Chen Jenny, Shishkin Alexander A, Zhu Xiaopeng, Kadri Sabah, Maza Itay, Guttman Mitchell, Hanna Jacob H, Regev Aviv, Garber Manuel
Broad Institute of MIT and Harvard, Cambridge, MA, 02142, USA.
Division of Health Sciences and Technology, Massachusetts Institute of Technology, Cambridge, MA, 02140, USA.
Genome Biol. 2016 Feb 2;17:19. doi: 10.1186/s13059-016-0880-9.
Recent advances in transcriptome sequencing have enabled the discovery of thousands of long non-coding RNAs (lncRNAs) across many species. Though several lncRNAs have been shown to play important roles in diverse biological processes, the functions and mechanisms of most lncRNAs remain unknown. Two significant obstacles lie between transcriptome sequencing and functional characterization of lncRNAs: identifying truly non-coding genes from de novo reconstructed transcriptomes, and prioritizing the hundreds of resulting putative lncRNAs for downstream experimental interrogation.
We present slncky, a lncRNA discovery tool that produces a high-quality set of lncRNAs from RNA-sequencing data and further uses evolutionary constraint to prioritize lncRNAs that are likely to be functionally important. Our automated filtering pipeline is comparable to manual curation efforts and more sensitive than previously published computational approaches. Furthermore, we developed a sensitive alignment pipeline for aligning lncRNA loci and propose new evolutionary metrics relevant for analyzing sequence and transcript evolution. Our analysis reveals that evolutionary selection acts in several distinct patterns, and uncovers two notable classes of intergenic lncRNAs: one showing strong purifying selection on RNA sequence and another where constraint is restricted to the regulation but not the sequence of the transcript.
Our results highlight that lncRNAs are not a homogenous class of molecules but rather a mixture of multiple functional classes with distinct biological mechanism and/or roles. Our novel comparative methods for lncRNAs reveals 233 constrained lncRNAs out of tens of thousands of currently annotated transcripts, which we make available through the slncky Evolution Browser.
转录组测序的最新进展使得在许多物种中发现了数千种长链非编码RNA（lncRNA）。尽管已证明几种lncRNA在多种生物学过程中发挥重要作用，但大多数lncRNA的功能和机制仍不清楚。在lncRNA的转录组测序和功能表征之间存在两个重大障碍：从从头重建的转录组中识别真正的非编码基因，以及从数百个推定的lncRNA中确定优先顺序以进行下游实验研究。
我们展示了slncky，这是一种lncRNA发现工具，它从RNA测序数据中生成一组高质量的lncRNA，并进一步利用进化约束对可能具有重要功能的lncRNA进行优先级排序。我们的自动过滤流程与人工筛选效果相当，并且比以前发表的计算方法更敏感。此外，我们开发了一种用于比对lncRNA基因座的敏感比对流程，并提出了与分析序列和转录本进化相关的新进化指标。我们的分析表明，进化选择以几种不同的模式起作用，并揭示了两类值得注意的基因间lncRNA：一类在RNA序列上表现出强烈的纯化选择，另一类的约束仅限于转录本的调控而非序列。
我们的结果突出表明，lncRNA不是一类同质分子，而是多种功能类别的混合物，具有不同的生物学机制和/或作用。我们用于lncRNA的新型比较方法在数以万计的当前注释转录本中揭示了233个受约束的lncRNA，我们通过slncky进化浏览器提供这些信息。
