Integration of genome-wide approaches identifies lncRNAs of adult neural stem cells and their progeny in vivo [RNA CaptureSeq]

Long noncoding RNAs (lncRNAs) have been described in cell lines and various whole tissues, but lncRNA analysis of development in vivo is limited. Here, we comprehensively analyze lncRNA expression for the adult mouse subventricular zone neural stem cell lineage. We utilize complementary genome-wide techniques including RNA-seq, RNA CaptureSeq, and ChIP-seq to associate specific lncRNAs with neural cell types, developmental processes, and human disease states. By integrating data from chromatin state maps, custom microarrays, and FACS purification of the subventricular zone lineage, we stringently identify lncRNAs with potential roles in adult neurogenesis. shRNA-mediated knockdown of two such lncRNAs, Six3os and Dlx1as, indicate roles for lncRNAs in the glial-neuronal lineage specification of multipotent adult stem cells. Our data and workflow thus provide a uniquely coherent in vivo lncRNA analysis and form the foundation of a user-friendly online resource for the study of lncRNAs in development and disease. RNA CaptureSeq was perfomed on RNA extracted from adult mouse subventricular zone to better characterize rare lncRNA isoforms. Genomic regions used to design capture array are included in the raw file target_regions.bed.长链非编码RNA（Long noncoding RNAs，lncRNAs）已在细胞系及多种完整组织中被报道，但针对体内发育过程的长链非编码RNA分析仍较为有限。本研究针对成年小鼠脑室下区（subventricular zone）的神经干细胞谱系，全面解析了长链非编码RNA的表达模式。我们采用互补的全基因组技术，包括RNA测序（RNA-seq）、RNA捕获测序（RNA CaptureSeq）与染色质免疫共沉淀测序（ChIP-seq），将特定长链非编码RNA与神经细胞类型、发育过程及人类疾病状态相关联。通过整合染色质状态图谱、定制微阵列以及脑室下区谱系的荧光激活细胞分选（fluorescence-activated cell sorting，FACS）纯化数据，我们严格筛选出了可能在成年神经发生中发挥功能的长链非编码RNA。针对其中两种长链非编码RNA——Six3os与Dlx1as——进行的短发夹RNA（short hairpin RNA，shRNA）介导的敲低实验结果表明，长链非编码RNA在多能成体干细胞的胶质细胞-神经元谱系特化过程中发挥重要作用。本研究的数据与分析流程为体内长链非编码RNA分析提供了一套高度连贯的研究方案，并为开发用于研究发育与疾病相关长链非编码RNA的友好型在线资源奠定了基础。我们对从成年小鼠脑室下区提取的RNA进行了RNA捕获测序（RNA CaptureSeq），以更好地鉴定稀有长链非编码RNA剪接异构体。用于设计捕获阵列的基因组区域已包含在原始文件target_regions.bed中。