Characterization of microRNA-offset RNAs in human embryonic stem cells (hESCs)

Small RNA molecules, including microRNAs (miRNAs), play critical roles in regulating pluripotency, proliferation and differentiation of embryonic stem cells. miRNA-offset RNAs (moRNAs) are similar in length to miRNAs, align to miRNA precursor (pre-miRNA) loci and are therefore believed to derive from processing of the pre-miRNA hairpin sequence. Recent next generation sequencing (NGS) studies have reported the presence of moRNAs in human neurons and cancer cells and in several tissues in mouse, including pluripotent stem cells. In order to gain additional knowledge about human moRNAs and their putative development-related expression, we applied NGS of small RNAs in human embryonic stem cells (hESCs) and fibroblasts. We found that certain moRNA isoforms are notably expressed in hESCs from loci coding for stem cell-selective or cancer-related miRNA clusters. In contrast, we observed only sparse moRNAs in fibroblasts. Consistent with earlier findings, most of the observed moRNAs derived from conserved loci and their expression did not appear to correlate with the expression of the adjacent miRNAs. We provide here the first report of moRNAs in hESCs, and their expression profile in in comparison to fibroblasts. Moreover, we expand the repertoire of hESC miRNAs. These findings provide an expansion on the known repertoire of small non-coding RNA contents in hESCs. Overall design: We performed small RNA-seq analysis from three samples: one from hESC line HS401, one from hESC line HS181 and one from human foreskin fibroblast line HFF-1小分子RNA，包括微RNA（microRNAs, miRNAs），在调控胚胎干细胞的多能性、增殖与分化过程中发挥关键作用。miRNA偏移RNA（miRNA-offset RNAs, moRNAs）长度与miRNAs相近，可比对至miRNA前体（pre-miRNA）基因座，因此被认为源自pre-miRNA发夹结构的加工过程。近年来的新一代测序（next generation sequencing, NGS）研究已在人类神经元、癌细胞以及包括多能干细胞在内的小鼠多种组织中报道了moRNAs的存在。为进一步解析人类moRNAs及其潜在的发育相关表达模式，我们对人类胚胎干细胞（human embryonic stem cells, hESCs）和成纤维细胞开展了小分子RNA的NGS测序分析。研究发现，部分moRNA异构体在源自干细胞选择性或癌症相关miRNA簇基因座的hESCs中显著表达。与之相反，我们仅在成纤维细胞中检测到少量moRNAs。与此前研究结果一致，绝大多数检测到的moRNAs源自保守基因座，且其表达水平与邻近miRNAs的表达并无明显关联。本研究首次报道了hESCs中的moRNAs，并对比分析了其与成纤维细胞的表达谱。此外，本研究还拓展了hESC的miRNA已知集合。上述研究结果进一步丰富了人们对hESCs中小非编码RNA组成的认知。实验设计概述：我们对三个样本开展了小分子RNA测序分析，样本分别来自hESC细胞系HS401、hESC细胞系HS181以及人包皮成纤维细胞系HFF-1