Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Original Article
  • Published:

ZEB1 induces LOXL2-mediated collagen stabilization and deposition in the extracellular matrix to drive lung cancer invasion and metastasis

Oncogene volume 36, pages 1925–1938 (2017)Cite this article

Subjects

Abstract

Lung cancer is the leading cause of cancer-related deaths, primarily due to distant metastatic disease. Metastatic lung cancer cells can undergo an epithelial-to-mesenchymal transition (EMT) regulated by various transcription factors, including a double-negative feedback loop between the microRNA-200 (miR-200) family and ZEB1, but the precise mechanisms by which ZEB1-dependent EMT promotes malignancy remain largely undefined. Although the cell-intrinsic effects of EMT are important for tumor progression, the reciprocal dynamic crosstalk between mesenchymal cancer cells and the extracellular matrix (ECM) is equally critical in regulating invasion and metastasis. Investigating the collaborative effect of EMT and ECM in the metastatic process reveals increased collagen deposition in metastatic tumor tissues as a direct consequence of amplified collagen gene expression in ZEB1-activated mesenchymal lung cancer cells. In addition, collagen fibers in metastatic lung tumors exhibit greater linearity and organization as a result of collagen crosslinking by the lysyl oxidase (LOX) family of enzymes. Expression of the LOX and LOXL2 isoforms is directly regulated by miR-200 and ZEB1, respectively, and their upregulation in metastatic tumors and mesenchymal cell lines is coordinated to that of collagen. Functionally, LOXL2, as opposed to LOX, is the principal isoform that crosslinks and stabilizes insoluble collagen deposition in tumor tissues. In turn, focal adhesion formation and FAK/SRC signaling is activated in mesenchymal tumor cells by crosslinked collagen in the ECM. Our study is the first to validate direct regulation of LOX and LOXL2 by the miR-200/ZEB1 axis, defines a novel mechanism driving tumor metastasis, delineates collagen as a prognostic marker, and identifies LOXL2 as a potential therapeutic target against tumor progression.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7

Similar content being viewed by others

References

  1. Siegel R, Ma J, Zou Z, Jemal A . Cancer statistics, 2014. CA Cancer J Clin 2014; 64: 9–29.

    Article  PubMed  Google Scholar 

  2. Gibbons DL, Lin W, Creighton CJ, Zheng S, Berel D, Yang Y et al. Expression signatures of metastatic capacity in a genetic mouse model of lung adenocarcinoma. PLoS One 2009; 4: e5401.

    Article  PubMed  PubMed Central  Google Scholar 

  3. Gibbons DL, Lin W, Creighton CJ, Rizvi ZH, Gregory PA, Goodall GJ et al. Contextual extracellular cues promote tumor cell EMT and metastasis by regulating miR-200 family expression. Genes Dev 2009; 23: 2140–2151.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Gregory PA, Bert AG, Paterson EL, Barry SC, Tsykin A, Farshid G et al. The miR-200 family and miR-205 regulate epithelial to mesenchymal transition by targeting ZEB1 and SIP1. Nat Cell Biol 2008; 10: 593–601.

    Article  CAS  PubMed  Google Scholar 

  5. Gregory PA, Bracken CP, Smith E, Bert AG, Wright JA, Roslan S et al. An autocrine TGF-beta/ZEB/miR-200 signaling network regulates establishment and maintenance of epithelial-mesenchymal transition. Mol Biol Cell 2011; 22: 1686–1698.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Yang J, Weinberg RA . Epithelial-mesenchymal transition: at the crossroads of development and tumor metastasis. Dev Cell 2008; 14: 818–829.

    Article  CAS  PubMed  Google Scholar 

  7. Wellner U, Schubert J, Burk UC, Schmalhofer O, Zhu F, Sonntag A et al. The EMT-activator ZEB1 promotes tumorigenicity by repressing stemness-inhibiting microRNAs. Nat Cell Biol 2009; 11: 1487–1495.

    Article  CAS  PubMed  Google Scholar 

  8. Ma L, Weinberg RA . Micromanagers of malignancy: role of microRNAs in regulating metastasis. Trends Genet 2008; 24: 448–456.

