A genomic approach to studying the life cycle of intron lariats

项目摘要/摘要 前-mRNA剪接是一个关键的和受调控的加工事件，其中内含子精确地 从新生的RNA转录本中摘录出来。多达三分之一的可遗传疾病突变 导致拼接缺陷。这项研究研究了分支点在决定剪接中的作用 在活体中利用了位置选择(3‘s)以及分支点对生命周期的影响 内含子。每个前mRNA剪接事件都会产生一个套索和剪接的外显子连接。当一个 人们对剪接外显子连接的了解很多，对连接蛋白几乎一无所知。通过 绘制人类基因组的所有分支点，我们正在开辟一个全新的领域 分析。通过文字记录数据确定分支点将有助于解释 临床测序数据。除了这些数据的内在价值外，成功完成 这一提议将检验一些关于基本催化的假设和认识 在处理真核基因的过程中发生在体内。在一个系统中研究这些中间体 广度将为数十万个加工过程带来生化水平的理解 事件。此外，每个内含子套索都有一个生命周期--通过剪接转录的 产品，通过脱枝和降解回收。内含子的循环对于 补充细胞内游离核苷酸水平并使剪接因子恢复活性 剪接体。一些内含子在作为非编码RNA(NcRNAs)剪接后有第二次生命。 当我们采样内含子的稳态水平时，我们对这两个过程都有了深入的了解。 这项提议试图通过探索某些内含子为什么 看起来很稳定。

Project Summary/Abstract Pre-mRNA splicing is a critical and regulated processing event where introns are precisely excised from nascent RNA transcripts. As many as one third of all heritable disease mutations result in splicing defects. This research studies the role of branchpoints in determining splice site selection (3'ss) is utilized in vivo and also the effect branchpoints have on the life cycle of the intron. Each pre-mRNA splicing event creates a lariat and spliced exon junction. While a great deal is known about splice exon junctions almost nothing is known about lariats. By mapping all branchpoints in the human genome, we are opening up a whole new area of analysis. The identification of branchpoints by transcript data will facilitate the interpretation of clinical sequencing data. In addition to the intrinsic value of this data, the successful completion of this proposal will test some hypothesis about the fundamental catalysis and recognition that occurs in vivo in the processing of eukaryotic genes. Studying these intermediates at a system wide level will bring a biochemical-level understanding to hundreds of thousands of processing events. Furthermore, each intron lariat has a lifecycle – created by splicing of a transcribed product, recycled by debranching and degradation. The recycling of introns is vital to replenishing the intracellular levels of free nucleotides and to return splicing factors into active spliceosomes. Some introns have a second life after splicing as non- coding RNAs (ncRNAs). As we are sampling steady state levels of introns we gain insight into both these processes. This proposal seeks to follow this lead by exploring some of the reasons why certain introns appear stabilized.