Ruan Lei, Yang Yi, Huang Yi, Ding Ling, Zhang Cuntai, Wu Xiaofen
Department of Gerontology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
Medicine (Baltimore). 2017 Aug;96(32):e7539. doi: 10.1097/MD.0000000000007539.
RAN guanine nucleotide release factor (RANGRF) encoding protein MOG1 plays an important role in cardiac arrhythmia, so we intended to investigate the regulatory miRNA of RANGRF and explore its potential regulatory mechanism in arrhythmogenesis.
Based on bioinformatic analysis, miR-3144-5p was predicted to be a regulatory miRNA of RANGRF, which were then validated through a dual-luciferase reporter plasmid assay. Subsequently, the expression level of miR-3144-5p in human cardiac myocytes (HCMs) was detected, followed by cell transfection with miR-3144-5p mimics. Transcriptome sequencing was then performed in HCMs with or without transfection. The sequencing results were subjected to bioinformatic analyses, including differentially expressed gene (DEG) analysis, functional enrichment analysis, protein-protein interaction (PPI) network analysis, miRNA-target gene analysis, and miRNA-transcription factor (TF)-target gene coregulatory network analysis.
There really existed a regulatory relation between miR-3144-5p and RANGRF. The expression level of miR-3144-5p was low in HCMs. After cell transfection, miR-3144-5p expression level significantly increased in HCMs. Bioinformatic analyses of the transcriptome sequencing results identified 300 upregulated and 271 downregulated DEGs between miR-3144-5p mimic and control group. The upregulated genes ISL1 and neuregulin 1 (NRG1) were significantly enriched in cardiac muscle cell myoblast differentiation (GO:0060379). CCL21 was one of the hub genes in the PPI network and also a target gene of miR-3144-5p. Moreover, the TF of v-Myc avian myelocytomatosis viral oncogene neuroblastoma-derived homolog (MYCN) was involved in the miR-3144-5p-TF-target gene coregulatory network and interacted with the target genes of miR-3144-5p.
ISL1, NRG1, CCL21, and MYCN were differentially expressed in the miR-3144-5p mimic group, suggesting that miR-3144-5p overexpression plays a role in HCMs by regulating these genes and TF. This study may provide new insight into the mechanisms behind the progression of cardiac arrhythmia.
编码RAN鸟嘌呤核苷酸释放因子（RANGRF）的蛋白质MOG1在心律失常中起重要作用，因此我们旨在研究RANGRF的调控性微小RNA（miRNA），并探讨其在心律失常发生中的潜在调控机制。
基于生物信息学分析，预测miR-3144-5p是RANGRF的调控性miRNA，随后通过双荧光素酶报告质粒检测进行验证。随后，检测人心脏心肌细胞（HCMs）中miR-3144-5p的表达水平，接着用miR-3144-5p模拟物转染细胞。然后对转染和未转染的HCMs进行转录组测序。对测序结果进行生物信息学分析，包括差异表达基因（DEG）分析、功能富集分析、蛋白质-蛋白质相互作用（PPI）网络分析、miRNA-靶基因分析以及miRNA-转录因子（TF）-靶基因共调控网络分析。
miR-3144-5p与RANGRF之间确实存在调控关系。HCMs中miR-3144-5p的表达水平较低。细胞转染后，HCMs中miR-3144-5p的表达水平显著升高。对转录组测序结果的生物信息学分析确定了miR-3144-5p模拟物组和对照组之间有300个上调和271个下调的DEG。上调基因ISL1和神经调节蛋白1（NRG1）在心肌细胞成肌细胞分化（GO:0060379）中显著富集。CCL21是PPI网络中的枢纽基因之一，也是miR-3144-5p的靶基因。此外，v-Myc禽成髓细胞瘤病毒癌基因神经母细胞瘤衍生同源物（MYCN）转录因子参与了miR-3144-5p-TF-靶基因共调控网络，并与miR-3144-5p的靶基因相互作用。
ISL1、NRG1、CCL21和MYCN在miR-3144-5p模拟物组中差异表达，表明miR-3144-5p过表达通过调控这些基因和转录因子在HCMs中发挥作用。本研究可能为心律失常进展背后的机制提供新的见解。
