Genetic and Molecular Analyses of Human LINE-1 Retrotransposition

人类 LINE-1 逆转录转座的遗传和分子分析

• 批准号: 8309244
• 负责人: JOHN V. MORAN
• 金额: $ 29.64万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-09-30 至 2015-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8309244
• 关键词: Affect; Alu Elements; Baculovirus Expression System; Beryllium; Biochemical; Biological; Biological Assay; Cells; Cultured Cells; Cytidine Deaminase; DNA Damage; DNA Repair; DNA Repair Pathway; DNA Sequence; DNA Transposable Elements; Development; Disease; Elements; Engineering; Event; Evolution; Family; Functional RNA; Funding; Gene Family; Genes; Genetic; Genetic Variation; Genome; Genomics; Germ Lines; Hereditary Disease; Host Defense; Human; Human Genome; In Vitro; Individual; Integration Host Factors; L1 Elements; Laboratories; Lead; Learning; Length; Location; Long Interspersed Elements; Mammalian Cell; Mediating; Messenger RNA; Minisatellite Repeats; Mobile Genetic Elements; Molecular; Molecular Genetics; Monitor; Mutation; Nonhomologous DNA End Joining; Paste substance; Pathway interactions; Process; Proteins; Protocols documentation; RNA; Reagent; Retrotransposition; Retrotransposon; Reverse Transcription; Short Interspersed Nucleotide Elements; Small Nucleolar RNA; Somatic Cell; Structure; System; U6 small nuclear RNA; Variant; human DNA; human disease; insight; meetings; member; public health relevance; reconstitution; response; success; tool

DESCRIPTION (provided by applicant): Long Interspersed Element-1 (LINE-1 or L1) is an abundant mobile genetic element that comprises approximately 17% of human DNA. Active human L1s are about 6 kb in length and they encode two proteins (ORF1p and ORF2p) that are critical for retrotransposition. Ongoing LINE-1 retrotransposition events contribute to inter-individual genetic variation, and, on occasion, LINE-1 insertions into genes can result in genetic disease. During the past decade, my laboratory has developed efficient assays to monitor LINE-1 retrotransposition in cultured mammalian cells. Here, we will use these assays in conjunction with genetic, molecular, and biochemical approaches to further elucidate the mechanism of L1 retrotransposition. We also will identify host proteins that affect LINE-1 retrotransposition in human cells. A basic mechanistic understanding of the process of L1 retrotransposition will lead to greater insight about how transposable element activity contributes to human disease, human genome evolution, and human diversity. PUBLIC HEALTH RELEVANCE: Long INterspersed Element-1 (LINE-1) is an abundant mobile genetic element that comprises 17% of human DNA. Active LINE-1 elements are able to retrotranspose or "jump," inserting themselves into a new genomic location by a copy and paste mechanism. On occasion, new LINE-1 insertions in the germ line, in early development, or in somatic cells can result in disease-producing mutations. Despite the mutagenic potential of LINE-1 elements, little is known about the molecular mechanism of LINE-1 retrotransposition and even less is known about cellular proteins that influence this process. During the past decade, my laboratory has developed a toolbox to study LINE- 1 retrotransposition in a controlled manner in mammalian cultured cells. These tools have allowed significant progress in this field by my lab and others. In this proposal, we will use genetic, molecular biological, and biochemical approaches to further elucidate the molecular mechanism of LINE-1 retrotransposition. We also will explore how mutations in host proteins involved in host defense and/or DNA repair pathways affect LINE-1 jumping. Through these studies, we will learn more about how this intriguing family of repetitive DNA sequences contributes to the structure and function of the human genome.

描述（由申请人提供）：长散布元件-1（LINE-1或L1）是一种丰富的移动的遗传元件，约占人类DNA的17%。活性人L1长约6 kb，它们编码两种对逆转录转座至关重要的蛋白质（ORF 1 p和ORF 2 p）。正在进行的LINE-1逆转录转座事件有助于个体间的遗传变异，有时，LINE-1插入基因可能导致遗传疾病。在过去的十年中，我的实验室已经开发出有效的检测方法来监测培养的哺乳动物细胞中LINE-1的逆转录转座。在这里，我们将使用这些检测结合遗传，分子和生化方法，以进一步阐明L1逆转录转座的机制。我们还将鉴定影响人类细胞中LINE-1逆转录转座的宿主蛋白。对L1反转录转座过程的基本机制理解将使我们更深入地了解转座因子活性如何促进人类疾病、人类基因组进化和人类多样性。 公共卫生相关性：长插入元件-1（LINE-1）是一种丰富的移动的遗传元件，占人类DNA的17%。活性LINE-1元件能够反转录转座或“跳跃”，通过复制和粘贴机制将其自身插入新的基因组位置。有时，在生殖系、早期发育或体细胞中插入新的LINE-1可能导致致病突变。尽管LINE-1元件具有致突变潜力，但对LINE-1逆转录转座的分子机制知之甚少，对影响这一过程的细胞蛋白质知之更少。在过去的十年中，我的实验室开发了一个工具箱，以研究LINE- 1逆转录转座在哺乳动物培养细胞中的控制方式。这些工具使我的实验室和其他人在这一领域取得了重大进展。在本研究中，我们将利用遗传学、分子生物学和生物化学的方法进一步阐明LINE-1逆转录转座的分子机制。我们还将探讨宿主防御和/或DNA修复途径中涉及的宿主蛋白质突变如何影响LINE-1跳跃。通过这些研究，我们将更多地了解这个有趣的重复DNA序列家族如何对人类基因组的结构和功能做出贡献。