人支气管上皮HBE细胞中p53相关的放射诱导表达长链非编码RNA的鉴定和生物功能预测分析  

Identification and bioinformatic function analysis of p53-related differentially expressed lncRNAs in γ-rays irradiatedhuman bronchial epithelial HBE cells

作　　者：胡赛 黄瑞雪[2] 周平坤 黄波[1] HU Sai;HUANG Rui-xue;ZHOU Ping-kun;HUANG Bo(School of Public Health,University of South China,Hengyang 421001,China;School of Public Health,Central South University,Changsha 410078,China;Beijing Key Laboratory for Radiobiology,Institute of Radiation Medicine,Academy of Military Medical Sciences,Academy of Military Sciences,Beijing 100850,China)

机构地区：[1]南华大学公共卫生学院,湖南衡阳421001 [2]中南大学公共卫生学院劳动卫生与环境卫生学系,湖南长沙410078 [3]军事科学院军事医学研究院辐射医学研究所,北京市放射生物学重点实验室,北京100850

出　　处：《中国药理学与毒理学杂志》2019年第8期587-594,共8页Chinese Journal of Pharmacology and Toxicology

基　　金：国家自然科学基金（U1803124）~~

摘　　要：目的鉴定人支气管上皮HBE细胞中p53相关的γ射线诱导表达的长链非编码RNA(lncRNA)分子,为研究与p53关联的lncRNA分子在放射损伤反应如上皮间充质转化中的作用机制提供信息。方法使用CRISPR-Cas9基因编辑技术构建p53敲除的人支气管上皮细胞(HBEp53-/-),Western印迹法检测P53表达水平,lncRNA芯片杂交分析lncRNA表达谱的改变,实时定量RT-PCR分析验证部分lncRNA分子的表达变化,生物信息学技术分析差异变化lncRNA的生物学功能。结果与野生型HBE细胞相比,HBEp53-/-细胞在照射后有239条lncRNA出现上调表达,287条出现下调表达。上调和下调表达前5位lnc RNA的差异表达变化经PCR得到验证。生物信息学GO分析表明,前10位差异表达的lncRNA的靶基因主要涉及β3肾上腺素受体结合蛋白、酪氨酸激酶活性蛋白、DNA结合蛋白、锌离子跨膜转运体蛋白、突触融合蛋白结合蛋白和泛素结合蛋白等分子功能。KEGG分析表明,差异表达lnc RNA的靶基因功能的生物体系统主要涉及感知、神经、免疫、内分泌、环境适应、发育和循环系统等。结论鉴定了人支气管上皮HBE细胞中与p53相关的放射诱导lncRNA分子;经生物信息学预测,发现这些lncRNA分子在细胞照射应答过程中,涉及多种生理功能,参与多个功能信号通路。OBJECTIVE To identify the differentially expressed long non-coding RNAs(lncRNAs)inγ-rays irradiated p53 knockout HBE(HBEp53-/-)cells,and to provide more baseline information for further investigation of p53 related lncRNAs in irradiation-induced injury,such as epithelial-mesenchymal transition in lung tissue epithelial cells.METHODS HBEp53-/-cells were established with CRISPR-cas9.Western blotting was used to detect the knockout efficacy of P53 protein,while real time RT-PCR was used to verify the selected expressions of lncRNAs.Bioinformatic analysis including GO and KEGG analysis was used to predict the lncRNA function and signaling pathway.RESULTS Compared to the irradiated HBE cells,239 lncRNAs were up-regulated and 289 down-regulated in irradiated HBEp53-/-cells.The top five of the up-regulated or down-regulated differentially expressed lncRNAs were further confirmed by RT-PCR analysis.Through bioinformatic prediction GO analysis,the targeted genes of lncRNAs with changed expressions were related to multiple biological processes and molecular functions,includingβ3 adrenergic receptor binding,protein tyrosine kinase activity,DNA binding,zic ion transmembrane transporter activity,ubiquitin binding and syntaxin binding.KEGG analysis revealed that the functional organismal systems involved included the sensory,nervous,immune,endocrine,environmental adaption,developmental,and circulatory systems.CONCLUSION p53-related radiation-inducible lncRNAs have been identified in HBEp53-/-cells.These identified lncRNAs may play critical roles in various biological processes and functional signaling pathways in cellular response to radiation injury.

关 键 词：Γ射线 长链非编码RNA P53 支气管上皮细胞 

分 类 号：Q691[生物学—生物物理学] R394-33[医药卫生—医学遗传学]