Eya2 overexpression promotes the invasion of human astrocytoma through the regulation of ERK/MMP9 signaling

Wen,Zhifeng; Liang,Chuansheng; Pan,Qichen; Wang,Yunjie

doi:10.3892/ijmm.2017.3132

Eya2 overexpression promotes the invasion of human astrocytoma through the regulation of ERK/MMP9 signaling

Authors:
- Zhifeng Wen
- Chuansheng Liang
- Qichen Pan
- Yunjie Wang
View Affiliations

Affiliations: Department of Neurosurgery, The First Affiliated Hospital of China Medical University, Shenyang, Liaoning 110001, P.R. China
Published online on: September 13, 2017 https://doi.org/10.3892/ijmm.2017.3132
Pages: 1315-1322
Copyright: © Wen et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

The overexpression of eyes absent (Eya) 2 has been found in several human cancers. However, its biological roles and clinical significance in human astrocytoma have not yet been explored. This study investigated the clinical significance and biological roles of Eya2 in human astrocytoma tissues and cell lines. Using immunohistochemistry, we found Eya2 overexpression in 33 out of 90 (36.7%) astrocytoma specimens. The rate of Eya2 overexpression was higher in grade III-IV (48.1%) than in grade Ⅰ+Ⅱ astrocytomas (21.1%). Transfection with an Eya2 expression plasmid was performed in A172 cells with a low endogenous expression of Eya2 and the knockdown of Eya2 was carried out in U251 cells with a high endogenous expression using siRNA. Eya2 overexpression induced A172 cell proliferation and invasion, while the knockdown of Eya2 using siRNA decreased the proliferation and invasion of U251 cells. In addition, we found that transfection with the Eya2 expression plasmid facilitated cell cycle progression, and that the knockdown of Eya2 inhibited cell cycle progression, accompanied by a change in the expression of cell cycle-related proteins, including cyclin D1 and cyclin E. Eya2 also positively regulated extracellular signal-regulated kinase (ERK) activity and matrix metalloproteinase (MMP)9 expression. The blockade of ERK signaling using an inhibitor abolished the effects of Eya2 on A172 cell invasion and MMP9 production. In addition, we found that there was a positive correlation between Eya2 and Six1 in the astrocytoma cell lines. Immunoprecipitation revealed that Eya2 interacted with Six1 protein in the U251 cell line, which exhibited a high expression of both proteins. Eya2 failed to upregulate MMP expression in the A172 cells in which Six1 was silenced. On the whole, our data indicate that Eya2 may serve as a potential oncoprotein in human astrocytoma. Eya2 regulates astrocytoma cell proliferation and invasion, possibly through the regulation of ERK signaling.

