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RAN GTPase is an effector of the invasive/metastatic phenotype induced by osteopontin

Oncogene volume 27, pages 7139–7149 (2008)Cite this article

Abstract

Osteopontin (OPN) is a phosphorylated glycoprotein that binds to α v-containing integrins and is important in malignant transformation and cancer. Previously, we have utilized suppressive subtractive hybridization between mRNAs isolated from the Rama 37 (R37) rat mammary cell line and a subclone rendered invasive and metastatic by stable transfection with an expression vector for OPN to identify RAN GTPase (RAN) as the most overexpressed gene, in addition to that of OPN. Here we show that transfection of noninvasive R37 cells with an expression vector for RAN resulted in increased anchorage-independent growth, cell attachment and invasion through Matrigel in vitro, and metastasis in syngeneic rats. This induction of a malignant phenotype was induced independently of the expression of OPN, and was reversed by specifically reducing the expression of RAN using small-interfering RNAs. By using a combination of mutant protein and inhibitors, it was found that RAN signal transduction occurred through the c-Met receptor and PI3 kinase. This study therefore identifies RAN as a novel effector of OPN-mediated malignant transformation and some of its downstream signaling events in a mammary epithelial model of cancer invasion/metastasis.

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Accession codes

Accessions

GenBank/EMBL/DDBJ

Abbreviations

Mab:

monoclonal antibody

OPN:

osteopontin

Rama:

rat mammary

References

  • Abe H, Kamai T, Shirataki H, Oyama T, Arai K, Yoshida K . (2008). High expression of Ran GTPase is associated with local invasion and metastasis of human clear cell renal cell carcinoma. Int J Cancer 122: 2391–2397.

    Article  CAS  PubMed  Google Scholar 

  • Azuma K, Sasada T, Takedatsu H, Shomura H, Koga M, Maeda Y et al. (2004). Ran, a small GTPase gene, encodes cytotoxic T lymphocyte (CTL) epitopes capable of inducing HLA-A33-restricted and tumor-reactive CTLs in cancer patients. Clin Cancer Res 10: 6695–6702.

    Article  CAS  PubMed  Google Scholar 

  • Berns K, Hijmans EM, Mullenders J, Brummelkamp TR, Velds A, Heimerikx M et al. (2004). A large-scale RNAi screen in human cells identifies new components of the p53 pathway. Nature 428: 431–437.

    Article  CAS  PubMed  Google Scholar 

  • Beviglia L, Matsumoto K, Lin CS, Ziober BL, Kramer RH . (1997). Expression of the c-Met/HGF receptor in human breast carcinoma: correlation with tumor progression. Int J Cancer 74: 301–309.

    Article  CAS  PubMed  Google Scholar 

  • Bischoff FR, Klebe C, Kretschmer J, Wittinghofer A, Ponstingl H . (1994). RanGAP1 induces GTPase activity of nuclear Ras-related Ran. Proc Natl Acad Sci USA 91: 2587–2591.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bramwell VH, Doig GS, Tuck AB, Wilson SM, Tonkin KS, Tomiak A et al. (2006). Serial plasma osteopontin levels have prognostic value in metastatic breast cancer. Clin Cancer Res 12: 3337–3343.

    Article  CAS  PubMed  Google Scholar 

  • Carey KL, Richards SA, Lounsbury KM, Macara IG . (1996). Evidence using a green fluorescent protein-glucocorticoid receptor chimera that the Ran/TC4 GTPase mediates an essential function independent of nuclear protein import. J Cell Biol 133: 985–996.

    Article  CAS  PubMed  Google Scholar 

  • Chen H, Ke Y, Oates AJ, Barraclough R, Rudland PS . (1997). Isolation of and effector for metastasis-inducing DNAs from a human metastatic carcinoma cell line. Oncogene 14: 1581–1588.

    Article  CAS  PubMed  Google Scholar 

  • Darzynkiewicz Z, Juan G, Bedner E . (2001). Determining cell cycle stages by flow cytometry. Curr Protoc Cell Biol, Chapter 8, Unit 8 4.

  • Das R, Philip S, Mahabeleshwar GH, Bulbule A, Kundu GC, Rangaswami H et al. (2005). Osteopontin: it's role in regulation of cell motility and nuclear factor kappa B-mediated urokinase type plasminogen activator expression. IUBMB Life 57: 441–447.

