Deletion of the short arm of chromosome 3 is one of the most frequent genetic alterations in many solid tumors including nasopharyngeal carcinoma (NPC), suggesting the existence of one or more tumor suppressor genes (TSGs) within the frequently deleted region. A putative TSG RBMS3 (RNA binding motif, single stranded interacting protein 3), located at 3p24-p23, has been identified in our previous study. Here, we reported that downregulation of RBMS3 was detected in 3/3 NPC cell lines and 13/15 (86.7%) primary NPC tissues. Functional studies using both overexpression and suppression systems demonstrated that RBMS3 has a strong tumor suppressive role in NPC. The tumor suppressive mechanism of RBMS3 was associated with its role in cell cycle arrest at the G1/S checkpoint by upregulating p53 and p21, downregulating cyclin E and CDK2, and the subsequent inhibition of Rb-ser780. Further analysis demonstrated that RBMS3 had a pro-apoptotic role in a mitochondrial-dependent manner via activation of caspase-9 and PARP. Finally, RBMS3 inhibited microvessel formation, which may be mediated by down-regulation of MMP2 and β-catenin and inactivation of its downstream targets, including cyclin-D1, c-Myc, MMP7, and MMP9. Taken together, our findings define a function for RBMS3 as an important tumor suppressor gene in NPC.
References
[1]
Jeannel D, Bouvier G, Huber A (1999) Nasopharyngeal carcinoma: an epidemiological approach to carcinogenesis. Cancer Surv 33: 125–155.
[2]
Parkin DM, Bray F, Ferlay J, Pisani P (2005) Global cancer statistics, 2002. CA Cancer J Clin 55: 74–108.
Yuan JM, Wang XL, Xiang YB, Gao YT, Ross RK, et al. (2000) Preserved foods in relation to risk of nasopharyngeal carcinoma in Shanghai, China. Int J Cancer 85: 358–363.
[5]
Liebowitz D (1994) Nasopharyngeal carcinoma: the Epstein-Barr virus association. Semin Oncol 21: 376–381.
[6]
Lo KW, Huang DP (2002) Genetic and epigenetic changes in nasopharyngeal carcinoma. Semin Cancer Biol 12: 451–462.
[7]
Henle G, Henle W (1976) Epstein-Barr virus-specific IgA serum antibodies as an outstanding feature of nasopharyngeal carcinoma. Int J Cancer 17: 1–7.
[8]
Kwong D, Lam A, Guan X, Law S, Tai A, et al. (2004) Chromosomal aberrations in esophageal squamous cell carcinoma among Chinese: gain of 12p predicts poor prognosis after surgery. Hum Pathol 35: 309–316.
[9]
Hesson LB, Cooper WN, Latif F (2007) Evaluation of the 3p21.3 tumour-suppressor gene cluster. Oncogene 26: 7283–7301.
[10]
Qiu GH, Tan LK, Loh KS, Lim CY, Srivastava G, et al. (2004) The candidate tumor suppressor gene BLU, located at the commonly deleted region 3p21.3, is an E2F-regulated, stress-responsive gene and inactivated by both epigenetic and genetic mechanisms in nasopharyngeal carcinoma. Oncogene 23: 4793–4806.
[11]
Qin YR, Fu L, Sham PC, Kwong DL, Zhu CL, et al. (2008) Single-nucleotide polymorphism-mass array reveals commonly deleted regions at 3p22 and 3p14.2 associate with poor clinical outcome in esophageal squamous cell carcinoma. Int J Cancer 123: 826–830.
[12]
Qiu GH, Salto-Tellez M, Ross JA, Yeo W, Cui Y, et al. (2008) The tumor suppressor gene DLEC1 is frequently silenced by DNA methylation in hepatocellular carcinoma and induces G1 arrest in cell cycle. J Hepatol 48: 433–441.
[13]
Fu L, Qin YR, Xie D, Hu L, Kwong DL, et al. (2007) Characterization of a novel tumor-suppressor gene PLC delta 1 at 3p22 in esophageal squamous cell carcinoma. Cancer Res 67: 10720–10726.
[14]
Hu XT, Zhang FB, Fan YC, Shu XS, Wong AH, et al. (2009) Phospholipase C delta 1 is a novel 3p22.3 tumor suppressor involved in cytoskeleton organization, with its epigenetic silencing correlated with high-stage gastric cancer. Oncogene 28: 2466–2475.
