Identification and initial functional characterization of a human vascular cell-enriched long noncoding RNA.
Identification and initial functional characterization of a human vascular cell-enriched long noncoding RNA.
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DOI：
10.1161/atvbaha.114.303240
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发表时间：
2014-06
期刊：
影响因子：
--
通讯作者：
Miano JM
中科院分区：
文献类型：
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作者：
Bell RD;Long X;Lin M;Bergmann JH;Nanda V;Cowan SL;Zhou Q;Han Y;Spector DL;Zheng D;Miano JM
长链非编码RNA （lncRNAs）是一类快速增长的RNA基因，其功能主要与基因表达的转录和转录后控制有关。关于lncRNA在人血管细胞中的表达和功能的信息缺乏。因此，我们开始在人类血管平滑肌细胞中鉴定新的lncRNA基因，并深入了解它们在平滑肌细胞表型控制中的作用。人类冠状动脉平滑肌细胞的RNA测序显示了31个未注释的lncRNA，包括一个血管细胞富集的lncRNA，我们称之为SENCR（平滑肌和内皮细胞富集迁移/分化相关的长链非编码RNA）。链特异性RT-PCR和cDNA末端的快速扩增表明，senr是从FLI1基因5 '端反义转录而来，并以两个剪接变异体的形式存在。RNA荧光原位杂交和生化分离研究表明，SENCR是一种细胞质lncRNA。与这一观察结果一致，敲低研究显示senr对FLI1或邻近基因表达几乎没有顺式作用作用。senr敲低后平滑肌细胞的rna测序实验显示，心肌素和许多平滑肌收缩基因的表达减少，而一些促迁移基因的表达增加。RT-PCR和Western blotting实验验证了SENCR敲低后的几个差异表达基因。抓伤的功能丧失研究和Boyden室实验支持senr作为平滑肌细胞迁移的抑制剂。SENCR是一种新的富含血管细胞的细胞质lncRNA，似乎可以稳定平滑肌细胞的收缩表型。
Long non-coding RNAs (lncRNAs) represent a rapidly growing class of RNA genes with functions related primarily to transcriptional and post-transcriptional control of gene expression. There is a paucity of information about lncRNA expression and function in human vascular cells. Thus, we set out to identify novel lncRNA genes in human vascular smooth muscle cells and to gain insight into their role in the control of smooth muscle cell phenotypes. RNA-sequencing of human coronary artery smooth muscle cells revealed 31 unannotated lncRNAs, including a vascular cell-enriched lncRNA we call SENCR (Smooth muscle and Endothelial cell enriched migration/differentiation-associated long Non-Coding RNA). Strand-specific RT-PCR and rapid amplification of cDNA ends indicate that SENCR is transcribed antisense from the 5’ end of the FLI1 gene and exists as two splice variants. RNA fluorescence in situ hybridization and biochemical fractionation studies demonstrate SENCR is a cytoplasmic lncRNA. Consistent with this observation, knockdown studies reveal little to no cis-acting effect of SENCR on FLI1 or neighboring gene expression. RNA-sequencing experiments in smooth muscle cells following SENCR knockdown disclose decreased expression of Myocardin and numerous smooth muscle contractile genes, while a number of pro-migratory genes are increased. RT-PCR and Western blotting experiments validate several differentially expressed genes following SENCR knockdown. Loss-of-function studies in scratch wound and Boyden chamber assays support SENCR as an inhibitor of smooth muscle cell migration. SENCR is a new vascular cell-enriched, cytoplasmic lncRNA that appears to stabilize the smooth muscle cell contractile phenotype.