Key Laboratory of Organ Regeneration and Transplantation of Ministry of Education, Cancer Center, The First Hospital of Jilin University, Changchun, Jilin 130061, P.R. China.
Stanford University Medical School, VA Palo Alto Health Care System, Palo Alto, California 94304, USA.
Genome Res. 2019 Sep;29(9):1521-1532. doi: 10.1101/gr.244996.118. Epub 2019 Jul 17.
Long noncoding RNAs (lncRNAs) can regulate the activity of target genes by participating in the organization of chromatin architecture. We have devised a "chromatin-RNA in situ reverse transcription sequencing" (CRIST-seq) approach to profile the lncRNA interaction network in gene regulatory elements by combining the simplicity of RNA biotin labeling with the specificity of the CRISPR/Cas9 system. Using gene-specific gRNAs, we describe a pluripotency-specific lncRNA interacting network in the promoters of and , two critical stem cell factors that are required for the maintenance of pluripotency. The promoter-interacting lncRNAs were specifically activated during reprogramming into pluripotency. Knockdown of these lncRNAs caused the stem cells to exit from pluripotency. In contrast, overexpression of the pluripotency-associated lncRNA activated the promoters of core stem cell factor genes and enhanced fibroblast reprogramming into pluripotency. These CRIST-seq data suggest that the and promoters are organized within a unique lncRNA interaction network that determines the fate of pluripotency during reprogramming. This CRIST approach may be broadly used to map lncRNA interaction networks at target loci across the genome.
长链非编码 RNA（lncRNA）可以通过参与染色质结构的组织来调节靶基因的活性。我们设计了一种“染色质-RNA 原位反转录测序”（CRIST-seq）方法，通过将 RNA 生物素标记的简单性与 CRISPR/Cas9 系统的特异性相结合，来描绘基因调控元件中的 lncRNA 相互作用网络。使用基因特异性 gRNA，我们描述了在 和 启动子中存在的多能性特异性 lncRNA 相互作用网络，这两个关键的干细胞因子对于维持多能性是必需的。启动子相互作用的 lncRNA 在重编程为多能性的过程中特异性激活。这些 lncRNA 的敲低导致干细胞退出多能性。相比之下，多能相关 lncRNA 的过表达激活了核心干细胞因子基因的启动子，并增强了成纤维细胞向多能性的重编程。这些 CRIST-seq 数据表明， 和 启动子在一个独特的 lncRNA 相互作用网络中组织，该网络决定了重编程过程中多能性的命运。这种 CRIST 方法可以广泛用于绘制基因组中靶位点的 lncRNA 相互作用网络。
