Bioinformatics analysis and experimental validation of microRNA-140-3p targeting cell division cycle associated 8 inhibiting invasion and metastasis of lung adenocarcinoma cells

ZHENG Quan; HU Ya-Qiong; BAI Jun; YIN Chong-Gao; LI Hong-Li; LIU Yu-Qing

doi:10.16098/j.issn.0529-1356.2021.04.014

PDF(18830 KB)

Acta Anatomica Sinica ›› 2021, Vol. 52 ›› Issue (4) : 589-600. DOI: 10.16098/j.issn.0529-1356.2021.04.014

Bioinformatics analysis and experimental validation of microRNA-140-3p targeting cell division cycle associated 8 inhibiting invasion and metastasis of lung adenocarcinoma cells

ZHENG Quan¹ HU Ya-qiong¹ BAI Jun¹ YIN Chong-gao² LI Hong-li³ LIU Yu-qing^1*#br#

Author information +

History +

Abstract

Objective To investigate the effect of microRNA(miR)-140-3p targeting cell division cycle associated 8(CDCA8) on invasion and metastasis of lung adenocarcinoma cells. Methods The differentially expressed miRNAs were analyzed by GEO2R in GEO database. The target genes of miR-140-3p were searched by TargetScan human7.2 and miRWalk databases. The hub gene was screened by Cytoscape 3.7.2 software. GEPIA database was used to query the expression levels of target gene in lung adenocarcinoma tissues and normal lung tissues, the expression levels in different stages of lung adenocarcinoma, and the relationship between the expression levels of target gene and the overall survival rate of lung adenocarcinoma patients. The survival analysis of mi-140-3p in lung adenocarcinoma and the correlation between miR-140-3p and CDCA8 expression levels were searched in starBase database. Real-time PCR was used to detect the expression levels of miR-140-3p in normal lung epithelial cells BEAS-2B and lung adenocarcinoma cells A549, as well as the efficiency of infection. Expression levels of CDCA8 mRNA and protein were detected by Real-time PCR and Western blotting experiments after overexpresion of miR-140-3p. Dual-luciferase reporter assay verified whether miR-140-3p directly binds to CDCA8. Transwell invasion assay detected the effect of overexpression of miR-140-3p and CDCA8 on the invasiveness of lung adenocarcinoma cells. Results Analysis result from GEO and other databases showed that the expression level of miR-140-3p in normal lung tissues was significantly higher than that in lung adenocarcinoma, and its predicted target gene CDCA8 expression level in lung adenocarcinoma was significantly higher than that in normal lung tissues, and CDCA8 was negatively correlated with the expression level of miR-140-3p in lung adenocarcinoma. The experimental result showed that the expression of miR-140-3p in A549 cells was significantly lower than that in BEAS-2B cells (P＜0.05). The expression level of miR-140-3p increased significantly after lentiviral infection (P＜0.05). CDCA8 mRNA and protein expression levels were significantly down-regulated after overexpression of miR-140-3p (P＜0.05). Dual-luciferase reporter assay result showed that miR-140-3p could directly bind to CDCA8 (P＜0.05). Compared with the control group, overexpression of miR-140-3p inhibited the invasion and metastasis of lung adenocarcinoma A549, while CDCA8 revrsed the inhibition of miR-140-3p on the invasion and metastasis of lung adenocarcinoma A549 (P＜0.05). Conclusion MiR-140-3p targeting CDCA8 inhibits the invasion and metastasis of lung adenocarcinoma cells.

Key words

Lung adenocarcinoma / MicroRNA-140-3p / Cell division cycle associated 8 / Invasion / Metastasis / Western blotting

Cite this article

EndNote

Ris (Procite)

Bibtex

Download Citations

ZHENG Quan HU Ya-qiong BAI Jun YIN Chong-gao LI Hong-li LIU Yu-qing. Bioinformatics analysis and experimental validation of microRNA-140-3p targeting cell division cycle associated 8 inhibiting invasion and metastasis of lung adenocarcinoma cells[J]. Acta Anatomica Sinica. 2021, 52(4): 589-600 https://doi.org/10.16098/j.issn.0529-1356.2021.04.014

References

［1］ Seijo LM, Peled N, Ajona D, et al. Biomarkers in lung cancer screening: achievements, promises, and challenges［J］. J Thorac Oncol,2019,14(3):343-357.

［2］ Wang J, Jia Y, Zhao S, et al. BIN1 reverses PD-L1-mediated immune escape by inactivating the c-MYC and EGFR/MAPK signaling pathways in nonsmall cell lung cancer［J］. Oncogene,2017,36(45):6235-6243.

［3］ Garzon R, Calin GA, Croce CM. MicroRNAs in cancer［J］. Annu Rev Med,2009,60(1):167-179.

［4］ Fan BL，Ji XH，Zhu WL. Effects of miR-100 on mitotic arrest and Polo-like kinase 1 protein expression in hepatocellular carcinoma cells［J］. Acta Acatomica Sinica,2014,45(1):70-73 (in Chinese)

范秉琳，嵇晓辉，朱武凌. MiR-100对肝癌细胞有丝分裂阻滞及Polo样激酶1蛋白表达的作用［J］. 解剖学报,2014,45(1):70-73.

［5］ Raponi M, Dossey L, Jatkoe T, et al. MicroRNA classifiers for predicting prognosis of squamous cell lung cancer［J］. Cancer Res,2009,69(14):5776-5783.

［6］ Kumar MS, Erkeland S , Pester RE, et al. Suppression of non-small cell lung tumor development by the let-7 microRNA family［J］. Proc Natl Acad Sci USA,2008,105(10):3903-3908.