    Article  CAS  PubMed  Google Scholar 

  9. Davalos V, Moutinho C, Villanueva A, Boque R, Silva P, Carneiro F et al. Dynamic epigenetic regulation of the microRNA-200 family mediates epithelial and mesenchymal transitions in human tumorigenesis. Oncogene 2012; 31: 2062–2074.

    Article  CAS  PubMed  Google Scholar 

  10. Kundu ST, Byers LA, Peng DH, Roybal JD, Diao L, Wang J et al. The miR-200 family and the miR-183~96~182 cluster target Foxf2 to inhibit invasion and metastasis in lung cancers. Oncogene 2015; 35: 173–186.

    Article  PubMed  PubMed Central  Google Scholar 

  11. Roybal JD, Zang Y, Ahn YH, Yang Y, Gibbons DL, Baird BN et al. miR-200 Inhibits lung adenocarcinoma cell invasion and metastasis by targeting Flt1/VEGFR1. Mol Cancer Res 2011; 9: 25–35.

    Article  CAS  PubMed  Google Scholar 

  12. Levental KR, Yu H, Kass L, Lakins JN, Egeblad M, Erler JT et al. Matrix crosslinking forces tumor progression by enhancing integrin signaling. Cell 2009; 139: 891–906.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Egeblad M, Rasch MG, Weaver VM . Dynamic interplay between the collagen scaffold and tumor evolution. Curr Opin Cell Biol 2010; 22: 697–706.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Streuli CH, Bailey N, Bissell MJ . Control of mammary epithelial differentiation - basement-membrane induces tissue-specific gene-expression in the absence of cell cell-interaction and morphological polarity. J Cell Biol 1991; 115: 1383–1395.

    Article  CAS  PubMed  Google Scholar 

  15. Gill BJ, Gibbons DL, Roudsari LC, Saik JE, Rizvi ZH, Roybal JD et al. A synthetic matrix with independently tunable biochemistry and mechanical properties to study epithelial morphogenesis and EMT in a lung adenocarcinoma model. Cancer Res 2012; 72: 6013–6023.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Gilkes DM, Semenza GL, Wirtz D . Hypoxia and the extracellular matrix: drivers of tumour metastasis. Nat Rev Cancer 2014; 14: 430–439.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Ungewiss C, Rizvi ZH, Roybal JD, Peng DH, Gold KA, Shin DH et al. The microRNA-200/Zeb1 axis regulates ECM-dependent beta1-integrin/FAK signaling, cancer cell invasion and metastasis through CRKL. Sci Rep 2016; 6: 18652.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Jung HY, Fattet L, Yang J . Molecular pathways: linking tumor microenvironment to epithelial-mesenchymal transition in metastasis. Clin Cancer Res 2015; 21: 962–968.

    Article  CAS  PubMed  Google Scholar 

  19. Shintani Y, Maeda M, Chaika N, Johnson KR, Wheelock MJ . Collagen I promotes epithelial-to-mesenchymal transition in lung cancer cells via transforming growth factor-beta signaling. Am J Respir Cell Mol Biol 2008; 38: 95–104.

    Article  CAS  PubMed  Google Scholar 

  20. Leight JL, Wozniak MA, Chen S, Lynch ML, Chen CS . Matrix rigidity regulates a switch between TGF-beta1-induced apoptosis and epithelial-mesenchymal transition. Mol Biol Cell 2012; 23: 781–791.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Paszek MJ, Zahir N, Johnson KR, Lakins JN, Rozenberg GI, Gefen A et al. Tensional homeostasis and the malignant phenotype. Cancer Cell 2005; 8: 241–254.

    Article  CAS  PubMed  Google Scholar 

  22. Erler JT, Bennewith KL, Nicolau M, Dornhofer N, Kong C, Le QT et al. Lysyl oxidase is essential for hypoxia-induced metastasis. Nature 2006; 440: 1222–1226.

    Article  CAS  PubMed  Google Scholar 

  23. Cox TR, Bird D, Baker AM, Barker HE, Ho MW, Lang G et al. LOX-mediated collagen crosslinking is responsible for fibrosis-enhanced metastasis. Cancer Res 2013; 73: 1721–1732.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Kagan HM, Li W . Lysyl oxidase: properties, specificity, and biological roles inside and outside of the cell. J Cell Biochem 2003; 88: 660–672.

