LINE-1 Retrotransposition in Human Embryonic Stem Cells

人类胚胎干细胞中的 LINE-1 逆转座

• 批准号: 8072570
• 负责人: JOHN V. MORAN
• 金额: $ 23.55万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-08-01 至 2013-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8072570
• 关键词: Affect; Alu Elements; Area; Bar Codes; Biological Assay; Biological Models; Cell Culture Techniques; Cell Line; Cells; Chromosomes; Colon Carcinoma; Cultured Cells; DNA Sequence; Development; Disease; Embryo; Embryonic Development; Engineering; Event; Evolution; Functional RNA; Genes; Genome; Genomic Instability; Germ Lines; Hemophilia A; Hereditary Disease; Human; Human Development; Human Engineering; Human Genome; In Vitro; L1 Elements; Laboratories; Libraries; Long Interspersed Elements; Mediating; Michigan; Molecular; Monitor; Muscular Dystrophies; Mutate; Mutation Spectra; Neuronal Differentiation; Parents; Pattern; Play; Process; Proteins; Protocols documentation; Pseudogenes; Rattus; Retrotransposition; Retrotransposon; Role; Short Interspersed Nucleotide Elements; Somatic Cell; Staging; System; Testing; Transgenic Mice; Universities; Variant; base; cell type; human DNA; human embryonic stem cell; in vitro Model; in vivo; nerve stem cell; relating to nervous system; tumorigenesis

DESCRIPTION (provided by applicant): Long Interspersed Element-1 (LINE-1 or L1) is an abundant retrotransposon that comprises ~17% of human DNA. The overwhelming majority of L1s can no longer move (i.e., retrotranspose). However, the average human genome contains ~80-100 retrotransposition-competent L1s (RC-L1s) and their mobility, in both germ line and somatic cells, has resulted in a variety of genetic disorders, including Hemophilia A, muscular dystrophy, and colon cancer. The proteins encoded by RC-L1s also likely are responsible for the mobilization of Alu elements, certain Short Interspersed Elements (SINEs) and the formation of processed pseudogenes, which together comprise ~13% of human DNA. Thus, either directly or by the promiscuous mobilization of cellular RNAs, L1-mediated retrotransposition events are responsible for at least one billion bases in human DNA and have had a tremendous impact on human genome evolution. Recent studies indicate that L1 retrotransposition events can occur during early embryogenesis and in neural progenitor cells. We hypothesize that hES cells represent a developmentally relevant in vitro model system to study the mechanisms by which L1- mediated retrotransposition events impact genome integrity at various stages of human development. An understanding of the basic molecular processes that impact genome integrity in hES cells is important to evaluate since these cells ultimately may be used therapeutically to treat a wide-range of diseases. The specific aims of this proposal are: 1. To determine the impact of L1 retrotransposition events in hES cells. 2. To determine the constellation of endogenous L1s that are mobilized in hES cells. 3. To determine how L1 retrotransposition impacts neuronal differentiation. Lay Narrative: LINE-1 elements are DNA sequences that can "jump" from one area of a chromosome into another by a process called retrotransposition. They comprise ~17% of the human DNA and have played a major role in genome evolution. The vast majorities of LINE-1 elements are mutated and can no longer jump; however, ~100 LINE-1 elements are still able to mobilize. Deleterious LINE-1 insertions into genes have caused diseases such as Hemophilia A, muscular dystrophy, and also are implicated in tumorigenesis. The proteins encoded by LINE-1 elements also are responsible for the mobilization of Alu elements, certain Short Interspersed Elements (SINEs) and the formation of processed pseudogenes, which together comprise at least 13% of human DNA. Thus, either directly or by the promiscuous mobilization of cellular RNAs, L1-mediated retrotransposition events are responsible for at least one billion bases in human DNA and have had a tremendous impact on human genome evolution. Despite these findings, it still is unclear how often, in what cell types, and when during development L1 retrotransposes in vivo. Recent studies indicate that L1 retrotransposition events can occur during early embryogenesis and in neural progenitor cells. We hypothesize that hES cells represent a developmentally relevant in vitro model system to study the mechanisms by which L1-mediated retrotransposition events impact genome integrity at various stages of human development. An understanding of the basic molecular processes that impact genome integrity in hES cells is important to evaluate since these cells ultimately may be used therapeutically to treat a wide-range of diseases.

描述（由申请人提供）：长散布元件-1（LINE-1 或 L1）是一种丰富的逆转录转座子，包含约 17% 的人类 DNA。绝大多数 L1 不能再移动（即逆转录转座）。然而，人类基因组平均包含约 80-100 个具有逆转录转座能力的 L1 (RC-L1)，它们在生殖系和体细胞中的迁移性导致了多种遗​​传性疾病，包括 A 型血友病、肌营养不良症和结肠癌。 RC-L1 编码的蛋白质也可能负责 Alu 元件、某些短散布元件 (SINE) 的动员以及加工假基因的形成，这些假基因总共构成约 13% 的人类 DNA。因此，无论是直接的还是通过细胞 RNA 的混杂动员，L1 介导的逆转录转座事件负责人类 DNA 中至少 10 亿个碱基，并对人类基因组进化产生了巨大影响。最近的研究表明，L1 逆转录转座事件可能发生在早期胚胎发生期间和神经祖细胞中。我们假设 hES 细胞代表了一种与发育相关的体外模型系统，用于研究 L1 介导的逆转录转座事件在人类发育各个阶段影响基因组完整性的机制。了解影响 hES 细胞基因组完整性的基本分子过程对于评估非常重要，因为这些细胞最终可用于治疗多种疾病。该提案的具体目标是： 1. 确定 L1 逆转录转座事件对 hES 细胞的影响。 2. 确定 hES 细胞中动员的内源性 L1 群。 3. 确定L1逆转录转座如何影响神经元分化。 通俗叙述：LINE-1 元件是 DNA 序列，可以通过称为逆转录转座的过程从染色体的一个区域“跳”到另一个区域。它们约占人类 DNA 的 17%，在基因组进化中发挥着重要作用。绝大多数LINE-1元件发生突变，无法再跳跃；然而，大约 100 个 LINE-1 元件仍然能够动员。有害的 LINE-1 插入基因会导致 A 型血友病、肌营养不良症等疾病，并且还与肿瘤发生有关。 LINE-1 元件编码的蛋白质还负责 Alu 元件、某些短散布元件 (SINE) 的动员以及加工假基因的形成，这些假基因共同构成了至少 13% 的人类 DNA。因此，无论是直接的还是通过细胞 RNA 的混杂动员，L1 介导的逆转录转座事件负责人类 DNA 中至少 10 亿个碱基，并对人类基因组进化产生了巨大影响。尽管有这些发现，但仍不清楚 L1 在体内逆转座的频率、细胞类型以及发育过程中的时间。最近的研究表明，L1 逆转录转座事件可能发生在早期胚胎发生期间和神经祖细胞中。我们假设 hES 细胞代表了一种与发育相关的体外模型系统，用于研究 L1 介导的逆转录转座事件在人类发育各个阶段影响基因组完整性的机制。了解影响 hES 细胞基因组完整性的基本分子过程对于评估非常重要，因为这些细胞最终可用于治疗多种疾病。