Bioinformatic Identification of miR-622 Key Target Genes and Experimental Validation of the miR-622-RNF8 Axis in Breast Cancer
Bioinformatic Identification of miR-622 Key Target Genes and Experimental Validation of the miR-622-RNF8 Axis in Breast Cancer
复制标题
乳腺癌中 miR-622 关键靶基因的生物信息学鉴定和 miR-622-RNF8 轴的实验验证
DOI：
10.3389/fonc.2019.01114
复制
发表时间：
2019-10-23
影响因子：
4.7
通讯作者：
Zhu, Lingyun
中科院分区：
文献类型：
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作者：
Liu, Chuanyang;Min, Lu;Zhu, Lingyun
乳腺癌是女性癌症相关死亡的主要原因。近几十年来，microRNAs(MiRNAs)是一种在转录后水平调节基因表达的短小非编码RNA，据报道参与了许多与肿瘤发生、肿瘤进展和转移相关的HUB基因的调控。然而，miRNAs调控乳腺癌转移的确切机制尚不清楚，这限制了开发新的、有效的治疗靶点的机会。在这里，我们的目标是确定癌症中miR-622相关的主要调控机制。首先，我们发现miR-622与各种癌症的不良预后显著相关。利用一个完整的miRNA预测过程，我们确定了77个有希望的靶点，并构建了一个蛋白质-蛋白质相互作用网络。此外，还进行了包括GO和KEGG途径分析在内的富集化分析，以确定miR-622的潜在功能，揭示了miR-622的调控网络和潜在功能。然后，我们在蛋白质-蛋白质相互作用网络中确定了一个由六个中枢基因组成的关键簇。基于TCGA和GEO数据集，进一步选择这些基因用于泛癌表达、预后和预测标记分析，以挖掘这些HUB基因的潜在临床价值。为了进一步验证我们的生物信息学结果，我们选择miR-622和RNF8的调节轴作为体外概念验证。miR-622和RNF8是最近报道的促进乳腺癌细胞EMT过程和乳腺癌转移的中枢基因之一。在体外，我们证实了miR-622对RNF8的直接调节，并发现所预测的miR-622-RNF8轴可以调节RNF8诱导的上皮-间充质转化、细胞迁移和细胞活力。这些结果通过救援实验得到了进一步证实。我们建立了一个闭环的miRNA-靶-表型研究模型，该模型集成了对miRNA靶基因的生物信息学分析和对所识别的关键miRNA-靶-表型轴的实验验证。我们不仅在计算机上鉴定了miR-622的HUB靶基因，而且首次揭示了miR-622在乳腺癌细胞EMT过程、存活和体外迁移中的调控机制。
Breast cancer is the leading cause of cancer-associated deaths among females. In recent decades, microRNAs (miRNAs), a type of short non-coding RNA that regulates gene expression at the post-transcription level, have been reported to participate in the regulation of many hub genes associated with tumorigenesis, tumor progression, and metastasis. However, the precise mechanism by which miRNAs regulate breast cancer metastasis remains poorly discussed, which limits the opportunity for the development of novel, effective therapeutic targets. Here, we aimed to determine the miR-622-related principal regulatory mechanism in cancer. First, we found that miR-622 was significantly related to a poor prognosis in various cancers. By utilizing an integrated miRNA prediction process, we identified 77 promising targets and constructed a protein-protein interaction network. Furthermore, enrichment analyses, including GO and KEGG pathway analyses, were performed to determine the potential function of miR-622, which revealed regulation networks and potential functions of miR-622. Then, we identified a key cluster comprised of six hub genes in the protein-protein interaction network. These genes were further chosen for pan-cancer expression, prognostic and predictive marker analyses based on the TCGA and GEO datasets to mine the potential clinical values of these hub genes. To further validate our bioinformatic results, the regulatory axis of miR-622 and RNF8, one of the hub genes recently reported to promote breast cancer cell EMT process and breast cancer metastasis, was selected as in vitro proof of concept. In vitro, we demonstrated the direct regulation of RNF8 by miR-622 and found that the predicted miR-622-RNF8 axis could regulate RNF8-induced epithelial-mesenchymal transition, cell migration, and cell viability. These results were further demonstrated with rescue experiments. We established a closed-loop miRNA-target-phenotype research model that integrated the bioinformatic analysis of the miRNA target genes and experimental validation of the identified key miRNA-target-phenotype axis. We not only identified the hub target genes of miR-622 in silico but also revealed the regulatory mechanism of miR-622 in breast cancer cell EMT process, viability, and migration in vitro for the first time.