[15]
Penkov D, Ni R, Else C, Pinol-Roma S, Ramirez F, et al. (2000) Cloning of a human gene closely related to the genes coding for the c-myc single-strand binding proteins. Gene 243: 27–36.
[16]
Arvanitis C, Felsher DW (2006) Conditional transgenic models define how MYC initiates and maintains tumorigenesis. Semin Cancer Biol 16: 313–317.
[17]
Fan CS, Wong N, Leung SF, To KF, Lo KW, et al. (2000) Frequent c-myc and Int-2 overrepresentations in nasopharyngeal carcinoma. Hum Pathol 31: 169–178.
[18]
Hui AB, Lo KW, Teo PM, To KF, Huang DP (2002) Genome wide detection of oncogene amplifications in nasopharyngeal carcinoma by array based comparative genomic hybridization. Int J Oncol 20: 467–473.
[19]
Fritz D, Stefanovic B (2007) RNA-binding protein RBMS3 is expressed in activated hepatic stellate cells and liver fibrosis and increases expression of transcription factor Prx1. J Mol Biol 371: 585–595.
[20]
Wang XH, Sun X, Meng XW, Lv ZW, Du YJ, et al. (2010) beta-catenin siRNA regulation of apoptosis- and angiogenesis-related gene expression in hepatocellular carcinoma cells: potential uses for gene therapy. Oncol Rep 24: 1093–1099.
[21]
Carnero A, Hannon GJ (1998) The INK4 family of CDK inhibitors. Curr Top Microbiol Immunol 227: 43–55.
[22]
Lu Z, Ghosh S, Wang Z, Hunter T (2003) Downregulation of caveolin-1 function by EGF leads to the loss of E-cadherin, increased transcriptional activity of beta-catenin, and enhanced tumor cell invasion. Cancer Cell 4: 499–515.
[23]
Sherr CJ, Roberts JM (1999) CDK inhibitors: positive and negative regulators of G1-phase progression. Genes Dev 13: 1501–1512.
[24]
Hoshino I, Matsubara H, Akutsu Y, Nishimori T, Yoneyama Y, et al. (2007) Tumor suppressor Prdx1 is a prognostic factor in esophageal squamous cell carcinoma patients. Oncol Rep 18: 867–871.
[25]
Giorgio M, Trinei M, Migliaccio E, Pelicci PG (2007) Hydrogen peroxide: a metabolic by-product or a common mediator of ageing signals? Nat Rev Mol Cell Biol 8: 722–728.
[26]
Pellegrini M, Pacini S, Baldari CT (2005) p66SHC: the apoptotic side of Shc proteins. Apoptosis 10: 13–18.
[27]
Petronilli V, Costantini P, Scorrano L, Colonna R, Passamonti S, et al. (1994) The voltage sensor of the mitochondrial permeability transition pore is tuned by the oxidation-reduction state of vicinal thiols. Increase of the gating potential by oxidants and its reversal by reducing agents. J Biol Chem 269: 16638–16642.
[28]
Bernardi P, Petronilli V, Di Lisa F, Forte M (2001) A mitochondrial perspective on cell death. Trends Biochem Sci 26: 112–117.
[29]
Bouck N, Stellmach V, Hsu SC (1996) How tumors become angiogenic. In: Advances in Cancer Research, Vol VandeWoude GFKG, editor. 69: 135–174.
[30]
Mukhopadhyay D, Tsiokas L, Sukhatme VP (1995) Wild-type p53 and v-Src exert opposing influences on human vascular endothelial growth factor gene expression. Cancer Res 55: 6161–6165.
[31]
Rak J, Mitsuhashi Y, Bayko L, Filmus J, Shirasawa S, et al. (1995) Mutant ras oncogenes upregulate VEGF/VPF expression: implications for induction and inhibition of tumor angiogenesis. Cancer Res 55: 4575–4580.
[32]
Kraemer M, Tournaire R, Dejong V, Montreau N, Briane D, et al. (1999) Rat embryo fibroblasts transformed by c-Jun display highly metastatic and angiogenic activities in vivo and deregulate gene expression of both angiogenic and antiangiogenic factors. Cell Growth Differ 10: 193–200.
[33]
Marconcini L, Marchio S, Morbidelli L, Cartocci E, Albini A, et al. (1999) c-fos-induced growth factor/vascular endothelial growth factor D induces angiogenesis in vivo and in vitro. Proc Natl Acad Sci U S A 96: 9671–9676.