    Article  CAS  PubMed  Google Scholar 

  • Davies BR, Davies MP, Gibbs FE, Barraclough R, Rudland PS . (1993). Induction of the metastatic phenotype by transfection of a benign rat mammary epithelial cell line with the gene for p9Ka, a rat calcium-binding protein, but not with the oncogene EJ-ras-1. Oncogene 8: 999–1008.

    CAS  PubMed  Google Scholar 

  • Denti S, Sirri A, Cheli A, Rogge L, Innamorati G, Putignano S et al. (2004). RanBPM is a phosphoprotein that associates with the plasma membrane and interacts with the integrin LFA-1. J Biol Chem 279: 13027–13034.

    Article  CAS  PubMed  Google Scholar 

  • Dunnington DJ, Hughes CM, Monaghan P, Rudland PS . (1983). Phenotypic instability of rat mammary tumor epithelial cells. J Natl Cancer Inst 71: 1227–1240.

    CAS  PubMed  Google Scholar 

  • El-Tanani M, Barraclough R, Wilkinson MC, Rudland PS . (2001). Metastasis-inducing DNA regulates the expression of the osteopontin gene by binding the transcription factor Tcf-4. Cancer Res 61: 5619–5629.

    CAS  PubMed  Google Scholar 

  • El-Tanani M, Platt-Higgins A, Rudland PS, Campbell FC . (2004). Ets gene PEA3 cooperates with β-catenin-Lef-1 and c-Jun in regulation of osteopontin transcription. J Biol Chem 279: 20794–20806.

    Article  CAS  PubMed  Google Scholar 

  • El-Tanani MK, Campbell FC, Crowe P, Erwin P, Harkin DP, Pharoah P et al. (2006a). BRCA1 suppresses osteopontin-mediated breast cancer. J Biol Chem 281: 26587–26601.

    Article  CAS  PubMed  Google Scholar 

  • El-Tanani MK, Campbell FC, Kurisetty V, Jin D, McCann M, Rudland PS . (2006b). The regulation and role of osteopontin in malignant transformation and cancer. Cytokine Growth Factor Rev 17: 463–474.

    Article  CAS  PubMed  Google Scholar 

  • Emberley ED, Gietz RD, Campbell JD, HayGlass KT, Murphy LC, Watson PH . (2002). RanBPM interacts with psoriasin in vitro and their expression correlates with specific clinical features in vivo in breast cancer. BMC Cancer 2: 28.

    Article  PubMed  PubMed Central  Google Scholar 

  • Furger KA, Allan AL, Wilson SM, Hota C, Vantyghem SA, Postenka CO et al. (2003). β(3) Integrin expression increases breast carcinoma cell responsiveness to the malignancy-enhancing effects of osteopontin. Mol Cancer Res 1: 810–819.

    CAS  PubMed  Google Scholar 

  • Hafizi S, Gustafsson A, Stenhoff J, Dahlback B . (2005). The RAN-binding protein RanBPM interacts with Axl and Sky receptor tyrosine kinases. Int J Biochem Cell Biol 37: 2344–2356.

    Article  CAS  PubMed  Google Scholar 

  • Hara S, Nakashiro K, Goda H, Hamakawa H . (2008). Role of Akt isoforms in HGF-induced invasive growth of human salivary gland cancer cells. Biochem Biophys Res Commun 370: 123–128.

    Article  CAS  PubMed  Google Scholar 

  • Johnston NI, Gunasekharan VK, Ravindranath A, O’Connell C, Johnston PG, El-Tanani MK . (2008). Osteopontin as a target for cancer therapy. Front Biosci 13: 4361–4372.

    Article  CAS  PubMed  Google Scholar 

  • Kilty IC, Barraclough R, Schmidt G, Rudland PS . (1999). Isolation of a potential neural stem cell line from the internal capsule of an adult transgenic rat brain. J Neurochem 73: 1859–1870.

    CAS  PubMed  Google Scholar 

  • Kurisetty VV, Johnston PG, Rudland PS, El-Tanani MK . (2008). Identification of genes differentially expressed between benign and osteopontin transformed rat mammary epithelial cells. BMC Research Notes; accepted for publication.

  • Lounsbury KM, Richards SA, Carey KL, Macara IG . (1996). Mutations within the Ran/TC4 GTPase. Effects on regulatory factor interactions and subcellular localization. J Biol Chem 271: 32834–32841.

