Detecting actively translated open reading frames in ribosome profiling data
Detecting actively translated open reading frames in ribosome profiling data
复制标题
DOI：
10.1038/nmeth.3688
复制
发表时间：
2016-02-01
期刊：
影响因子：
48
通讯作者：
Ohler, Uwe
中科院分区：
文献类型：
--
作者：
Calviello, Lorenzo;Mukherjee, Neelanjan;Ohler, Uwe
RNA 测序方案可以量化从转录到蛋白质合成的基因表达调控。核糖体分析 (Ribo-seq) 绘制整个转录组中翻译核糖体的位置。我们开发了 RiboTaper（可在 https://ohlerlab.mdc-berlin.de/software/ 获取），这是一种严格的统计方法，可根据 Ribo-seq 数据的特征性三核苷酸周期性来识别翻译区域。我们使用 RiboTaper 和 HEK293 细胞的深度 Ribo-seq 数据得出广泛的翻译图谱，涵盖超过 11,000 个蛋白质编码基因的开放阅读框 (ORF) 注释。我们还发现了数百个上游 ORF 和带注释的非编码基因 (ncORF) 中 ORF 的不同核糖体特征。质谱数据证实 RiboTaper 实现了细胞蛋白质组的出色覆盖。尽管以这种方式验证了数十种新型肽产品，但目前注释的长非编码 RNA 中很少有似乎编码稳定的多肽。 RiboTaper 是一种强大的方法，可从 Ribo-seq 数据中全面从头识别常用的 ORORF。
RNA-sequencing protocols can quantify gene expression regulation from transcription to protein synthesis. Ribosome profiling (Ribo-seq) maps the positions of translating ribosomes over the entire transcriptome. We have developed RiboTaper (available at https://ohlerlab.mdc-berlin.de/software/), a rigorous statistical approach that identifies translated regions on the basis of the characteristic three-nucleotide periodicity of Ribo-seq data. We used RiboTaper with deep Ribo-seq data from HEK293 cells to derive an extensive map of translation that covered open reading frame (ORF) annotations for more than 11,000 protein-coding genes. We also found distinct ribosomal signatures for several hundred upstream ORFs and ORFs in annotated noncoding genes (ncORFs). Mass spectrometry data confirmed that RiboTaper achieved excellent coverage of the cellular proteome. Although dozens of novel peptide products were validated in this manner, few of the currently annotated long noncoding RNAs appeared to encode stable polypeptides. RiboTaper is a powerful method for comprehensive de novo identification of actively used ORORFs from Ribo-seq data.