Abstract

Non-coding RNAs (ncRNAs) are powerful regulators of gene expression but medium-sized (50-300 nts in length) ncRNAs (msRNAs) are barely picked-up precisely by RNA-sequencing. Here we describe msRNA-sequencing (msRNAseq), a modified protocol that associated with a computational analyses pipeline identified about ~1800 msRNA loci, including over 300 putatively novel msRNAs, in human and murine cells. We focused on the identification and initial characterization of three POLIII-derived transcripts. The validation of these uncharacterized msRNAs identified an ncRNA in antisense orientation from the POLR3E locus transcribed by POLIII. This msRNA, termed POLAR (POLR3E Antisense RNA), has a strikingly short half-life, localizes to paraspeckles (PSPs) and associates with PSP-associated proteins indicating that msRNAseq identifies functional msRNAs. Thus, our analyses will pave the way for analyzing the roles of msRNAs in cells, development and diseases.

摘要

非编码RNA（ncRNA）是基因表达的有力调节剂，但是中等大小的ncRNA（msRNAs）长度仅为50-300 nt，而RNA测序几乎不能精确地将其拾取。在这里，我们描述了msRNA测序（msRNAseq），这是一种经过修改的协议，与计算分析管道相关联，可识别人和鼠类细胞中约1800个msRNA位点，包括300多个假定的新型msRNA。我们专注于三种POLIII衍生的转录物的鉴定和初步表征。这些未表征的msRNA的验证从POLIII转录的POLR3E基因座中以反义方向鉴定了ncRNA 。此msRNA，称为POLAR（POL R3E甲ntisense řNA），其半衰期非常短，位于散斑（PSP）上，并与PSP相关蛋白缔合，表明msRNAseq可以识别功能性msRNA。因此，我们的分析将为分析msRNA在细胞，发育和疾病中的作用铺平道路。