Integrated analysis of miRNA and mRNA expression profiles in p53-edited PFF cells
Integrated analysis of miRNA and mRNA expression profiles in p53-edited PFF cells
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p53 编辑的 PFF 细胞中 miRNA 和 mRNA 表达谱的综合分析
DOI：
10.1080/15384101.2020.1742852
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发表时间：
2020-03
期刊：
影响因子：
4.3
通讯作者：
Yuyun Xing
中科院分区：
文献类型：
--
作者：
Chuanmin Qiao;Weiwei Liu;Haoyun Jiang;Maozhang He;Qiang Yang;Yuyun Xing
摘要p53基因是人类肿瘤中最常发生突变的基因，超过一半的肿瘤在该位点发生突变。R248和R249（对应于猪R241和R242）是在肝、肺、乳腺和一些其他癌症中频繁突变的热点突变之一。在这项研究中，通过CRISPR-Cas9技术在猪胎儿成纤维细胞（PFF）中敲除或点编辑（R241和R242转化为241 W和242 S）p53基因。miRNA和mRNA的高通量测序揭示了p53敲除（p53-KO）细胞中总共225个差异表达的miRNA（DEM）和738个差异表达的基因（DEG），以及点修饰（p53- 241 W242 S）细胞中总共211个DEM和722个DEG。p53-KO/p53-WT和p53- 241 W242 S/p53-WT miRNAs数据集共有28个标注的DEM存在重叠，其中miR-34 c、miR-218、miR-205、miR-105-1、miR-105-2、miR-206、miR-224和miR-429在p53调控网络中起重要作用。在p53-KO和p53- 241 W242 S细胞中的前10个DEG中，大多数基因被报道参与肿瘤、细胞增殖或细胞迁移。p53-KO和p53- 241 W242 S细胞的增殖率明显高于p53-WT细胞（P < 0.01）。总之，p53基因的遗传修饰显著影响PFF细胞中大量基因和miRNAs的表达水平。p53基因编辑的PFF细胞可作为研究p53信号网络的非肿瘤细胞模型，并可作为体细胞核移植的供体细胞，目的是建立具有相应p53突变的猪模型。缩略语：CRISPR-Cas9：规则间隔短回文重复序列相关蛋白9; PFF：猪胎儿成纤维细胞; SCNT：体细胞核移植; RNA测序：小RNA测序和mRNA测序; DEG：差异表达的mRNA; DEM：差异表达的miRNA。
ABSTRACT p53 is the most frequently mutated gene in human cancers, with over half of all tumors harboring mutation at this locus. R248 and R249 (corresponding to porcine R241 and R242), are among the hotspot mutations frequently mutated in liver, lung, breast, and some other cancers. In this study, p53 gene was knocked out or point-edited (R241 and R242 were converted to 241W and 242S) in porcine fetal fibroblast (PFF) cells via CRISPR-Cas9 technique. High throughput sequencing of miRNA and mRNA uncovered a total of 225 differentially expressed miRNAs (DEMs) and 738 differentially expressed genes (DEGs) in the p53 knockout (p53-KO) cells, and a total of 211 DEMs and 722 DEGs in the point-modified (p53-241W242S) cells. Totally 28 annotated DEMs were found to overlap between p53-KO/p53-WT and p53-241W242S/p53-WT miRNAs datasets, of which miR-34 c, miR-218, miR-205, miR-105-1, miR-105-2, miR-206, miR-224 and miR-429 play important roles in p53 regulatory network. Among the top 10 DEGs in p53-KO and p53-241W242S cells, most genes were reported to be involved in tumors, cell proliferation or cell migration. p53-KO and p53-241W242S cells showed a significantly higher (P < 0.01) proliferation rate compared with p53-WT cells. In conclusion, genetic modifications of p53 gene significantly affect the expression levels of a large number of genes and miRNAs in the PFF cells. The p53-edited PFF cells could be used as non-tumor cell models for investigating the p53 signaling network, and as donor cells for somatic nuclear transfer, with the aim to develop porcine models with the corresponding p53 mutations. Abbreviations: CRISPR-Cas9: Clustered regularly interspaced short palindromic repeats-associated protein 9; PFF: porcine fetal fibroblasts; SCNT: somatic cell nuclear transfer; RNA sequencing: small RNA sequencing and mRNA sequencing; DEGs: differentially expressed mRNAs; DEMs: differentially expressed miRNAs.
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