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: 7941810
• 负责人: THOMAS W GLOVER
• 金额: $ 49.99万
• 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 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7941810
• 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）： 本申请涉及广泛的挑战领域，（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：“基因组的微观缺失和复制因其在许多复杂的人类疾病中的潜在作用而引起越来越多的关注。特别令人感兴趣的是自发性CNV，定义为存在于受影响个体中，但在双亲中均不存在的CNV。对自发性CNV如何产生的理解有限。需要进行研究，以确定环境暴露是否会影响拷贝数变异和其他结构变异的风险，这些变异与复杂疾病有关。鉴于这一研究领域的早期阶段，研究应侧重于体外暴露细胞的变化”。近年来，拷贝数变异（copy number variants，CNVs）广泛分布于人类基因组中，是遗传变异的重要组成部分，在表型多样性、复杂疾病和进化中起着不可或缺的作用。在健康个体中，超过1300个频率>1%的CNV被描述为缺失或重复，大小从几Kb到超过Mb，随着更多数据的可用，这个数字肯定会增加。类似的自发性CNV现在众所周知是遗传和发育障碍的主要原因，包括精神发育迟滞、自闭症、精神分裂症、癫痫、腭裂和许多其他疾病。对特发性精神发育迟滞和发育障碍的研究发现，在5-17%的受影响个体中存在新生CNV，这表明突变率很高。类似的拷贝数改变也在许多癌症中以高频率发现，其中可能在起始或进展中起作用。因此，CNV是正常遗传变异和进化中的关键因素，并且是遗传和发育障碍以及癌症中非常重要的一类突变。对于CNVs如何产生、所涉及的细胞机制和风险因素以及环境因子对其形成的影响的理解非常有限。与所有突变类别一样，几乎可以肯定的是，环境损伤可以诱导或增加新的和有害的CNV的风险，然而我们对这些事件的机制和频率知之甚少。由侧翼重复序列或节段性重复介导的减数分裂不平等重组[或非等位基因同源重组（NAHR）]导致许多复发性、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的大多数，是通过与异常DNA复制和/或DNA损伤的非同源修复偶联的机制产生的。这表明一个意想不到的有丝分裂，而不是减数分裂，细胞起源的许多CNVs，并有一些重要的影响，环境暴露的作用，在其形成和发展的体外模型系统，他们的研究。我们的目标是确定环境因素在CNVs形成中的作用，并对这些频繁突变产生的机制获得新的见解。我们已经组建了一支强大的研究团队，并开发了一个正常的人类细胞培养模型系统，加上领先的基因组分析技术，使我们处于独特的位置，以应对这一及时的挑战。使用这个系统，我们发现，aphidicolin诱导的复制应激导致一个显着的高频率的从头CNVs。这些发现使我们假设环境诱导的复制应激和/或DNA双链断裂是导致人类生殖系和癌细胞中有丝分裂细胞分裂期间CNVs的两个主要因素。为了验证这一假设，我们将在全基因组范围内表征环境因子诱导的CNV，并直接比较这两种非排他性CNV形成模型的强度，从而首次提供了两种不同类别的环境因子的基因组表现的高分辨率目录，最有可能与CNV形成相关。这些是（1）导致复制应激的试剂，其可能通过二次断裂或复制模板转换导致CNV，和（2）直接诱导DNA双链断裂（DNA DSB）的试剂，其可能通过断裂末端的不适当连接导致CNV。具体来说，我们将研究羟基脲和叶酸的压力，通过不同的机制比aphidicolin抑制复制，电离辐射和博莱霉素，直接导致DNA双链断裂。我们的“铲准备”体外模型系统加上高分辨率基因组微阵列和下一代测序将使我们能够确定这些药物对CNVs的频率，频谱，分布和结构以及其他亚显微结构变异的影响。综合结果将解决我们对一类非常重要的突变的知识中的一个主要空白，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子。