View Figures

View References

1	Siegel R, Naishadham D and Jemal A: Cancer statistics, 2012. CA Cancer J Clin. 62:10–29. 2012. View Article : Google Scholar : PubMed/NCBI
2	Krupkova O Jr, Loja T, Redova M, Neradil J, Zitterbart K, Sterba J and Veselska R: Analysis of nuclear nestin localization in cell lines derived from neurogenic tumors. Tumour Biol. 32:631–639. 2011. View Article : Google Scholar : PubMed/NCBI
3	Zadran S, Amighi A, Otiniano E, Wong K and Zadran H: ENTPD5-mediated modulation of ATP results in altered metabolism and decreased survival in gliomablastoma multiforme. Tumour Biol. 33:2411–2421. 2012. View Article : Google Scholar : PubMed/NCBI
4	Chi F, Wu R, Jin X, Jiang M and Zhu X: HER2 induces cell proliferation and invasion of non-small-cell lung cancer by upregulating COX-2 expression via MEK/ERK signaling pathway. Onco Targets Ther. 9:2709–2716. 2016.PubMed/NCBI
5	Jiang Q, Pan Y, Cheng Y and Li H, Liu D and Li H: Lunasin suppresses the migration and invasion of breast cancer cells by inhibiting matrix metalloproteinase-2/-9 via the FAK/Akt/ERK and NF-κB signaling pathways. Oncol Rep. 36:253–262. 2016. View Article : Google Scholar : PubMed/NCBI
6	Ko HS, Park BJ, Choi SK, Kang HK, Kim A, Kim HS, Park IY and Shin JC: STAT3 and ERK signaling pathways are implicated in the invasion activity by oncostatin M through induction of matrix metalloproteinases 2 and 9. Yonsei Med J. 57:761–768. 2016. View Article : Google Scholar : PubMed/NCBI
7	Dong DD, Zhou H and Li G: ADAM15 targets MMP9 activity to promote lung cancer cell invasion. Oncol Rep. 34:2451–2460. 2015. View Article : Google Scholar : PubMed/NCBI
8	Kalhori V and Törnquist K: MMP2 and MMP9 participate in S1P-induced invasion of follicular ML-1 thyroid cancer cells. Mol Cell Endocrinol. 404:113–122. 2015. View Article : Google Scholar : PubMed/NCBI
9	Jacob A, Jing J, Lee J, Schedin P, Gilbert SM, Peden AA, Junutula JR and Prekeris R: Rab40b regulates trafficking of MMP2 and MMP9 during invadopodia formation and invasion of breast cancer cells. J Cell Sci. 126:4647–4658. 2013. View Article : Google Scholar : PubMed/NCBI
10	Silver SJ, Davies EL, Doyon L and Rebay I: Functional dissection of eyes absent reveals new modes of regulation within the retinal determination gene network. Mol Cell Biol. 23:5989–5999. 2003. View Article : Google Scholar : PubMed/NCBI
11	Kumar JP and Moses K: EGF receptor and Notch signaling act upstream of Eyeless/Pax6 to control eye specification. Cell. 104:687–697. 2001. View Article : Google Scholar : PubMed/NCBI
12	Kenyon KL, Ranade SS, Curtiss J, Mlodzik M and Pignoni F: Coordinating proliferation and tissue specification to promote regional identity in the Drosophila head. Dev Cell. 5:403–414. 2003. View Article : Google Scholar : PubMed/NCBI
13	Krishnan N, Jeong DG, Jung SK, Ryu SE, Xiao A, Allis CD, Kim SJ and Tonks NK: Dephosphorylation of the C-terminal tyrosyl residue of the DNA damage-related histone H2A.X is mediated by the protein phosphatase eyes absent. J Biol Chem. 284:16066–16070. 2009. View Article : Google Scholar : PubMed/NCBI
14	Farabaugh SM, Micalizzi DS, Jedlicka P, Zhao R and Ford HL: Eya2 is required to mediate the pro-metastatic functions of Six1 via the induction of TGF-β signaling, epithelial-mesenchymal transition, and cancer stem cell properties. Oncogene. 31:552–562. 2012.
15	Zhang L, Yang N, Huang J, Buckanovich RJ, Liang S, Barchetti A, Vezzani C, O'Brien-Jenkins A, Wang J, Ward MR, et al: Transcriptional coactivator Drosophila eyes absent homologue 2 is up-regulated in epithelial ovarian cancer and promotes tumor growth. Cancer Res. 65:925–932. 2005.PubMed/NCBI
16	Bierkens M, Krijgsman O, Wilting SM, Bosch L, Jaspers A, Meijer GA, Meijer CJ, Snijders PJ, Ylstra B and Steenbergen RD: Focal aberrations indicate EYA2 and hsa-miR-375 as oncogene and tumor suppressor in cervical carcinogenesis. Genes Chromosomes Cancer. 52:56–68. 2013. View Article : Google Scholar
17	Bai G, Chu J, Eli M, Bao Y and Wen H: PAQR3 overexpression suppresses the aggressive phenotype of esophageal squamous cell carcinoma cells via inhibition of ERK signaling. Biomed Pharmacother. 94:813–819. 2017. View Article : Google Scholar : PubMed/NCBI
18	Li J, Chen S and Ni B: Upregulation of HINT2 inhibits non-small cell lung cancer cell invasion via COX-2/PGE2-mediated activation of β-catenin signaling. Oncol Res. Aug 11–2017.Epub ahead of print. View Article : Google Scholar
