Original Article

Desmoglein 3 promotes cancer cell migration and invasion by regulating activator protein 1 and protein kinase C-dependent-Ezrin activation

  • Oncogene (2014) 33, 23632374 (01 May 2014)
  • doi:10.1038/onc.2013.186
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Abstract

Desmoglein 3 (Dsg3), the pemphigus vulgaris antigen, has recently been shown to be upregulated in squamous cell carcinoma (SCC) and has been identified as a good tumor-specific marker for clinical staging of cervical sentinel lymph nodes in head and neck SCC. However, little is known about its biological function in cancer. The actin-binding protein Ezrin and the activator protein 1 (AP-1) transcription factor are implicated in cancer progression and metastasis. Here, we report that Dsg3 regulates the activity of c-Jun/AP-1 as well as protein kinase C (PKC)-mediated phosphorylation of Ezrin-Thr567, which contributes to the accelerated motility of cancer cells. Ectopic expression of Dsg3 in cancer cell lines caused enhanced phosphorylation at Ezrin-Thr567 with concomitant augmented membrane protrusions, cell spreading and invasive phenotype. We showed that Dsg3 formed a complex with Ezrin at the plasma membrane that was required for its proper function of interacting with F-actin and CD44 as Dsg3 knockdown impaired these associations. The increased Ezrin phosphorylation in Dsg3-overexpressing cells could be abrogated substantially by various pharmacological inhibitors for Ser/Thr kinases, including PKC and Rho kinase that are known to activate Ezrin. Furthermore, a marked increase in c-Jun S63 phosphorylation, among others, was found in Dsg3-overexpressing cells and the activation of c-Jun/AP-1 was further supported by a luciferase reporter assay. Taken together, our study identifies a novel Dsg3-mediated c-Jun/AP-1 regulatory mechanism and PKC-dependent Ezrin phosphorylation that could be responsible for Dsg3-associated cancer metastasis.

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References

  1. 1.

    , , . Autoantibodies against a novel epithelial cadherin in pemphigus vulgaris, a disease of cell adhesion. Cell 1991; 67: 869–877.

  2. 2.

    , , , . Pemphigus vulgaris antigen is a desmosomal desmoglein. Dermatology 1994; 189(Suppl 1): 24–26.

  3. 3.

    , , , , . Extracellular domain of pemphigus vulgaris antigen (desmoglein 3) mediates weak homophilic adhesion. J Invest Dermatol 1994; 103: 609–615.

  4. 4.

    , , , , , et al. Desmoglein endocytosis and desmosome disassembly are coordinated responses to pemphigus autoantibodies. J Biol Chem 2006; 281: 7623–7634.

  5. 5.

    , , , , , . Pemphigus vulgaris IgG-induced desmoglein-3 endocytosis and desmosomal disassembly are mediated by a clathrin- and dynamin-independent mechanism. J Biol Chem 2008; 283: 18303–18313.

  6. 6.

    , , , , , et al. P120-catenin is a novel desmoglein 3 interacting partner: identification of the p120-catenin association site of desmoglein 3. Exp Cell Res 2008; 314: 1683–1692.

  7. 7.

    , , , , , et al. Pemphigus vulgaris identifies plakoglobin as key suppressor of c-Myc in the skin. EMBO J 2006; 25: 3298–3309.

  8. 8.

    , , , , , et al. Non-junctional human desmoglein 3 acts as an upstream regulator of Src in E-cadherin adhesion, a pathway possibly involved in the pathogenesis of pemphigus vulgaris. J Pathol 2012; 227: 81–93.

  9. 9.

    , , , , , et al. Desmoglein 3 acting as an upstream regulator of Rho GTPases, Rac-1/Cdc42 in the regulation of actin organisation and dynamics. Exp Cell Res 2012; 318: 2269–2283.

  10. 10.

    , , , , , et al. Desmoglein 3, via an interaction with E-cadherin, is associated with activation of Src. PLoS One 2010; 5: e14211.

  11. 11.

    , . Desmosomes: a role in cancer? Br J Cancer 2007; 96: 1783–1787.

  12. 12.

    , , , , , et al. Immunohistochemical study of desmosomes in oral squamous cell carcinoma: correlation with cytokeratin and E-cadherin staining, and with tumour behaviour. J Pathol 1998; 184: 369–381.

  13. 13.

    , , , , , . A molecular study of desmosomes identifies a desmoglein isoform switch in head and neck squamous cell carcinoma. J Oral Pathol Med 2011; 40: 67–76.

  14. 14.

    , , , , , et al. DSG3 is overexpressed in head neck cancer and is a potential molecular target for inhibition of oncogenesis. Oncogene 2007; 26: 467–476.

  15. 15.