    Article  CAS  PubMed  Google Scholar 

  25. Barker HE, Cox TR, Erler JT . The rationale for targeting the LOX family in cancer. Nat Rev Cancer 2012; 12: 540–552.

    Article  CAS  PubMed  Google Scholar 

  26. Gao Y, Xiao Q, Ma H, Li L, Liu J, Feng Y et al. LKB1 inhibits lung cancer progression through lysyl oxidase and extracellular matrix remodeling. Proc Natl Acad Sci USA 2010; 107: 18892–18897.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Han X, Li F, Fang Z, Gao Y, Li F, Fang R et al. Transdifferentiation of lung adenocarcinoma in mice with Lkb1 deficiency to squamous cell carcinoma. Nat Commun 2014; 5: 3261.

    Article  PubMed  Google Scholar 

  28. Byers LA, Diao L, Wang J, Saintigny P, Girard L, Peyton M et al. An epithelial-mesenchymal transition gene signature predicts resistance to EGFR and PI3K inhibitors and identifies Axl as a therapeutic target for overcoming EGFR inhibitor resistance. Clin Cancer Res 2013; 19: 279–290.

    Article  CAS  PubMed  Google Scholar 

  29. Barry-Hamilton V, Spangler R, Marshall D, McCauley S, Rodriguez HM, Oyasu M et al. Allosteric inhibition of lysyl oxidase-like-2 impedes the development of a pathologic microenvironment. Nat Med 2010; 16: 1009–1017.

    Article  CAS  PubMed  Google Scholar 

  30. Pickup MW, Laklai H, Acerbi I, Owens P, Gorska AE, Chytil A et al. Stromally derived lysyl oxidase promotes metastasis of transforming growth factor-beta-deficient mouse mammary carcinomas. Cancer Res 2013; 73: 5336–5346.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Peng L, Ran YL, Hu H, Yu L, Liu Q, Zhou Z et al. Secreted LOXL2 is a novel therapeutic target that promotes gastric cancer metastasis via the Src/FAK pathway. Carcinogenesis 2009; 30: 1660–1669.

    Article  CAS  PubMed  Google Scholar 

  32. Ahn YH, Gibbons DL, Chakravarti D, Creighton CJ, Rizvi ZH, Adams HP et al. ZEB1 drives prometastatic actin cytoskeletal remodeling by downregulating miR-34a expression. J Clin Invest 2012; 122: 3170–3183.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Chen L, Gibbons DL, Goswami S, Cortez MA, Ahn YH, Byers LA et al. Metastasis is regulated via microRNA-200/ZEB1 axis control of tumour cell PD-L1 expression and intratumoral immunosuppression. Nat Commun 2014; 5: 5241.

    Article  CAS  PubMed  Google Scholar 

  34. Yang Y, Ahn YH, Chen Y, Tan X, Guo L, Gibbons DL et al. ZEB1 sensitizes lung adenocarcinoma to metastasis suppression by PI3K antagonism. J Clin Invest 2014; 124: 2696–2708.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Yang Y, Ahn YH, Gibbons DL, Zang Y, Lin W, Thilaganathan N et al. The Notch ligand Jagged2 promotes lung adenocarcinoma metastasis through a miR-200-dependent pathway in mice. J Clin Invest 2011; 121: 1373–1385.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Palamakumbura AH, Trackman PC . A fluorometric assay for detection of lysyl oxidase enzyme activity in biological samples. Anal Biochem 2002; 300: 245–251.

    Article  CAS  PubMed  Google Scholar 

  37. Vadasz Z, Kessler O, Akiri G, Gengrinovitch S, Kagan HM, Baruch Y et al. Abnormal deposition of collagen around hepatocytes in Wilson's disease is associated with hepatocyte specific expression of lysyl oxidase and lysyl oxidase like protein-2. J Hepatol 2005; 43: 499–507.

    Article  CAS  PubMed  Google Scholar 

  38. Hollosi P, Yakushiji JK, Fong KS, Csiszar K, Fong SF . Lysyl oxidase-like 2 promotes migration in noninvasive breast cancer cells but not in normal breast epithelial cells. Int J Cancer 2009; 125: 318–327.

