Environmental Risk Factors for Copy Number Variation in Human Chromosomes

人类染色体拷贝数变异的环境风险因素

• 批准号: 7817619
• 负责人: THOMAS W GLOVER
• 金额: $ 48.56万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-28 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7817619
• 关键词: Accounting; Address; Affect; Aphidicolin; Area; Attention; Autistic Disorder; Biological Models; Bleomycin; Cataloging; Catalogs; Categories; Cell Culture Techniques; Cell division; Cells; Chromosomal Rearrangement; Chromosome Breakage; Chromosomes; Cleft Palate; Complex; Congenital Abnormality; Congenital Heart Defects; Copy Number Polymorphism; Coupled; DNA; DNA Damage; DNA Double Strand Break; DNA Repair; DNA Sequence Rearrangement; DNA biosynthesis; Data; Data Set; Detection; Development; Disease; Dose; Environmental Exposure; Environmental Risk Factor; Epilepsy; Event; Evolution; Experimental Models; Exposure to; Floxuridine; Folate; Folic Acid Antagonists; Frequencies; Genes; Genetic; Genetic Recombination; Genetic Variation; Genome; Genomics; Goals; Hereditary Disease; Human; Human Chromosomes; Human Genome; In Vitro; Individual; Ionizing radiation; Knowledge; Lead; Lymphoid; Malignant Neoplasms; Mediating; Meiosis; Mental Retardation; Mental disorders; Methods; Microscope; Microscopic; Mitotic; Modeling; Monitor; Mutate; Mutation; Nature; Normal Cell; Paper; Parents; Partner in relationship; Pattern; Play; Population; Positioning Attribute; Prevention; Process; Publishing; Recovery; Recurrence; Recurrent disease; Research; Research Personnel; Resolution; Ribonucleotide Reductase Inhibitor; Risk; Risk Factors; Role; Schizophrenia; Staging; Stress; Structure; System; Techniques; Technology; Terminal Repeat Sequences; Testing; Time; Variant; base; cancer cell; chromosome replication; comparative; cost; developmental disease; egg; environmental agent; genetic risk factor; genome-wide; homologous recombination; human disease; hydroxyurea; in vitro Model; in vivo; insight; interest; next generation; novel; sperm cell; structural genomics; tool

DESCRIPTION (provided by applicant): This application addresses broad Challenge Area, (08) Genomics and Specific Challenge Topic, 08-ES-106: The role of environmental exposure in copy number variation (CNV): "Microscopic deletions and replications of the genome have attracted increasing attention for their potential role in many complex human diseases. Of particular interest are spontaneous CNVs, defined as those present in an affected individual, but absent in both parents. There is limited understanding of how spontaneous CNVs arise. Studies are needed that will determine whether environmental exposures can affect risk for copy number variation and other structural variations that have been implicated in complex diseases. Given the early stage of this research area, studies should focus on changes in cells exposed in vitro". In recent years, copy number variants (CNVs) have been found to be widely distributed throughout the human genome where they represent an important component of genetic variation and play an integral role in phenotypic diversity, complex disease and evolution. Over 1300 CNVs with frequencies of >1% have been described in healthy individuals that exist as deletions or duplications ranging in size from a few Kb to over a Mb, and this number will surely increase as more data become available. Similar spontaneous CNVs are now well known to be a major cause of genetic and developmental disorders, including mental retardation, autism, schizophrenia, epilepsy, cleft palate and many others. Studies of idiopathic mental retardation and developmental disorders have found de novo CNVs in 5-17% of affected individuals, suggesting a high mutation rate. Similar copy number alterations are also found at high frequency in many cancers where a role in initiation or progression is likely. CNVs are thus a key factor in normal genetic variation and evolution and are a very important class of mutation in genetic and developmental disorders and cancer. There is very limited understanding of how CNVs arise, the cellular mechanisms and risk factors that are involved, and the effects of environmental agents on their formation. As with all mutation classes, it is almost certain that environmental insults can induce or increase the risk for new and deleterious CNVs, however we have little knowledge of the mechanisms and frequency of such events. Meiotic unequal recombination [or non-allelic homologous recombination (NAHR)] mediated by flanking repeated sequences or segmental duplications leads to many recurrent, disease-related CNVs. However, there is growing evidence that many or most normal and sporadic, nonrecurrent CNVs, which account for the majority of disease-associated CNVs in humans and those in cancers arise via mechanisms coupled to aberrant DNA replication and/or non-homologous repair of DNA damage. This suggests an unexpected mitotic, rather than meiotic, cell origin for many CNVs and has a number of important implications for the role of environmental exposures in their formation and the development of in vitro model systems for their study. The Challenge, and our goal, is to determine the role of environmental factors in the formation of CNVs and to gain novel insight into the mechanisms by which these frequent mutations are generated. We have assembled a strong team of investigators and have developed a normal human cell culture model system coupled with leading edge genome analysis technologies, placing us in a unique position to address this timely Challenge. Using this system we have found that aphidicolin-induced replication stress leads to a remarkably high frequency of de novo CNVs. These findings lead us to hypothesize that environmentally-induced replication stress and/or DNA double strand breaks are two major factors leading to CNVs during mitotic cell divisions in the human germline and in cancer cells. To test this hypothesis, we will characterize environmental agent-induced CNVs at the genome-wide scale and directly compare the strength of these two non-exclusive models for CNV formation, thus providing for the first time a high resolution catalog of genomic manifestations of two different categories of environmental agents most likely to be associated with CNV formation. These are (1) agents that lead to replication stress, which might lead to CNVs through secondary breakage or replicative template switching, and (2) agents that directly induce DNA double-strand breaks (DNA DSBs), which might lead to CNVs through inappropriate joining of broken ends. Specifically, we will examine hydroxyurea and folate stress, which inhibit replication through different mechanisms than aphidicolin, and on ionizing radiation and bleomycin, which lead directly to DNA DSBs. Our "shovel ready" in vitro model system coupled with high resolution genomic microarrays and next-generation sequencing will allow us to determine the effects of these agents on the frequency, spectrum, distribution and structure of CNVs and other submicroscopic structural variations. The combined results will address a major gap in our knowledge about a very important class of mutations and allow predictions of the environmental agents that confer the greatest risk for ongoing structural alteration of the human genome.