    Article  CAS  PubMed  Google Scholar 

  • Margalit DN, Romberg L, Mets RB, Hebert AM, Mitchison TJ, Kirschner MW et al. (2004). Targeting cell division: small-molecule inhibitors of FtsZ GTPase perturb cytokinetic ring assembly and induce bacterial lethality. Proc Natl Acad Sci USA 101: 11821–11826.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Maulik G, Shrikhande A, Kijima T, Ma PC, Morrison PT, Salgia R . (2002). Role of the hepatocyte growth factor receptor, c-Met, in oncogenesis and potential for therapeutic inhibition. Cytokine Growth Factor Rev 13: 41–59.

    Article  CAS  PubMed  Google Scholar 

  • Morgan-Lappe SE, Tucker LA, Huang X, Zhang Q, Sarthy AV, Zakula D et al. (2007). Identification of Ras-related nuclear protein, targeting protein for xenopus kinesin-like protein 2, and stearoyl-CoA desaturase 1 as promising cancer targets from an RNAi-based screen. Cancer Res 67: 4390–4398.

    Article  CAS  PubMed  Google Scholar 

  • Moye VE, Barraclough R, West C, Rudland PS . (2004). Osteopontin expression correlates with adhesive and metastatic potential in metastasis-inducing DNA-transfected rat mammary cell lines. Br J Cancer 90: 1796–1802.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Nishitani H, Hirose E, Uchimura Y, Nakamura M, Umeda M, Nishii K et al. (2001). Full-sized RanBPM cDNA encodes a protein possessing a long stretch of proline and glutamine within the N-terminal region, comprising a large protein complex. Gene 272: 25–33.

    Article  CAS  PubMed  Google Scholar 

  • Oates AJ, Barraclough R, Rudland PS . (1996). The identification of osteopontin as a metastasis-related gene product in a rodent mammary tumour model. Oncogene 13: 97–104.

    CAS  PubMed  Google Scholar 

  • Ouellet V, Guyot MC, Le Page C, Filali-Mouhim A, Lussier C, Tonin PN et al. (2006). Tissue array analysis of expression microarray candidates identifies markers associated with tumor grade and outcome in serous epithelial ovarian cancer. Int J Cancer 119: 599–607.

    Article  CAS  PubMed  Google Scholar 

  • Peruzzi B, Bottaro DP . (2006). Targeting the c-Met signaling pathway in cancer. Clin Cancer Res 12: 3657–3660.

    Article  CAS  PubMed  Google Scholar 

  • Pisani P, Parkin DM, Bray F, Ferlay J . (1999). Estimates of the worldwide mortality from 25 cancers in 1990. Int J Cancer 83: 18–29.

    Article  CAS  PubMed  Google Scholar 

  • Rudland PS, Platt-Higgins A, El-Tanani M, de Silva Rudland S, Barraclough R, Winstanley JH et al. (2002). Prognostic significance of the metastasis-associated protein osteopontin in human breast cancer. Cancer Res 62: 3417–3427.

    CAS  PubMed  Google Scholar 

  • Sakata H, Stahl SJ, Taylor WG, Rosenberg JM, Sakaguchi K, Wingfield PT et al. (1997). Heparin binding and oligomerization of hepatocyte growth factor/scatter factor isoforms. Heparan sulfate glycosaminoglycan requirement for Met binding and signaling. J Biol Chem 272: 9457–9463.

    Article  CAS  PubMed  Google Scholar 

  • Sarbassov DD, Guertin DA, Ali SM, Sabatini DM . (2005). Phosphorylation and regulation of Akt/PKB by the rictor-mTOR complex. Science 307: 1098–1101.

    Article  CAS  PubMed  Google Scholar 

  • Sazer S, Dasso M . (2000). The ran decathlon: multiple roles of Ran. J Cell Sci 113 (Pt 7): 1111–1118.

    CAS  PubMed  Google Scholar 

  • Shinohara ML, Kim HJ, Kim JH, Garcia VA, Cantor H . (2008). Alternative translation of osteopontin generates intracellular and secreted isoforms that mediate distinct biological activities in dendritic cells. Proc Natl Acad Sci U S A 105: 7235–7239.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Shukla S, Maclennan GT, Hartman DJ, Fu P, Resnick MI, Gupta S . (2007). Activation of PI3K-Akt signaling pathway promotes prostate cancer cell invasion. Int J Cancer 121: 1424–1432.

