Genetic and Molecular Analyses of Human LINE-1 Retrotransposition

摘要 长插入元件-1（LINE-1或L1）转座元件衍生的序列包含 大约17%的人类基因组DNA。尽管绝大多数LINE-1都是 在进化过程中，由于突变过程的影响，一个普通的人类基因组大约包含 100个有活性的LINE-1可以通过RNA中间体移动到新的基因组位置， 反转录转位在生殖系中，在早期发育过程中，或在选择性染色体中， 体细胞继续产生个体间和个体内的遗传变异，并可能导致散发病例 人类遗传疾病，包括血友病A，血友病B，杜氏肌营养不良症，和某些 癌的我们认为，对LINE-1生物学的机械理解对于理解 对人类疾病、人类遗传变异和人类进化都有贡献。这一提议是第三次 竞争性更新申请GM 060518，并寻求持续的支持，以继续我们的LINE-1研究 生物学我们将在移动的遗传元素领域积累经验和知识， 从进化相关的反转录转座子的移动机制的研究中获得，并采用LINE- 1个培养细胞逆转录转座测定结合结合遗传， 分子生物学、生物化学、多组学和计算方法来回答前沿问题 在LINE-1生物学中。该提案包括两个目标，它们建立在我们广泛的试剂工具箱基础上， 以前的调查结果和初步数据。在目标1中，我们将阐明LINE-1的机制方面 反转录转位在目标2中，我们将确定宿主蛋白如何促进或限制LINE-1 反转录转位该项目将受益于莫兰和 密歇根大学的基德实验室和我们杰出的研究团队。圆满完成 上述特定目的的研究将增加我们对LINE-1如何移动到新的基因组位置的理解， 宿主已经进化出防止LINE-1逆转录转座的机制，以及LINE-1如何利用 细胞蛋白质来帮助其逆转录转座。这些知识也将提供关于LINE-1如何 反转录转座有助于人类疾病、人类遗传变异和人类基因组进化。

Abstract Long INterspersed Element-1 (LINE-1 or L1) transposable element derived sequences comprise approximately 17% of human genomic DNA. Although the overwhelming majority of LINE-1s have been rendered immobile by mutational processes over evolutionary time, an average human genome contains approximately 100 active LINE-1s that can mobilize to new genomic locations via an RNA intermediate by a process known as retrotransposition. The retrotransposition of active LINE-1s in the germline, during early development, or in select somatic cells continues to generate inter- and intra-individual genetic variation and can lead to sporadic cases of human genetic diseases, including hemophilia A, hemophilia B, Duchenne muscular dystrophy, and certain cancers. We posit that a mechanistic understanding of LINE-1 biology is essential to understand the forces that contribute to human disease, human genetic variation, and human evolution. This proposal represents the third competing renewal application of GM060518, and seeks ongoing support to continue our studies of LINE-1 biology. We will build upon our experience and knowledge in the field of mobile genetic elements, use insights gained from studies of the mobility mechanisms of evolutionarily related retrotransposons, and employ a LINE- 1 cultured cell retrotransposition assay in conjunction with an integrated strategy that combines genetic, molecular biological, biochemical, multi-omic, and computational approaches to answer cutting-edge questions in LINE-1 biology. This proposal comprises two Aims, which build upon our extensive toolbox of reagents, previous findings, and preliminary data. In Aim 1, we will elucidate mechanistic aspects of LINE-1 retrotransposition. In Aim 2, we will determine how host proteins act to facilitate or restrict LINE-1 retrotransposition. This project will benefit from a long-standing successful collaboration between the Moran and Kidd laboratories at the University of Michigan and our outstanding research teams. The successful completion of the above Specific Aims will increase our understanding how LINE-1s mobilize to new genomic locations, how the host has evolved mechanisms to prevent unabated LINE-1 retrotransposition, and how LINE-1 has exploited cellular proteins to aid in its retrotransposition. This knowledge also will provide insights into how LINE-1 retrotransposition contributes to human disease, human genetic variation, and human genome evolution.