Impact of Mucin1 knockdown on the phenotypic characteristics of the human hepatocellular carcinoma cell line SMMC-7721

Li,Qiongshu; Wang,Fengli; Liu,Guomu; Yuan,Hongyan; Chen,Tanxiu; Wang,Juan; Xie,Fei; Zhai,Ruiping; Wang,Fang; Guo,Yingying; Ni,Weihua; Tai,Guixiang

doi:10.3892/or.2014.3136

Impact of Mucin1 knockdown on the phenotypic characteristics of the human hepatocellular carcinoma cell line SMMC-7721

Authors:
- Qiongshu Li
- Fengli Wang
- Guomu Liu
- Hongyan Yuan
- Tanxiu Chen
- Juan Wang
- Fei Xie
- Ruiping Zhai
- Fang Wang
- Yingying Guo
- Weihua Ni
- Guixiang Tai
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Affiliations: Department of Immunology, College of Basic Medical Science, Jilin University, Changchun, Jilin 130021, P.R. China
Published online on: April 14, 2014 https://doi.org/10.3892/or.2014.3136
Pages: 2811-2819

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Abstract

Mucin1 (MUC1) is a transmembrane glycoprotein that plays a key role as an oncogene in the tumorigenesis of many human adenocarcinomas. However, the role of MUC1 in human hepatocellular carcinoma (HCC) progression remains unclear. In the present study, we silenced MUC1 to investigate its effect on the human HCC cell line SMMC-7721 and found that knockdown of MUC1 signiﬁcantly inhibited cell proliferation, enhanced cell-cell aggregation and induced apoptosis. No significant differences were found in in vitro migration or invasion. We also observed that knockdown of MUC1 decreased the translocation of β‑catenin to the nucleus, reduced the activity of T cell factor and blocked the expression of cyclin D1 and c-Myc. In addition, MUC1 knockdown enhanced the expression of E-cadherin, a molecular chaperone of β‑catenin that plays an important role in cell-cell aggregation. In vivo assays demonstrated that there was no tumor growth in mice injected with MUC1-silenced cells. Global gene expression analysis showed that a series of genes encoding molecules in the Wnt/β‑catenin, nuclear factor-κB (NF-κB), mitogen-activated protein kinase (MAPK), insulin, transforming growth factor β (TGF-β) and vascular endothelial growth factor (VEGF) signaling pathways were all influenced by the knockdown of MUC1, and these may contribute to the phenotypic alterations observed. Collectively, our results indicate that MUC1 plays a key role in HCC tumorigenesis.