    Article  CAS  PubMed  Google Scholar 

  • Singhal H, Bautista DS, Tonkin KS, O’Malley FP, Tuck AB, Chambers AF et al. (1997). Elevated plasma osteopontin in metastatic breast cancer associated with increased tumor burden and decreased survival. Clin Cancer Res 3: 605–611.

    CAS  PubMed  Google Scholar 

  • Su L, Mukherjee AB, Mukherjee BB . (1995). Expression of antisense osteopontin RNA inhibits tumor promoter-induced neoplastic transformation of mouse JB6 epidermal cells. Oncogene 10: 2163–2169.

    CAS  PubMed  Google Scholar 

  • Tuck AB, Chambers AF, Allan AL . (2007). Osteopontin overexpression in breast cancer: knowledge gained and possible implications for clinical management. J Cell Biochem 102: 859–868.

    Article  CAS  PubMed  Google Scholar 

  • Tuck AB, Elliott BE, Hota C, Tremblay E, Chambers AF . (2000). Osteopontin-induced, integrin-dependent migration of human mammary epithelial cells involves activation of the hepatocyte growth factor receptor (Met). J Cell Biochem 78: 465–475.

    Article  CAS  PubMed  Google Scholar 

  • Tuck AB, Hota C, Wilson SM, Chambers AF . (2003). Osteopontin-induced migration of human mammary epithelial cells involves activation of EGF receptor and multiple signal transduction pathways. Oncogene 22: 1198–1205.

    Article  CAS  PubMed  Google Scholar 

  • Tuck AB, O’Malley FP, Singhal H, Harris JF, Tonkin KS, Kerkvliet N et al. (1998). Osteopontin expression in a group of lymph node negative breast cancer patients. Int J Cancer 79: 502–508.

    Article  CAS  PubMed  Google Scholar 

  • Ue T, Yokozaki H, Kitadai Y, Yamamoto S, Yasui W, Ishikawa T et al. (1998). Co-expression of osteopontin and CD44v9 in gastric cancer. Int J Cancer 79: 127–132.

    Article  CAS  PubMed  Google Scholar 

  • Wang D, Li Z, Messing EM, Wu G . (2002). Activation of Ras/Erk pathway by a novel MET-interacting protein RanBPM. J Biol Chem 277: 36216–36222.

    Article  CAS  PubMed  Google Scholar 

  • Wang G, Platt-Higgins A, Carroll J, de Silva Rudland S, Winstanley J, Barraclough R et al. (2006). Induction of metastasis by S100P in a rat mammary model and its association with poor survival of breast cancer patients. Cancer Res 66: 1199–1207.

    Article  CAS  PubMed  Google Scholar 

  • Wennerberg K, Rossman KL, Der CJ . (2005). The Ras superfamily at a glance. J Cell Sci 118: 843–846.

    Article  CAS  PubMed  Google Scholar 

  • Yuan Y, Fu C, Chen H, Wang X, Deng W, Huang BR . (2006). The RAN-binding protein RanBPM interacts with TrkA receptor. Neurosci Lett 407: 26–31.

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

This work was supported by grants from Research and Development Office, Queen's University Belfast, Northern Ireland, UK and The Cancer and Polio Research Fund, Wirral, UK. DGF's post is funded by the Northwest Cancer Research Fund, Liverpool, UK.

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Authors and Affiliations

  1. Centre for Cancer Research and Cell Biology, Queen's University Belfast, Belfast, UK

    V V Kurisetty, P G Johnston, N Johnston, P Erwin, P Crowe, F C Campbell & M K El-Tanani

  2. Cancer and Polio Research Fund Laboratories, School of Biological Sciences, University of Liverpool, Liverpool, UK

    D G Fernig, I P Anderson & P S Rudland

Authors
  1. V V Kurisetty
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  2. P G Johnston
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  3. N Johnston
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  4. P Erwin
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  7. F C Campbell
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  8. I P Anderson
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  10. M K El-Tanani
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Correspondence to M K El-Tanani.

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Supplementary Information accompanies the paper on the Oncogene website (http://www.nature.com/onc)

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Kurisetty, V., Johnston, P., Johnston, N. et al. RAN GTPase is an effector of the invasive/metastatic phenotype induced by osteopontin. Oncogene 27, 7139–7149 (2008). https://doi.org/10.1038/onc.2008.325

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