The DNMT1 associated lncRNA Dali is an epigenetic regulator of neural differentiation [4]. Mus musculus

Many intergenic long noncoding RNA (lncRNA) loci regulate the expression of adjacent protein coding genes. Less clear is whether intergenic lncRNAs commonly regulate transcription by modulating chromatin at genomically distant loci. Here, we report both genomically local and distal RNA-dependent roles of Dali, a conserved central nervous system expressed intergenic lncRNA. Dali is transcribed downstream of the Pou3f3 transcription factor gene and its depletion disrupts the differentiation of neuroblastoma cells. Locally, Dali transcript regulates transcription of the Pou3f3 locus. Distally, it preferentially targets active promoters and regulates expression of neural differentiation genes, in part through physical association with the POU3F3 protein. Dali interacts with the DNMT1 DNA methyltransferase in mouse and human and regulates DNA methylation status of CpG island-associated promoters in trans. These results demonstrate, for the first time, that a single intergenic lncRNA controls the activity and methylation of genomically distal regulatory elements to modulate large-scale transcriptional programmes. Overall design: The genome-wide binding profile of Dali was determined using Capture Hybridisation Analysis of RNA Targets (CHART)-Seq technique (Simon et al, Proc Natl Acad Sci U S A 108: 20497-20502, 2011) in N2A cells. Following the CHART-seq protocol, we used RNase H elution to recover genomic DNA associated with endogenous Dali transcripts. We identified Dali binding sites in comparison both to an input DNA sample from N2A cells and to DNA recovered using the control E.coli LacZ oligonucleotide. Please note that n2a_input, n2a_LacZ data was published previously [GSE52571]

众多基因间长链非编码RNA（intergenic long noncoding RNA, lncRNA）位点可调控邻近蛋白编码基因的表达。目前尚不明确的是，基因间lncRNA是否普遍通过调控基因组远端位点的染色质状态来调控转录过程。本研究针对保守表达于中枢神经系统的基因间lncRNA Dali，揭示了其同时具备基因组局部及远端RNA依赖性功能。Dali转录于Pou3f3转录因子基因的下游区域，其表达敲除会破坏神经母细胞瘤细胞的分化过程。局部层面，Dali转录本可调控Pou3f3基因座的转录。远端层面，Dali优先靶向活性启动子并调控神经分化相关基因的表达，其部分机制依赖于与POU3F3蛋白的物理结合。Dali可在小鼠及人类细胞中与DNMT1 DNA甲基转移酶相互作用，并通过反式调控方式调控CpG岛相关启动子的DNA甲基化状态。本研究结果首次证实，单个基因间lncRNA可通过调控基因组远端调控元件的活性与甲基化状态，进而调控大规模转录程序。整体实验设计：本研究采用RNA靶标捕获杂交分析测序（Capture Hybridisation Analysis of RNA Targets, CHART-seq）技术，在N2A细胞中解析了Dali的全基因组结合图谱（引用文献：Simon等，《美国国家科学院院刊》，2011年，第108卷，20497-20502页）。按照CHART-seq实验流程，本研究采用核糖核酸酶H（RNase H）洗脱法回收与内源性Dali转录本结合的基因组DNA。通过与N2A细胞的输入DNA样本、以及使用对照大肠杆菌LacZ寡核苷酸回收的DNA进行比对，研究团队鉴定出Dali的结合位点。请注意，n2a_input与n2a_LacZ的相关数据已在前期研究中发表[GSE52571]