PDF(Pubmed)
Abstract：
RNA-seq has brought forth significant discoveries regarding aberrations in RNA processing, implicating these RNA variants in a variety of diseases. Aberrant splicing and single nucleotide variants (SNVs) in RNA have been demonstrated to alter transcript stability, localization, and function. In particular, the upregulation of ADAR, an enzyme that mediates adenosine-to-inosine editing, has been previously linked to an increase in the invasiveness of lung adenocarcinoma cells and associated with splicing regulation. Despite the functional importance of studying splicing and SNVs, the use of short-read RNA-seq has limited the community\'s ability to interrogate both forms of RNA variation simultaneously.
We employ long-read sequencing technology to obtain full-length transcript sequences, elucidating cis-effects of variants on splicing changes at a single molecule level. We develop a computational workflow that augments FLAIR, a tool that calls isoform models expressed in long-read data, to integrate RNA variant calls with the associated isoforms that bear them. We generate nanopore data with high sequence accuracy from H1975 lung adenocarcinoma cells with and without knockdown of ADAR. We apply our workflow to identify key inosine isoform associations to help clarify the prominence of ADAR in tumorigenesis.
Ultimately, we find that a long-read approach provides valuable insight toward characterizing the relationship between RNA variants and splicing patterns.
We employ long-read sequencing technology to obtain full-length transcript sequences, elucidating cis-effects of variants on splicing changes at a single molecule level. We develop a computational workflow that augments FLAIR, a tool that calls isoform models expressed in long-read data, to integrate RNA variant calls with the associated isoforms that bear them. We generate nanopore data with high sequence accuracy from H1975 lung adenocarcinoma cells with and without knockdown of ADAR. We apply our workflow to identify key inosine isoform associations to help clarify the prominence of ADAR in tumorigenesis.
Ultimately, we find that a long-read approach provides valuable insight toward characterizing the relationship between RNA variants and splicing patterns.
摘要：
RNA-seq带来了关于RNA加工中的畸变的重大发现，这些RNA变异与多种疾病有关。RNA中的异常剪接和单核苷酸变体（SNV）已被证明可以改变转录物的稳定性，本地化,和功能。特别是,ADAR的上调，一种介导腺苷到肌苷编辑的酶，先前已与肺腺癌细胞的侵袭性增加有关，并与剪接调节有关。尽管研究剪接和SNV的功能重要性，短读RNA-seq的使用限制了社区同时询问两种形式的RNA变异的能力。
我们采用长读测序技术来获得全长转录物序列，在单分子水平上阐明变体对剪接变化的顺式效应。我们开发了一个计算工作流程来增强FLAIR，调用以长读数据表示的同工型模型的工具，将RNA变体调用与携带它们的相关同种型整合在一起。我们从具有和不具有ADAR敲低的H1975肺腺癌细胞产生具有高序列准确性的纳米孔数据。我们应用我们的工作流程来确定关键的肌苷同工型关联，以帮助阐明ADAR在肿瘤发生中的重要性。
最终,我们发现长篇阅读方法为表征RNA变体和剪接模式之间的关系提供了有价值的见解。
我们采用长读测序技术来获得全长转录物序列，在单分子水平上阐明变体对剪接变化的顺式效应。我们开发了一个计算工作流程来增强FLAIR，调用以长读数据表示的同工型模型的工具，将RNA变体调用与携带它们的相关同种型整合在一起。我们从具有和不具有ADAR敲低的H1975肺腺癌细胞产生具有高序列准确性的纳米孔数据。我们应用我们的工作流程来确定关键的肌苷同工型关联，以帮助阐明ADAR在肿瘤发生中的重要性。
最终,我们发现长篇阅读方法为表征RNA变体和剪接模式之间的关系提供了有价值的见解。