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1	Kufe DW: Mucins in cancer: function, prognosis and therapy. Nat Rev Cancer. 9:874–885. 2009. View Article : Google Scholar : PubMed/NCBI
2	Duffy MJ: CA 15-3 and related mucins as circulating markers in breast cancer. Ann Clin Biochem. 36(Pt 5): 579–586. 1999. View Article : Google Scholar : PubMed/NCBI
3	Apostolopoulos V, Pietersz GA, Tsibanis A, et al: Pilot phase III immunotherapy study in early-stage breast cancer patients using oxidized mannan-MUC1 [ISRCTN71711835]. Breast Cancer Res. 8:R272006.
4	Sugiura D, Aida S, Denda-Nagai K, et al: Differential effector mechanisms induced by vaccination with MUC1 DNA in the rejection of colon carcinoma growth at orthotopic sites and metastases. Cancer Sci. 99:2477–2484. 2008. View Article : Google Scholar : PubMed/NCBI
5	Zhang S, Zhang HS, Cordon-Cardo C, Ragupathi G and Livingston PO: Selection of tumor antigens as targets for immune attack using immunohistochemistry: protein antigens. Clin Cancer Res. 4:2669–2676. 1998.PubMed/NCBI
6	Gendler S, Taylor-Papadimitriou J, Duhig T, Rothbard J and Burchell J: A highly immunogenic region of a human polymorphic epithelial mucin expressed by carcinomas is made up of tandem repeats. J Biol Chem. 263:12820–12823. 1988.PubMed/NCBI
7	Ligtenberg MJ, Kruijshaar L, Buijs F, van Meijer M, Litvinov SV and Hilkens J: Cell-associated episialin is a complex containing two proteins derived from a common precursor. J Biol Chem. 267:6171–6177. 1992.PubMed/NCBI
8	Kufe DW: Targeting the human MUC1 oncoprotein: a tale of two proteins. Cancer Biol Ther. 7:81–84. 2008. View Article : Google Scholar : PubMed/NCBI
9	Singh PK and Hollingsworth MA: Cell surface-associated mucins in signal transduction. Trends Cell Biol. 16:467–476. 2006. View Article : Google Scholar : PubMed/NCBI
10	Carraway KL III, Funes M, Workman HC and Sweeney C: Contribution of membrane mucins to tumor progression through modulation of cellular growth signaling pathways. Curr Top Dev Biol. 78:1–22. 2007. View Article : Google Scholar : PubMed/NCBI
11	Yu LG, Andrews N, Zhao Q, et al: Galectin-3 interaction with Thomsen-Friedenreich disaccharide on cancer-associated MUC1 causes increased cancer cell endothelial adhesion. J Biol Chem. 282:773–781. 2007. View Article : Google Scholar : PubMed/NCBI
12	Rahn JJ, Shen Q, Mah BK and Hugh JC: MUC1 initiates a calcium signal after ligation by intercellular adhesion molecule-1. J Biol Chem. 279:29386–29390. 2004. View Article : Google Scholar : PubMed/NCBI
13	Rahn JJ, Chow JW, Horne GJ, et al: MUC1 mediates transendothelial migration in vitro by ligating endothelial cell ICAM-1. Clin Exp Metastasis. 22:475–483. 2005. View Article : Google Scholar : PubMed/NCBI
14	Shen Q, Rahn JJ, Zhang J, et al: MUC1 initiates Src-CrkL- Rac1/Cdc42-mediated actin cytoskeletal protrusive motility after ligating intercellular adhesion molecule-1. Mol Cancer Res. 6:555–567. 2008. View Article : Google Scholar : PubMed/NCBI
15	Zhao Q, Barclay M, Hilkens J, et al: Interaction between circulating galectin-3 and cancer-associated MUC1 enhances tumour cell homotypic aggregation and prevents anoikis. Mol Cancer. 9:1542010. View Article : Google Scholar : PubMed/NCBI
16	Huang L, Chen D, Liu D, Yin L, Kharbanda S and Kufe D: MUC1 oncoprotein blocks glycogen synthase kinase 3beta-mediated phosphorylation and degradation of beta-catenin. Cancer Res. 65:10413–10422. 2005. View Article : Google Scholar : PubMed/NCBI
17	Li Y, Kuwahara H, Ren J, Wen G and Kufe D: The c-Src tyrosine kinase regulates signaling of the human DF3/MUC1 carcinoma-associated antigen with GSK3 beta and beta-catenin. J Biol Chem. 276:6061–6064. 2001. View Article : Google Scholar : PubMed/NCBI
18	Pandey P, Kharbanda S and Kufe D: Association of the DF3/MUC1 breast cancer antigen with Grb2 and the Sos/Ras exchange protein. Cancer Res. 55:4000–4003. 1995.PubMed/NCBI
19	Wei X, Xu H and Kufe D: Human mucin 1 oncoprotein represses transcription of the p53 tumor suppressor gene. Cancer Res. 67:1853–1858. 2007. View Article : Google Scholar : PubMed/NCBI
20	Singh PK, Behrens ME, Eggers JP, et al: Phosphorylation of MUC1 by Met modulates interaction with p53 and MMP1 expression. J Biol Chem. 283:26985–26995. 2008. View Article : Google Scholar : PubMed/NCBI
21	Schroeder JA, Thompson MC, Gardner MM and Gendler SJ: Transgenic MUC1 interacts with epidermal growth factor receptor and correlates with mitogen-activated protein kinase activation in the mouse mammary gland. J Biol Chem. 276:13057–13064. 2001. View Article : Google Scholar : PubMed/NCBI
22	Lau SK, Shields DJ, Murphy EA, et al: EGFR-mediated carcinoma cell metastasis mediated by integrin alphavbeta5 depends on activation of c-Src and cleavage of MUC1. PLoS One. 7:e367532012. View Article : Google Scholar : PubMed/NCBI
23	Ahmad R, Raina D, Trivedi V, et al: MUC1 oncoprotein activates the IkappaB kinase beta complex and constitutive NF-kappaB signalling. Nat Cell Biol. 9:1419–1427. 2007. View Article : Google Scholar : PubMed/NCBI