［7］ Phan NN, Wang CY, Li KL, et al. Distinct expression of CDCA3, CDCA5, and CDCA8 leads to shorter relapse free survival in breast cancer patient［J］. Oncotarget,2018,9(6):6977-6992.

［8］ Chang JL, Chen TH, Wang CF, et al. Borealin/Dasra B is a cell cycle-regulated chromosomal passenger protein and its nuclear accumulation is linked to poor prognosis for human gastric cancer［J］. Exp Cell Res,2006,312(7):962-973.

［9］ Hayama S, Daigo Y, Yamabuki T, et al. Phosphorylation and activation of cell division cycle associated 8 by aurora kinase B plays a significant role in human lung carcinogenesis［J］. Cancer Res,2007,67(9):4113-4122.

［10］ Garber ME, Troyanskaya OG, Schluens K, et al. Diversity of gene expression in adenocarcinoma of the lung［J］. Proc Natl Acad Sci USA,2001,98(24):13784-13789.

［11］ Landi MT, Dracheva T, Rotunno M, et al. Gene expression signature of cigarette smoking and its role in lung adenocarcinoma development and survival［J］. PLoS One,2008,3(2):e1651.

［12］ Hou J, Aerts J, den Hamer B, et al. Gene expression-based classification of non-small cell lung carcinomas and survival prediction［J］. PLoS One,2010,5(4): e10312.

［13］ Selamat SA, Chung BS, Girard L, et al. Genome-scale analysis of DNA methylation in lung adenocarcinoma and integration with mRNA expression［J］. Genome Res,2012,22(7):1197-1211.

［14］ Okayama H, Kohno T, Ishii Y, et al. Identification of genes upregulated in ALK-positive and EGFR/KRAS/ALK-negative lung adenocarcinomas［J］. Cancer Res,2012,72(1):100-111.

［15］ Zhang GX，Xu XW，Meng B，et al. MiR-186-5p inhibits epithelial mesenchymal transition in lung adenocarcinoma cells through modulating PTTG1［J］. Chinese Journal of Biochemistry and Molecular Biology, 2017, 33(4): 380-385 (in Chinese)

张国新，徐新伟，孟斌，等. MiR-186-5p通过靶向调控PTTG1抑制肺腺癌细胞的上皮-间质转化［J］. 中国生物化学与分子生物学报,2017,33(4):380-385.

［16］ Zhang B, Li H, Yin C, et al. Dock1 promotes the mesenchymal transition of glioma and is modulated by MiR-31［J］. Neuropathol Appl Neurobiol,2017,43(5):419-432.

［17］ Yin C, Mou Q, Pan X, et al. MiR-577 suppresses epithelial-mesenchymal transition and metastasis of breast cancer by targeting Rab25［J］. Thorac Cancer,2018,9(4):472-479.

［18］ Vasudevan S, Tong Y, Steitz JA. Switching from repression to activation: microRNAs can up-regulate translation［J］. Science,2007,318(5858):1931-1934.

［19］ Kai Y, Peng W, Ling W, et al. Reciprocal effects between microRNA-140-5p and ADAM10 suppress migration and invasion of human tongue cancer cells［J］. Biochem Biophys Res Commun,2014,448(3):308-314.

［20］ Yuan Y, Shen Y, Xue L, et al. miR-140 suppresses tumor growth and metastasis of non-small cell lung cancer by targeting insulin-like growth factor 1 receptor［J］. PLoS one,2013,8(9):e73604.

［21］ Ma J, Zhang F, Sun P. miR-140-3p impedes the proliferation of human cervical cancer cells by targeting RRM2 to induce cell-cycle arrest and early apoptosis［J］. Bioorg Med Chem,2020,28(3):115283.

［22］ Zhou Y, Wang B, Wang Y, et al. miR-140-3p inhibits breast cancer proliferation and migration by directly regulating the expression of tripartite motif 28［J］. Oncol Lett,2019,17(4):3835-3841.

［23］ Dong W, Yao C, Teng X, et al. MiR-140-3p suppressed cell growth and invasion by downregulating the expression of ATP8A1 in non-small cell lung cancer［J］. Tumour Biol,2016,37(3):2973-2985.

［24］ Liu ZhJ, Dou LT, Zhuang Zh, et al. Screening and survival analysis of differentially expressed genes in papillary thyroid carcinoma［J］. Acta Anatomica Sinica, 2020,51(1),51-57. (in Chinese)

刘中娇，窦龙涛，庄振，等. 乳头状甲状腺癌差异表达基因的筛选及生存分析［J］. 解剖学报,2020,51(1):51-57.

［25］ Kong X, Zhang G, Huo Y, et al. MicroRNA-140-3p inhibits proliferation, migration and invasion of lung cancer cells by targeting ATP6AP2［J］. Int J Clin Exp Pathol,2015,8(10):12845.

［26］ Ci C, Tang B, Lyu D, et al. Overexpression of CDCA8 promotes the malignant progression of cutaneous melanoma and leads to poor prognosis［J］. Int Jo Mol Med,2019,43(1):404-412.

［27］ Wang Y, Zhao Z, Bao X, et al. Borealin/Dasra B is overexpressed in colorectal cancers and contributes to proliferation of cancer cells［J］. Med Oncol,2014,31(11):248.

［28］ Bi Y, Chen S, Jiang J, et al. CDCA8 expression and its clinical relevance in patients with bladder cancer［J］. Medicine,2018,97(34): e11899.