    , , , , , et al. Desmoglein 3 is overexpressed in inverted papilloma and squamous cell carcinoma of sinonasal cavity. Laryngoscope 2010; 120: 26–29.

  16. 16.

    , , , , , et al. The role of desmoglein-3 in the diagnosis of squamous cell carcinoma of the lung. Am J Pathol 2009; 174: 1629–1637.

  17. 17.

    , , , , , et al. DSG3 as a biomarker for the ultrasensitive detection of occult lymph node metastasis in oral cancer using nanostructured immunoarrays. Oral Oncol 2012; 49: 93–101.

  18. 18.

    , , , , , et al. Molecular staging of cervical lymph nodes in squamous cell carcinoma of the head and neck. Cancer Res 2005; 65: 2147–2156.

  19. 19.

    , , , , , et al. Pemphigus vulgaris antigen mRNA quantification for the staging of sentinel lymph nodes in head and neck cancer. Br J Cancer 2010; 102: 181–187.

  20. 20.

    , , , , , et al. Intraoperative qRT-PCR for detection of lymph node metastasis in head and neck cancer. Clin Cancer Res 2011; 17: 1858–1866.

  21. 21.

    . Rapid phosphorylation and reorganization of ezrin and spectrin accompany morphological changes induced in A-431 cells by epidermal growth factor. J Cell Biol 1989; 108: 921–930.

  22. 22.

    . Regulation of cortical structure by the ezrin-radixin-moesin protein family. Curr Opin Cell Biol 1999; 11: 109–116.

  23. 23.

    , , , , , . ERM family members as molecular linkers between the cell surface glycoprotein CD44 and actin-based cytoskeletons. J Cell Biol 1994; 126: 391–401.

  24. 24.

    , . Identification and functional analysis of the ezrin-binding site in the hyaluronan receptor, CD44. Curr Biol 1998; 8: 705–708.

  25. 25.

    , , . Identification of EBP50: A PDZ-containing phosphoprotein that associates with members of the ezrin-radixin-moesin family. J Cell Biol 1997; 139: 169–179.

  26. 26.

    , , , , , et al. Phosphoinositide binding and phosphorylation act sequentially in the activation mechanism of ezrin. J Cell Biol 2004; 164: 653–659.

  27. 27.

    , , , . Rho-dependent and -independent activation mechanisms of ezrin/radixin/moesin proteins: an essential role for polyphosphoinositides in vivo. J Cell Sci 2002; 115: 2569–2580.

  28. 28.

    , , , , , et al. High turnover of ezrin T567 phosphorylation: conformation, activity, and cellular function. Am J Physiol Cell Physiol 2007; 293: C874–C884.

  29. 29.

    , , , , , et al. Protein kinase C regulates ezrin-radixin-moesin phosphorylation in canine osteosarcoma cells. Vet Comp Oncol 2011; 9: 207–218.

  30. 30.

    , . Actin microdomains on endothelial cells: association with CD44, ERM proteins, and signaling molecules during quiescence and wound healing. Histochem Cell Biol 2004; 121: 361–369.

  31. 31.

    , , , , . A novel PKC-regulated mechanism controls CD44 ezrin association and directional cell motility. Nat Cell Biol 2002; 4: 399–407.

  32. 32.

    , , , , , et al. Ezrin is a downstream effector of trafficking PKC-integrin complexes involved in the control of cell motility. EMBO J 2001; 20: 2723–2741.

  33. 33.

    , , , , , et al. The actin-cytoskeleton linker protein ezrin is regulated during osteosarcoma metastasis by PKC. Oncogene 2009; 28: 792–802.

  34. 34.

    , , , , , . Atypical protein kinase C (iota) activates ezrin in the apical domain of intestinal epithelial cells. J Cell Sci 2008; 121: 644–654.

  35. 35.

    , , , , , et al. Rho kinase phosphorylation promotes ezrin-mediated metastasis in hepatocellular carcinoma. Cancer Res 2011; 71: 1721–1729.

  36. 36.

    , , , , , et al. Rho-kinase phosphorylates COOH-terminal threonines of ezrin/radixin/moesin (ERM) proteins and regulates their head-to-tail association. J Cell Biol 1998; 140: 647–657.

  37. 37.

    , , . Downregulated AP-1 activity is associated with inhibition of Protein-Kinase-C-dependent CD44 and ezrin localisation and upregulation of PKC theta in A431 cells. J Cell Sci 2002; 115: 2713–2724.

  38. 38.

    , , , . A possible mechanism for ezrin to establish epithelial cell polarity. Am J Physiol Cell Physiol 2010; 299: C431–C443.

  39. 39.