24	Ahmad R, Raina D, Joshi MD, et al: MUC1-C oncoprotein functions as a direct activator of the nuclear factor-kappaB p65 transcription factor. Cancer Res. 69:7013–7021. 2009. View Article : Google Scholar : PubMed/NCBI
25	Hu XF, Yang E, Li J and Xing PX: MUC1 cytoplasmic tail: a potential therapeutic target for ovarian carcinoma. Expert Rev Anticancer Ther. 6:1261–1271. 2006. View Article : Google Scholar : PubMed/NCBI
26	Kufe DW: Functional targeting of the MUC1 oncogene in human cancers. Cancer Biol Ther. 8:1197–1203. 2009. View Article : Google Scholar : PubMed/NCBI
27	Klinge CM, Radde BN, Imbert-Fernandez Y, et al: Targeting the intracellular MUC1 C-terminal domain inhibits proliferation and estrogen receptor transcriptional activity in lung adenocarcinoma cells. Mol Cancer Ther. 10:2062–2071. 2011. View Article : Google Scholar
28	Hattrup CL, Bradley JM, Kotlarczyk KL, et al: The MUC1 cytoplasmic tail and tandem repeat domains contribute to mammary oncogenesis in FVB mice. Breast Cancer (Auckl). 1:57–63. 2008.PubMed/NCBI
29	Wang L, Ma J, Liu F, et al: Expression of MUC1 in primary and metastatic human epithelial ovarian cancer and its therapeutic significance. Gynecol Oncol. 105:695–702. 2007. View Article : Google Scholar : PubMed/NCBI
30	Kohlgraf KG, Gawron AJ, Higashi M, et al: Contribution of the MUC1 tandem repeat and cytoplasmic tail to invasive and metastatic properties of a pancreatic cancer cell line. Cancer Res. 63:5011–5020. 2003.PubMed/NCBI
31	Inagaki Y, Tang W, Xu H, et al: Sustained aberrant localization of KL-6 mucin and beta-catenin at the invasion front of human gastric cancer cells. Anticancer Res. 31:535–542. 2011.PubMed/NCBI
32	Bozkaya G, Korhan P, Cokakli M, et al: Cooperative interaction of MUC1 with the HGF/c-Met pathway during hepatocarcinogenesis. Mol Cancer. 11:642012. View Article : Google Scholar : PubMed/NCBI
33	Yuan SF, Li KZ, Wang L, et al: Expression of MUC1 and its significance in hepatocellular and cholangiocarcinoma tissue. World J Gastroenterol. 11:4661–4666. 2005.PubMed/NCBI
34	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(−Delta Delta C(T)) method. Methods. 25:402–408. 2001.
35	Veeravalli KK, Chetty C, Ponnala S, et al: MMP-9, uPAR and cathepsin B silencing downregulate integrins in human glioma xenograft cells in vitro and in vivo in nude mice. PLoS One. 5:e115832010. View Article : Google Scholar : PubMed/NCBI
36	Wang F, Li Q, Ni W, et al: Expression of human full-length MUC1 inhibits the proliferation and migration of a B16 mouse melanoma cell line. Oncol Rep. 30:260–268. 2013.PubMed/NCBI
37	Costa NR, Paulo P, Caffrey T, Hollingsworth MA and Santos-Silva F: Impact of MUC1 mucin downregulation in the phenotypic characteristics of MKN45 gastric carcinoma cell line. PLoS One. 6:e269702011. View Article : Google Scholar : PubMed/NCBI
38	Tsutsumida H, Swanson BJ, Singh PK, et al: RNA interference suppression of MUC1 reduces the growth rate and metastatic phenotype of human pancreatic cancer cells. Clin Cancer Res. 12:2976–2987. 2006. View Article : Google Scholar : PubMed/NCBI
39	Raina D, Kharbanda S and Kufe D: The MUC1 oncoprotein activates the anti-apoptotic phosphoinositide 3-kinase/Akt and Bcl-xL pathways in rat 3Y1 fibroblasts. J Biol Chem. 279:20607–20612. 2004. View Article : Google Scholar : PubMed/NCBI
40	Raina D, Ahmad R, Chen D, Kumar S, Kharbanda S and Kufe D: MUC1 oncoprotein suppresses activation of the ARF-MDM2-p53 pathway. Cancer Biol Ther. 7:1959–1967. 2008. View Article : Google Scholar : PubMed/NCBI
41	Agata N, Ahmad R, Kawano T, Raina D, Kharbanda S and Kufe D: MUC1 oncoprotein blocks death receptor-mediated apoptosis by inhibiting recruitment of caspase-8. Cancer Res. 68:6136–6144. 2008. View Article : Google Scholar : PubMed/NCBI
42	Ahmad R, Alam M, Rajabi H and Kufe D: The MUC1-C oncoprotein binds to the BH3 domain of the pro-apoptotic BAX protein and blocks BAX function. J Biol Chem. 287:20866–20875. 2012. View Article : Google Scholar : PubMed/NCBI
43	Li Y, Liu D, Chen D, Kharbanda S and Kufe D: Human DF3/MUC1 carcinoma-associated protein functions as an oncogene. Oncogene. 22:6107–6110. 2003. View Article : Google Scholar : PubMed/NCBI
44	Wen Y, Caffrey TC, Wheelock MJ, Johnson KR and Hollingsworth MA: Nuclear association of the cytoplasmic tail of MUC1 and beta-catenin. J Biol Chem. 278:38029–38039. 2003. View Article : Google Scholar : PubMed/NCBI
45	Li Y, Yi H, Yao Y, et al: The cytoplasmic domain of MUC1 induces hyperplasia in the mammary gland and correlates with nuclear accumulation of beta-catenin. PLoS One. 6:e191022011. View Article : Google Scholar : PubMed/NCBI
46	Yuan Z, Wong S, Borrelli A and Chung MA: Downregulation of MUC1 in cancer cells inhibits cell migration by promoting E-cadherin/catenin complex formation. Biochem Biophys Res Commun. 362:740–746. 2007. View Article : Google Scholar : PubMed/NCBI