Silencing of Mobile Genetic Elements by Small RNAs

描述（由申请人提供）：该项目的目标是表征转座子沉默途径，特别关注小rna的内源性作用。自20世纪80年代末以来，通过使用经典遗传工具来询问与转座机制相关的问题，已经在秀丽隐杆线虫中很好地表征了转座子。1999年，人们发现内源性小RNA通路调节转座子，并进行了几次筛选，以分离出种系中具有活跃转座子的突变体。然而，尽管在这一途径的基因中已经分离出许多突变体，但很少有研究对单个蛋白质的作用进行机械表征。此外，由于大多数转座子运动的分析是在高通量测序出现之前完成的，因此没有检查转座子运动的全基因组数据。除转座子外，许多其他类型的转录本也通过内源性RNA途径加工，包括异常转录本、复制和假设基因以及非编码区，然而，这些转录本被识别并进入这些途径的机制是完全未知的。通过探测这些途径的各个组成部分，检查它们与什么种类的RNA相关联，以及它们的酶功能对什么过程有贡献，该项目将对转座子沉默有更全面的了解。本文提出的实验旨在确定调节转座子动员的因素，包括基因组特征、宿主蛋白和环境刺激，目的是阐明该途径中已知和新成分的作用机制。这项工作的结果将对理解活性转座子如何影响基因表达和改变基因组结构具有重要意义，这可能导致包括癌症在内的许多疾病。Ruvkun实验室是研究秀丽隐杆线虫中RNAi机制对转座子调控的理想场所。该实验室不仅在小RNA领域拥有广泛的专业知识，而且在应激反应途径，高通量遗传筛选，蛋白质纯化和生物化学方面也有广泛的专业知识。实验室环境包含了一群对秀丽隐杆线虫生物学基本方面感兴趣的非常积极和科学多样化的研究人员。该实验室两侧有两个研究神经生物学和发病机理的优秀秀丽隐杆线虫实验室，并由一个高度协作的部门提供支持，该部门拥有RNA生物学和染色质结构方面的专业知识。我计划在剩余的博士后任期内利用所有这些资源。在我的研究生涯中，我一直专注于染色质和RNA生物学，这对理解和预防癌症至关重要。具体来说，对RNA沉默和转座子沉默的更深入了解可能会导致癌症诊断或治疗的进步。我致力于学术研究的事业，并计划围绕此提案的目标建立一个实验室。

DESCRIPTION (provided by applicant): The goal of this project is to characterize the transposon-silencing pathway, specifically focusing on the endogenous role of small RNAs. Since the late 1980's, transposons have been well characterized in C. elegans through the use of classical genetic tools to interrogate questions related to mechanisms of transposition. In 1999, it was discovered that endogenous small RNA pathways regulate transposons and several screens were performed to isolate mutants with active transposition in the germline. However, although many mutants have been isolated in genes attributed to this pathway, little work has been done to mechanistically characterize the roles of individual proteins. Additionally, because most analyses of transposon movement were done prior to the advent of high-throughput sequencing, there is no genome-wide data examining transposon mobilization. In addition to transposons, many other types of transcripts are processed through the endogenous RNA pathways, including aberrant transcripts, duplicated and hypothetical genes, and non-coding regions, however the mechanism by which these transcripts are recognized and routed into these pathways is completely unknown. By probing the individual components of these pathways, examining what species of RNA they associate with, and to what process their enzymatic functions contribute, this project will achieve a more complete understanding of transposon silencing. The proposed experiments herein aim to identify factors that regulate transposon mobilization, including genomic features, host proteins, and environmental stimuli with the goal of elucidating the mechanisms of action of the known and novel components of this pathway. The results of this work will have significant implications in understanding how active transposons affect gene expression and alter genome structure, which can result in many diseases including cancer. The Ruvkun lab is an ideal place to study regulation of transposons by RNAi machinery in C. elegans. The lab not only has extensive expertise in the small RNA field, but also in stress response pathways, high- throughput genetic screens, protein purification, and biochemistry. The laboratory environment contains an extremely motivated and scientifically diverse group of researchers interested in fundamental aspects of C. elegans biology. The lab is flanked by two excellent C. elegans labs studying neurobiology and pathogenesis and is supported by a highly collaborative department that houses labs with expertise in RNA biology and chromatin structure. I plan to utilize all of these resources during the rest of my post-doctoral tenure. Throughout my research career, I have been focused on chromatin and RNA biology, which are fundamentally important in the understanding and prevention of cancer. Specifically, greater understanding of RNA silencing and transposon quiescence may lead to advances in cancer diagnoses or therapeutics. I am committed to a career in academic research and plan to start a lab focused on the aims in this proposal.