    , , , , . Ezrin is an effector of hepatocyte growth factor-mediated migration and morphogenesis in epithelial cells. J Cell Biol 1997; 138: 423–434.

  40. 40.

    . Ezrin, a key component in tumor metastasis. Trends Mol Med 2004; 10: 201–204.

  41. 41.

    , , , , , et al. The membrane-cytoskeleton linker ezrin is necessary for osteosarcoma metastasis. Nat Med 2004; 10: 182–186.

  42. 42.

    , , , , , et al. Ezrin promotes ovarian carcinoma cell invasion and its retained expression predicts poor prognosis in ovarian carcinoma. Int J Gynecol Pathol 2006; 25: 121–130.

  43. 43.

    , , , , , . Expression of ezrin is associated with invasion and dedifferentiation of hepatitis B related hepatocellular carcinoma. BMC Cancer 2009; 9: 233.

  44. 44.

    , , , , , et al. Intense cytoplasmic ezrin immunoreactivity predicts poor survival in colorectal cancer. Hum Pathol 2008; 39: 1737–1743.

  45. 45.

    , , , , , . Ezrin promotes invasion and metastasis of pancreatic cancer cells. J Transl Med 2010; 8: 61.

  46. 46.

    , , , . Expression of Ezrin and E-cadherin in nasopharyngeal carcinoma and its significance. Zhong Nan Da Xue Xue Bao Yi Xue Ban 2010; 35: 969–975.

  47. 47.

    , , , , , et al. Expression of the membrane-cytoskeletal linker Ezrin in salivary gland adenoid cystic carcinoma. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2011; 112: 96–104.

  48. 48.

    , , , , . Knockdown of ezrin via RNA interference suppresses Helicobacter pylori-enhanced invasion of gastric cancer cells. Cancer Biol Ther 2011; 11: 746–752.

  49. 49.

    , , , , , et al. Roles of ezrin in the growth and invasiveness of esophageal squamous carcinoma cells. Int J Cancer 2009; 124: 2549–2558.

  50. 50.

    , , , , , . RNAi-mediated silencing of ezrin gene reverses malignant behavior of human gastric cancer cell line SGC-7901. J Dig Dis 2009; 10: 258–264.

  51. 51.

    , , , . Directed actin polymerization is the driving force for epithelial cell-cell adhesion. Cell 2000; 100: 209–219.

  52. 52.

    , , , , , . Development of a quantitative method to analyse tumour cell invasion in organotypic culture. J Pathol 2005; 205: 468–475.

  53. 53.

    , . The ARP2/3 complex: an actin nucleator comes of age. Nat Rev Mol Cell Biol 2006; 7: 713–726.

  54. 54.

    , , , , , et al. N,N'-Dinitrosopiperazine-mediated ezrin phosphorylation via activating Rho kinase and protein kinase C involves in metastasis of nasopharyngeal carcinoma 6-10B cells. J Biol Chem 2011; 286: 36956–36967.

  55. 55.

    , , , . Expression of Ezrin and phosphorylated Ezrin (pEzrin) in pancreatic ductal adenocarcinoma. Cancer Invest 2010; 28: 242–247.

  56. 56.

    , , . Organizing the cell cortex: the role of ERM proteins. Nat Rev Mol Cell Biol 2010; 11: 276–287.

  57. 57.

    , , , , , . Androgen induction of prostate cancer cell invasion is mediated by ezrin. J Biol Chem 2006; 281: 29938–29948.

  58. 58.

    , . Heterotypic and homotypic associations between ezrin and moesin, two putative membrane-cytoskeletal linking proteins. Proc Natl Acad Sci USA 1993; 90: 10846–10850.

  59. 59.

    , , , , , . Protein kinase C alpha/beta inhibitor Go6976 promotes formation of cell junctions and inhibits invasion of urinary bladder carcinoma cells. Cancer Res 2004; 64: 5693–5701.

  60. 60.

    , , , , . Protein kinase C activation disrupts epithelial apical junctions via ROCK-II dependent stimulation of actomyosin contractility. BMC Cell Biol 2009; 10: 36.

  61. 61.

    , , , , , et al. Ezrin, radixin, and moesin are phosphorylated in response to 2-methoxyestradiol and modulate endothelial hyperpermeability. Am J Respir Cell Mol Biol 2011; 45: 1185–1194.

  62. 62.

    , . ROCK and Dia have opposing effects on adherens junctions downstream of Rho. Nat Cell Biol 2002; 4: 408–415.

  63. 63.

    , , , , . Rational design and characterization of a Rac GTPase-specific small molecule inhibitor. Proc Natl Acad Sci USA 2004; 101: 7618–7623.

  64. 64.

    , , , , , et al. Desmosome signaling. Inhibition of p38MAPK prevents pemphigus vulgaris IgG-induced cytoskeleton reorganization. J Biol Chem 2005; 280: 23778–23784.