    Article  CAS  PubMed  Google Scholar 

  39. Zhan P, Shen XK, Qian Q, Zhu JP, Zhang Y, Xie HY et al. Down-regulation of lysyl oxidase-like 2 (LOXL2) is associated with disease progression in lung adenocarcinomas. Med Oncol 2012; 29: 648–655.

    Article  CAS  PubMed  Google Scholar 

  40. Payne SL, Hendrix MJ, Kirschmann DA . Paradoxical roles for lysyl oxidases in cancer–a prospect. J Cell Biochem 2007; 101: 1338–1354.

    Article  CAS  PubMed  Google Scholar 

  41. Hong HH, Pischon N, Santana RB, Palamakumbura AH, Chase HB, Gantz D et al. A role for lysyl oxidase regulation in the control of normal collagen deposition in differentiating osteoblast cultures. J Cell Physiol 2004; 200: 53–62.

    Article  CAS  PubMed  Google Scholar 

  42. Peinado H, Moreno-Bueno G, Hardisson D, Perez-Gomez E, Santos V, Mendiola M et al. Lysyl oxidase-like 2 as a new poor prognosis marker of squamous cell carcinomas. Cancer Res 2008; 68: 4541–4550.

    Article  CAS  PubMed  Google Scholar 

  43. Kalluri R, Weinberg RA . The basics of epithelial-mesenchymal transition. J Clin Invest 2009; 119: 1420–1428.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Alderton GK . Metastasis: epithelial to mesenchymal and back again. Nat Rev Cancer 2013; 13: 3.

    Article  PubMed  Google Scholar 

  45. Xiao D, He J . Epithelial mesenchymal transition and lung cancer. J Thorac Dis 2010; 2: 154–159.

    CAS  PubMed  PubMed Central  Google Scholar 

  46. Baker AM, Cox TR, Bird D, Lang G, Murray GI, Sun XF et al. The role of lysyl oxidase in SRC-dependent proliferation and metastasis of colorectal cancer. J Natl Cancer Inst 2011; 103: 407–424.

    Article  CAS  PubMed  Google Scholar 

  47. Barker HE, Bird D, Lang G, Erler JT . Tumor-secreted LOXL2 activates fibroblasts through FAK signaling. Mol Cancer Res 2013; 11: 1425–1436.

    Article  CAS  PubMed  Google Scholar 

  48. Barker HE, Chang J, Cox TR, Lang G, Bird D, Nicolau M et al. LOXL2-mediated matrix remodeling in metastasis and mammary gland involution. Cancer Res 2011; 71: 1561–1572.

    Article  CAS  PubMed  Google Scholar 

  49. Fong SF, Dietzsch E, Fong KS, Hollosi P, Asuncion L, He Q et al. Lysyl oxidase-like 2 expression is increased in colon and esophageal tumors and associated with less differentiated colon tumors. Genes Chromosomes Cancer 2007; 46: 644–655.

    Article  CAS  PubMed  Google Scholar 

  50. Lucero HA, Kagan HM . Lysyl oxidase: an oxidative enzyme and effector of cell function. Cell Mol Life Sci 2006; 63: 2304–2316.

    Article  CAS  PubMed  Google Scholar 

  51. Canesin G, Cuevas EP, Santos V, Lopez-Menendez C, Moreno-Bueno G, Huang Y et al. Lysyl oxidase-like 2 (LOXL2) and E47 EMT factor: novel partners in E-cadherin repression and early metastasis colonization. Oncogene 2015; 34: 951–964.

    Article  CAS  PubMed  Google Scholar 

  52. Peinado H, Del Carmen Iglesias-de la Cruz M, Olmeda D, Csiszar K, Fong KS, Vega S et al. A molecular role for lysyl oxidase-like 2 enzyme in snail regulation and tumor progression. EMBO J 2005; 24: 3446–3458.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Akbani R, Ng PK, Werner HM, Shahmoradgoli M, Zhang F, Ju Z et al. A pan-cancer proteomic perspective on The Cancer Genome Atlas. Nat Commun 2014; 5: 3887.