描述（由申请人提供）： 该应用程序解决了广泛的挑战领域，(08) 基因组学和特定挑战主题，08-ES-106：环境暴露在拷贝数变异 (CNV) 中的作用：“基因组的微观删除和复制因其潜力而受到越来越多的关注人们特别感兴趣的是自发性 CNV，其定义为受影响个体中存在但父母双方均不存在的自发性 CNV。需要进行研究来确定环境暴露是否会产生影响。风险鉴于该研究领域处于早期阶段，研究应集中于体外暴露的细胞的变化。近年来，人们发现拷贝数变异（CNV）广泛分布在整个人类基因组中，它们是遗传变异的重要组成部分，在表型多样性、复杂疾病和进化中发挥着不可或缺的作用。据描述，健康个体中有超过 1300 个频率 >1% 的 CNV，它们以缺失或重复的形式存在，大小从几 Kb 到超过 1 Mb，随着更多数据的出现，这个数字肯定会增加。现在众所周知，类似的自发 CNV 是遗传和发育障碍的主要原因，包括智力低下、自闭症、精神分裂症、癫痫、腭裂等。对特发性精神发育迟滞和发育障碍的研究发现，5-17% 的受影响个体中存在新生 CNV，表明突变率很高。在许多癌症中也频繁发现类似的拷贝数改变，这些改变可能在起始或进展中发挥作用。因此，CNV 是正常遗传变异和进化的关键因素，也是遗传和发育障碍以及癌症中非常重要的一类突变。对于 CNV 的产生方式、所涉及的细胞机制和风险因素以及环境因素对其形成的影响，人们的了解非常有限。与所有突变类别一样，几乎可以肯定的是，环境损害会诱发或增加新的有害 CNV 的风险，但我们对此类事件的机制和频率知之甚少。由侧翼重复序列或片段重复介导的减数分裂不等重组[或非等位基因同源重组(NAHR)]会导致许多复发的、与疾病相关的CNV。然而，越来越多的证据表明，许多或大多数正常和散发的非复发性 CNV（占人类和癌症中疾病相关 CNV 的大部分）是通过与异常 DNA 复制和/或 DNA 非同源修复相关的机制产生的。损害。这表明许多 CNV 的细胞起源是出乎意料的有丝分裂，而不是减数分裂，并且对于环境暴露在其形成中的作用以及其研究的体外模型系统的开发具有许多重要意义。我们的挑战和目标是确定环境因素在 CNV 形成中的作用，并获得对这些频繁突变产生机制的新见解。我们组建了一支强大的研究人员团队，并开发了一个正常的人类细胞培养模型系统，并结合了领先的基因组分析技术，使我们处于独特的地位来应对这一及时的挑战。使用该系统，我们发现阿非迪霉素诱导的复制应激导致从头 CNV 的出现频率非常高。这些发现使我们推测，环境诱导的复制应激和/或 DNA 双链断裂是人类种系和癌细胞有丝分裂细胞分裂过程中导致 CNV 的两个主要因素。为了检验这一假设，我们将在全基因组范围内表征环境因素诱导的 CNV，并直接比较这两种非排他性 CNV 形成模型的强度，从而首次提供两种基因组表现的高分辨率目录。最有可能与 CNV 形成相关的不同类别的环境因素。这些是 (1) 导致复制应激的试剂，可能通过二次断裂或复制模板切换导致 CNV，以及 (2) 直接诱导 DNA 双链断裂 (DNA DSB) 的试剂，可能通过不适当的方式导致 CNV断头的连接。具体来说，我们将检查羟基脲和叶酸应激（它们通过与阿菲迪霉素不同的机制抑制复制）以及电离辐射和博莱霉素（直接导致 DNA DSB）。我们的“随时可用”体外模型系统与高分辨率基因组微阵列和下一代测序相结合，将使我们能够确定这些药物对 CNV 的频率、频谱、分布和结构以及其他亚显微结构变异的影响。综合结果将弥补我们对一类非常重要的突变的认识上的重大差距，并允许预测对人类基因组持续结构改变带来最大风险的环境因素。