19	Song J, Guan Z, Li M, Sha S, Song C, Gao Z and Zhao Y: MicroRNA-154 inhibits the growth and invasion of gastric cancer cells by targeting DIXDC1/WNT signaling. Oncol Res. Aug 11–2017.Epub ahead of print.
20	Liao Y, Yuan S, Chen X, Zhu P, Li J, Qin L and Liao W: Up-regulation of BRCA1-associated RING domain 1 promotes hepatocellular carcinoma progression by targeting Akt signaling. Sci Rep. 7:76492017. View Article : Google Scholar : PubMed/NCBI
21	Jin H, Xie Q, Guo X, Xu J, Wang A, Li J, Zhu J, Wu XR, Huang H and Huang C: p63α protein upregulates heat shock protein 70 expression via E2F1 transcription factor 1, promoting Wasf3/Wave3/MMP9 signaling and bladder cancer invasion. J Biol Chem. Aug 9–2017.Epub ahead of print. View Article : Google Scholar
22	Patrick AN, Cabrera JH, Smith AL, Chen XS, Ford HL and Zhao R: Structure-function analyses of the human SIX1-EYA2 complex reveal insights into metastasis and BOR syndrome. Nat Struct Mol Biol. 20:447–453. 2013. View Article : Google Scholar : PubMed/NCBI
23	Fu J, Xu X, Kang L, Zhou L, Wang S, Lu J, Cheng L, Fan Z, Yuan B, Tian P, et al: miR-30a suppresses breast cancer cell proliferation and migration by targeting Eya2. Biochem Biophys Res Commun. 445:314–319. 2014. View Article : Google Scholar : PubMed/NCBI
24	Abdullah JM, Ahmad F, Ahmad KA, Ghazali MM, Jaafar H, Ideris A, Ali AM, Omar AR, Yusoff K, Lila MA, et al: Molecular genetic analysis of BAX and cyclin D1 genes in patients with malignant glioma. Neurol Res. 29:239–242. 2007. View Article : Google Scholar : PubMed/NCBI
25	Arato-Ohshima T1 and Sawa H: Over-expression of cyclin D1 induces glioma invasion by increasing matrix metalloproteinase activity and cell motility. Int J Cancer. 83:387–392. 1999. View Article : Google Scholar : PubMed/NCBI
26	Velpula KK, Rehman AA, Chelluboina B, Dasari VR, Gondi CS, Rao JS and Veeravalli KK: Glioma stem cell invasion through regulation of the interconnected ERK, integrin α6 and N-cadherin signaling pathway. Cell Signal. 24:2076–2084. 2012. View Article : Google Scholar : PubMed/NCBI
27	Das G, Shiras A, Shanmuganandam K and Shastry P: Rictor regulates MMP-9 activity and invasion through Raf-1-MEK-ERK signaling pathway in glioma cells. Mol Carcinog. 50:412–423. 2011. View Article : Google Scholar : PubMed/NCBI
28	Lakka SS, Jasti SL, Gondi C, Boyd D, Chandrasekar N, Dinh DH, Olivero WC, Gujrati M and Rao JS: Downregulation of MMP-9 in ERK-mutated stable transfectants inhibits glioma invasion in vitro. Oncogene. 21:5601–5608. 2002. View Article : Google Scholar : PubMed/NCBI
29	Blevins MA, Towers CG, Patrick AN, Zhao R and Ford HL: The SIX1-EYA transcriptional complex as a therapeutic target in cancer. Expert Opin Ther Targets. 19:213–225. 2015. View Article : Google Scholar : PubMed/NCBI
30	Li Z, Tian T, Lv F, Chang Y, Wang X, Zhang L, Li X, Li L, Ma W, Wu J, et al: Six1 promotes proliferation of pancreatic cancer cells via upregulation of cyclin D1 expression. PLoS One. 8:e592032013. View Article : Google Scholar : PubMed/NCBI
31	Ono H, Imoto I, Kozaki K, Tsuda H, Matsui T, Kurasawa Y, Muramatsu T, Sugihara K and Inazawa J: SIX1 promotes epithelial-mesenchymal transition in colorectal cancer through ZEB1 activation. Oncogene. 31:4923–4934. 2012. View Article : Google Scholar : PubMed/NCBI
32	Tian T, Li A, Lu H, Luo R, Zhang M and Li Z: Six1 promotes glioblastoma cell proliferation and invasion by upregulation of connective tissue growth factor. Am J Cancer Res. 5:1823–1830. 2015.PubMed/NCBI
33	Iwanaga R, Wang CA, Micalizzi DS, Harrell JC, Jedlicka P, Sartorius CA, Kabos P, Farabaugh SM, Bradford AP and Ford HL: Expression of Six1 in luminal breast cancers predicts poor prognosis and promotes increases in tumor initiating cells by activation of extracellular signal-regulated kinase and transforming growth factor-beta signaling pathways. Breast Cancer Res. 14:R1002012. View Article : Google Scholar : PubMed/NCBI
34	Coletta RD, Christensen K, Reichenberger KJ, Lamb J, Micomonaco D, Huang L, Wolf DM, Müller-Tidow C, Golub TR, Kawakami K, et al: The Six1 homeoprotein stimulates tumorigenesis by reactivation of cyclin A1. Proc Natl Acad Sci USA. 101:6478–6483. 2004. View Article : Google Scholar : PubMed/NCBI