    Article  CAS  PubMed  Google Scholar 

  54. Hoadley KA, Yau C, Wolf DM, Cherniack AD, Tamborero D, Ng S et al. Multiplatform analysis of 12 cancer types reveals molecular classification within and across tissues of origin. Cell 2014; 158: 929–944.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Sato M, Larsen JE, Lee W, Sun H, Shames DS, Dalvi MP et al. Human lung epithelial cells progressed to malignancy through specific oncogenic manipulations. Mol Cancer Res 2013; 11: 638–650.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Oliver WC, Pharr GM . An improved technique for determining hardness and elastic-modulus using load and displacement sensing indentation experiments. J Mater Res 1992; 7: 1564–1583.

    Article  CAS  Google Scholar 

  57. Rossi MA, Abreu MA, Santoro LB . Connective tissue skeleton of the human heart: a demonstration by cell-maceration scanning electron microscope method. Circulation 1998; 97: 934–935.

    Article  CAS  PubMed  Google Scholar 

  58. Chaudhuri BB, Kundu P, Sarkar N . Detection and gradation of oriented texture. Pattern Recogn Lett 1993; 14: 147–153.

    Article  Google Scholar 

Download references

Acknowledgements

We thank Dr G Goodall (University of Adelaide, Australia) for the kind gift of the pTRIPz-miR-200 expression constructs. We thank Dr M Dickinson (Baylor College of Medicine, Houston, TX, USA) for providing access to the multiphoton microscope for SHG analysis. We would like to thank members of the Gibbons lab for their assistance and critical reading of the manuscript. This work was supported by NCI K08 CA151661 (DLG), an MD Anderson Cancer Center Physician Scientist Award (DLG), Rexanna’s Foundation for Fighting Lung Cancer (DLG) and CPRIT grant RP120713 P2 (DLG and JG-A). DP was supported by a CPRIT Graduate Scholar Training Grant (RP140106). JW and PT are supported by Lung SPORE (P50 CA070907), Cancer Center Support Grant (CCSG CA016672) and Mary K Chapman Foundation. CJC was supported by CPRIT grant RP120713 C2 and NCI grant CA125123. LAB and DLG are R Lee Clark Fellows of the University of Texas MD Anderson Cancer Center, supported by the Jeane F Shelby Scholarship Fund.

Author information

Authors and Affiliations

  1. Department of Thoracic/Head and Neck Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA

    D H Peng, C Ungewiss, L A Byers, C Behrens & D L Gibbons

  2. The University of Texas Graduate School of Biomedical Sciences, Houston, TX, USA

    D H Peng & C Ungewiss

  3. Department of Bioinformatics and Computational Biology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA

    P Tong, J Wang & C J Creighton

  4. Department of Translational Molecular Pathology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA

    J R Canales, P A Villalobos, N Uraoka, B Mino & I I Wistuba

  5. Department of Bioengineering, Rice University, Houston, TX, USA

    R I Han, C A Wanna, M Fahrenholtz & K J Grande-Allen

  6. Department of Medicine and Dan L Duncan Cancer Center, Baylor College of Medicine, Houston, TX, USA

    C J Creighton

  7. Department of Molecular and Cellular Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA

    D L Gibbons

Authors
  1. D H Peng
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. C Ungewiss
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. P Tong
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. L A Byers
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. J Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. J R Canales
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. P A Villalobos
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. N Uraoka
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. B Mino
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. C Behrens
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. I I Wistuba
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. R I Han
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. C A Wanna
    View author publications

    You can also search for this author in PubMed Google Scholar

  14. M Fahrenholtz
    View author publications

    You can also search for this author in PubMed Google Scholar

  15. K J Grande-Allen
    View author publications

    You can also search for this author in PubMed Google Scholar

  16. C J Creighton
    View author publications

    You can also search for this author in PubMed Google Scholar

  17. D L Gibbons
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to D L Gibbons.

Ethics declarations

Competing interests

The authors declare no conflict of interest.

Additional information

Supplementary Information accompanies this paper on the Oncogene website

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Peng, D., Ungewiss, C., Tong, P. et al. ZEB1 induces LOXL2-mediated collagen stabilization and deposition in the extracellular matrix to drive lung cancer invasion and metastasis. Oncogene 36, 1925–1938 (2017). https://doi.org/10.1038/onc.2016.358

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/onc.2016.358

This article is cited by

Search

Advanced search

Quick links