[34]
Gnarra JR, Zhou S, Merrill MJ, Wagner JR, Krumm A, et al. (1996) Post-transcriptional regulation of vascular endothelial growth factor mRNA by the product of the VHL tumor suppressor gene. Proc Natl Acad Sci U S A 93: 10589–10594.
[35]
Claudio PP, Stiegler P, Howard CM, Bellan C, Minimo C, et al. (2001) RB2/p130 gene-enhanced expression down-regulates vascular endothelial growth factor expression and inhibits angiogenesis in vivo. Cancer Res 61: 462–468.
[36]
Ahn GO, Brown JM (2008) Matrix metalloproteinase-9 is required for tumor vasculogenesis but not for angiogenesis: role of bone marrow-derived myelomonocytic cells. Cancer Cell 13: 193–205.
[37]
van Noort M, Meeldijk J, van der Zee R, Destree O, Clevers H (2002) Wnt signaling controls the phosphorylation status of beta-catenin. J Biol Chem 277: 17901–17905.
[38]
Iwai S, Yonekawa A, Harada C, Hamada M, Katagiri W, et al. (2010) Involvement of the Wnt-beta-catenin pathway in invasion and migration of oral squamous carcinoma cells. Int J Oncol 37: 1095–1103.
[39]
Taurin S, Sandbo N, Qin Y, Browning D, Dulin NO (2006) Phosphorylation of beta-catenin by cyclic AMP-dependent protein kinase. J Biol Chem 281: 9971–9976.
[40]
Morrison JA, Gulley ML, Pathmanathan R, Raab-Traub N (2004) Differential signaling pathways are activated in the Epstein-Barr virus-associated malignancies nasopharyngeal carcinoma and Hodgkin lymphoma. Cancer Res 64: 5251–5260.
[41]
Zeng ZY, Zhou YH, Zhang WL, Xiong W, Fan SQ, et al. (2007) Gene expression profiling of nasopharyngeal carcinoma reveals the abnormally regulated Wnt signaling pathway. Hum Pathol 38: 120–133.
[42]
Luo J, Xiao J, Tao Z, Li X (1997) Detection of c-myc gene expression in nasopharyngeal carcinoma by nonradioactive in situ hybridization and immunohistochemistry. Chin Med J (Engl) 110: 229–232.
[43]
Porter MJ, Field JK, Leung SF, Lo D, Lee JC, et al. (1994) The detection of the c-myc and ras oncogenes in nasopharyngeal carcinoma by immunohistochemistry. Acta Otolaryngol 114: 105–109.
[44]
Fang YJ, Lu ZH, Wang GQ, Pan ZZ, Zhou ZW, et al. (2009) Elevated expressions of MMP7, TROP2, and survivin are associated with survival, disease recurrence, and liver metastasis of colon cancer. Int J Colorectal Dis 24: 875–884.
[45]
Yue W, Sun Q, Landreneau R, Wu C, Siegfried JM, et al. (2009) Fibulin-5 suppresses lung cancer invasion by inhibiting matrix metalloproteinase-7 expression. Cancer Res 69: 6339–6346.
[46]
Li Y, Chen L, Nie CJ, Zeng TT, Liu H, et al. (2011) Downregulation of RBMS3 is associated with poor prognosis in esophageal squamous cell carcinoma. Cancer Res 71: 6106–6115.
[47]
Hui AB, Cheung ST, Fong Y, Lo KW, Huang DP (1998) Characterization of a new EBV-associated nasopharyngeal carcinoma cell line. Cancer Genet Cytogenet 101: 83–88.
[48]
Cheung ST, Huang DP, Hui AB, Lo KW, Ko CW, et al. (1999) Nasopharyngeal carcinoma cell line (C666–1) consistently harbouring Epstein-Barr virus. Int J Cancer 83: 121–126.
[49]
Sizhong Z, Xiukung G, Yi Z (1983) Cytogenetic studies on an epithelial cell line derived from poorly differentiated nasopharyngeal carcinoma. Int J Cancer 31: 587–590.
[50]
Li HM, Man C, Jin Y, Deng W, Yip YL, et al. (2006) Molecular and cytogenetic changes involved in the immortalization of nasopharyngeal epithelial cells by telomerase. Int J Cancer 119: 1567–1576.
[51]
Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(T)(-Delta Delta C) method. Methods 25: 402–408.
[52]
Fox SB (2001) Microscopic assessment of angiogenesis in tumors. Methods Mol Med 46: 29–46.