  65. 65.

    , , , . Transcription factors control invasion: AP-1 the first among equals. Oncogene 2007; 26: 1–10.

  66. 66.

    , , , , , et al. Regulation of a multigenic invasion programme by the transcription factor, AP-1: re-expression of a down-regulated gene, TSC-36, inhibits invasion. Oncogene 2000; 19: 5348–5358.

  67. 67.

    , , , . Co-operative effect of c-Src and ezrin in deregulation of cell-cell contacts and scattering of mammary carcinoma cells. J Cell Biochem 2004; 92: 16–28.

  68. 68.

    , , , , , et al. Initiation of malignancy by duodenal contents reflux and the role of ezrin in developing esophageal squamous cell carcinoma. Cancer Sci 2010; 101: 624–630.

  69. 69.

    , , , , , et al. Ezrin/radixin/moesin (ERM) proteins bind to a positively charged amino acid cluster in the juxta-membrane cytoplasmic domain of CD44, CD43, and ICAM-2. J Cell Biol 1998; 140: 885–895.

  70. 70.

    , , , . Outside-in signaling through integrins and cadherins: a central mechanism to control epidermal growth and differentiation? J Invest Dermatol 2008; 128: 501–516.

  71. 71.

    , , , . Desmosomes and disease: pemphigus and bullous impetigo. Curr Opin Cell Biol 2004; 16: 536–543.

  72. 72.

    , , . A pathogenic autoantibody, pemphigus vulgaris-IgG, induces phosphorylation of desmoglein 3, and its dissociation from plakoglobin in cultured keratinocytes. Eur J Immunol 1999; 29: 2233–2240.

  73. 73.

    , , , , . Actin reorganization contributes to loss of cell adhesion in pemphigus vulgaris. Am J Physiol Cell Physiol 2010; 299: C606–C613.

  74. 74.

    , , . Amphetamine-induced ERM Proteins Phosphorylation Is through PKCbeta Activation in PC12 Cells. Korean J Physiol Pharmacol 2011; 15: 245–249.

  75. 75.

    , , , , . LOK is a major ERM kinase in resting lymphocytes and regulates cytoskeletal rearrangement through ERM phosphorylation. Proc Natl Acad Sci USA 2009; 106: 4707–4712.

  76. 76.

    , , , . Hypotonicity causes actin reorganization and recruitment of the actin-binding ERM protein moesin in membrane protrusions in collecting duct principal cells. Am J Physiol Cell Physiol 2007; 292: C1476–C1484.

  77. 77.

    , , , , , et al. Ezrin/radixin/moesin proteins are phosphorylated by TNF-alpha and modulate permeability increases in human pulmonary microvascular endothelial cells. J Immunol 2006; 176: 1218–1227.

  78. 78.

    , , , . Translocation of diacylglycerol kinase theta from cytosol to plasma membrane in response to activation of G protein-coupled receptors and protein kinase C. J Biol Chem 2005; 280: 9870–9878.

  79. 79.

    , , , , . Ezrin regulates E-cadherin-dependent adherens junction assembly through Rac1 activation. Mol Biol Cell 2003; 14: 2181–2191.

  80. 80.

    , , , , , et al. Psoriasin (S100A7) associates with integrin beta6 subunit and is required for alphavbeta6-dependent carcinoma cell invasion. Oncogene 2011; 30: 1422–1435.

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Acknowledgements

We thank Dr MT Teh, Professor K Parkinson, Dr SM Tsang, Dr MS Ikram and members of CDOS for helpful discussion and technical assistance. This work was supported by a British Skin Foundation-funded studentship awarded to HW and in part by the Institute of Dentistry as well as The Facial Surgery Research Foundation-Saving Faces.

Author information

Affiliations

  1. Queen Mary University of London, Barts and The London School of Medicine and Dentistry, Centre for Clinical and Diagnostic Oral Sciences, Institute of Dentistry, London, UK

    • L Brown
    • , A Waseem
    • , J Szary
    • , E Gunic
    • , T Mannan
    • , M Unadkat
    •  & H Wan
  2. Department of Pharmaceutical and Biological Chemistry, UCL School of Pharmacy, London, UK

    • I N Cruz
    •  & M Yang
  3. Division of Biomedical Sciences, St George’s, University of London, Cranmer Terrace, London, UK

    • F Valderrama
  4. Queen Mary University of London, Barts and The London School of Medicine and Dentistry, Centre for Cutaneous Research, Blizard Institute, London, UK

    • E A O′Toole

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Competing interests

The authors declare no conflict of interest.

Corresponding author

Correspondence to H Wan.

Supplementary information

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