism and schizophrenia severity (Aim 1) and risk (Aim 2) and in micro-/macrocephaly (Aim 3) using methods we have used successfully (ddPCR, 1q21-targeted arrayCGH) as well as through application of novel methods that have the potential to significantly improve DUF1220 copy number analysis (Aim 4). For example, we have recently optimized and validated our sequence read-depth method for measuring DUF1220 copy number at high resolution and precision from whole genome sequence (WGS) data. We will apply this approach to WGS data from 1) the Autism 10K project in collaboration with Dr. Stephen Scherer, and 2) to several additional large WGS datasets that are emerging for autism and schizophrenia. In addition, we have established a subcontract with our collaborator, Dr. Pui Kwok, to apply Irys, an optical mapping method for automated genome mapping of long (>150kb) single DNA molecules, to assess DUF1220 copy number variation in these disorders. The Irys approach should be aided by our development of a CRISPR-Cas9 method which allows DUF1220-specific labeling of genomic DNA. These approaches, including our exploration of long-read sequencing technologies, will be facilitated by utilization of a new, more accurate and gap-free 1q21 assembly we recently helped generate. Finally, we will build on our recent progress linking DUF1220 function to increases in neuron number by testing whether DUF1220 promotes neural stem cell (H9-derived) proliferation in a dosage-dependent manner and identifying cellular pathways affected by progressively increasing DUF1220 dosage.

概括 DUF1220 蛋白结构域经历了最大的人类谱系特异性拷贝数增加基因组中任何蛋白质编码区（人类单倍体基因组中约 290 个拷贝），并主要映射到 1q21，包含许多 DUF1220 拷贝的拷贝数变异重复连接的区域自闭症、精神分裂症、小头畸形和大头畸形。 DUF1220 拷贝数表现出广泛的高斯分布人类群体中的分布，并且是未经检查的功能等位基因变异的丰富来源。以前我们表明 DUF1220 拷贝数（剂量）与人脑扩张以及正常和病理性的有关大脑大小的变化。在过去的一年里，我们已经证明 DUF1220（CON1 亚型）剂量与自闭症症状严重程度的增加呈线性相关（现已通过复制研究证实），并且与精神分裂症的严重程度和风险成反比。这些发现是值得注意的，因为它们意味着剂量同一蛋白质结构域家族（DUF1220）内的变异可能与人类大脑进化、自闭症有关和精神分裂症，并且这些过程可能在遗传和机制上相互关联。基于这些重要发现，我们将扩大对自闭症和 DUF1220 拷贝数的研究使用我们拥有的方法评估精神分裂症的严重程度（目标 1）和风险（目标 2）以及小/大头畸形（目标 3）成功使用（ddPCR、1q21 靶向 arrayCGH）以及通过应用新方法有可能显着改善 DUF1220 拷贝数分析（目标 4）。例如，我们有最近优化并验证了我们用于测量 DUF1220 拷贝数的序列读取深度方法全基因组序列 (WGS) 数据的高分辨率和精确度。我们将把这种方法应用于 WGS 数据来自 1) 与 Stephen Scherer 博士合作的 Autism 10K 项目，以及 2) 其他几个项目正在出现的关于自闭症和精神分裂症的大型 WGS 数据集。此外，我们还设立了一个与我们的合作者 Pui Kwok 博士分包，应用 Irys，一种光学测绘方法，用于自动化长 (>150kb) 单 DNA 分子的基因组作图，以评估这些分子中的 DUF1220 拷贝数变异失调。 Irys 方法应该得到我们开发的 CRISPR-Cas9 方法的帮助，该方法允许基因组 DNA 的 DUF1220 特异性标记。这些方法，包括我们对长读的探索测序技术将通过使用新的、更准确且无间隙的 1q21 组装来促进我们最近帮助生成了。最后，我们将在最近将 DUF1220 功能链接到通过测试 DUF1220 是否促进神经干细胞（H9 衍生）增殖来增加神经元数量以剂量依赖的方式并识别受逐渐增加影响的细胞途径 DUF1220用量。