纳米氧化铈暴露下大型溞miR-190和MARK2基因的表达和功能分析

Expression and Function Analysis of miR-190 and MARK2 Gene in Daphnia magna Exposed to Nano Cerium Dioxide

随着越来越多的纳米氧化铈(nCeO2)进入水环境,增加了水生生物与nCeO2接触的健康风险。为了研究微小RNA-190 (miR-190)的功能以及nCeO2暴露对大型溞体内miR-190及其靶基因MARK2表达的影响,我们利用miRbase数据库和MEGA7.0软件分析miR-190序列在各物种中的系统发生关系。Targetscan、miRanda和RNAhybrid筛选鉴定miR-190的靶基因。通过GO和KEGG聚类分析揭示miR-190靶基因的生物学作用。qPCR分析nCeO2暴露对大型溞体内miR-190和MARK2转录水平表达的影响。我们发现,14个物种的miR-190序列有16个碱基完全相同,miR-190序列在进化过程中高度保守。miR-190的852个靶基因主要位于细胞膜、细胞质中,富集在细胞内信号转导、DNA模板和转录调控、蛋白质磷酸化等生物学过程,且靶基因参与胞吞作用、ABC转运器和FoxO等细胞内重要的信号通路。在nCeO2暴露24、48 h后,miR-190在大型溞中的转录表达下调,而MARK2的mRNA表达量则明显上升。结果提示miR-190与MARK2在大型溞中的表达存在负调控关系,miR-190可能通过调控MARK2的转录表达来影响相关的细胞生命活动,本研究为解析nCeO2对水生无脊椎动物的毒性效应提供科学参考。

As more and more cerium oxide nanoparticles (nCeO2) enter the aquatic environment, the health risk of aquatic organisms exposed to nCeO2 increases. To investi-gate the function of microRNA-190 (miR-190) and the effect of nCeO2 exposure on the expression of miR-190 and its target gene MARK2 in Daphnia magna, we analyzed the phylogenetic relationships of miR-190 sequences across species using the miRbase database and MEGA 7.0 software. Tar-getscan, miRanda and RNA hybrid tools were performed to identify miR-190 targeted genes. Bio-logical roles of miR-190 target genes were revealed by GO and KEGG clustering analyses. qPCR analyzed the effects of nCeO2 exposure on the expression of miR-190 and MARK2 transcript levels in Daphnia magna. We found that the miR-190 sequences of 14 species were identical in 16 bases and that the miR-190 sequences were highly conserved during evolution. The 852 target genes of miR-190 are mainly located in the cell membrane and cytoplasm, and are enriched in biological processes such as intracellular signaling, DNA template and transcriptional regulation, protein phosphorylation, etc. Moreover, the target genes are involved in important intracellular signaling pathways such as cytosolization, ABC transporter and FoxO. After 24 and 48 h of nCeO2 exposure, the transcript expression of miR-190 was down-regulated in Daphnia magna, whereas the mRNA expression of MARK2 increased significantly. The results suggested that there was a negative regulatory relationship between miR-190 and MARK2 expression in Daphnia magna, and that miR-190 may affect related cellular life activities by regulating the transcriptional expression of MARK2. This study provides a scientific reference for analyzing the toxic effects of nCeO2 on aquatic invertebrates.