Article | Published:

Stratifin regulates stabilization of receptor tyrosine kinases via interaction with ubiquitin-specific protease 8 in lung adenocarcinoma

Oncogenevolume 37pages53875402 (2018) | Download Citation

Abstract

Previously we have reported that stratifin (SFN, 14-3-3 sigma) acts as a novel oncogene, accelerating the tumor initiation and progression of lung adenocarcinoma. Here, pull-down assay and LC-MS/MS analysis revealed that ubiquitin-specific protease 8 (USP8) specifically bound to SFN in lung adenocarcinoma cells. Both USP8 and SFN showed higher expression in human lung adenocarcinoma than in normal lung tissue, and USP8 expression was significantly correlated with SFN expression. Expression of SFN, but not of USP8, was associated with histological subtype, pathological stage, and poor prognosis. USP8 stabilizes receptor tyrosine kinases (RTKs) such as EGFR and MET by deubiquitination, contributing to the proliferative activity of many human cancers including non-small cell lung cancer. In vitro, USP8 binds to SFN and they co-localize at the early endosomes in lung adenocarcinoma cells. Moreover, USP8 or SFN knockdown leads to downregulation of tumor cellular proliferation and upregulation of apoptosis, p-EGFR or p-MET, which are related to the degradation pathway, and accumulation of ubiquitinated RTKs, leading to lysosomal degradation. Additionally, mutant USP8, which is unable to bind to SFN, reduces the expression of RTKs and p-STAT3. We also found that interaction with SFN is critical for USP8 to exert its autodeubiquitination function and avoid dephosphorylation by PP1. Our findings demonstrate that SFN enhances RTK stabilization through abnormal USP8 regulation in lung adenocarcinoma, suggesting that SFN could be a more suitable therapeutic target for lung adenocarcinoma than USP8.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

References

  1. 1.

    Siegel RL, Miller KD, Jemal A. Cancer statistics, 2016. CA Cancer J Clin. 2016;66:7–30.

  2. 2.

    Noguchi M, Morikawa A, Kawasaki M, Matsuno Y, Yamada T, Hirohashi S, et al. Small adenocarcinoma of the lung. Histologic characteristics and prognosis. Cancer. 1995;75:2844–52.

  3. 3.

    Kakinuma R, Noguchi M, Ashizawa K, Kuriyama K, Maeshima AM, Koizumi N, et al. Natural history of pulmonary subsolid nodules: a prospective multicenter study. J Thorac Oncol. 2016;11:1012–28.

  4. 4.

    Shiba-Ishii A, Kano J, Morishita Y, Sato Y, Minami Y, Noguchi M. High expression of stratifin is a universal abnormality during the course of malignant progression of early-stage lung adenocarcinoma. Int J Cancer. 2011;129:2445–53.

  5. 5.

    Shiba-Ishii A, Kim Y, Shiozawa T, Iyama S, Satomi K, Kano J, et al. Stratifin accelerates progression of lung adenocarcinoma at an early stage. Mol Cancer. 2015;14:142.

  6. 6.

    Aghazadeh Y, Papadopoulos V. The role of the 14-3-3 protein family in health, disease, and drug development. Drug Discov Today. 2016;21:278–87.

  7. 7.

    Yaffe MB, Rittinger K, Volinia S, Caron PR, Aitken A, Leffers H, et al. The structural basis for 14-3-3:phosphopeptide binding specificity. Cell. 1997;91:961–71.

  8. 8.

    Hermeking H. The 14-3-3 cancer connection. Nat Rev Cancer. 2003;3:931–43.

  9. 9.

    Smith AJ, Daut J, Schwappach B. Membrane proteins as 14-3-3 clients in functional regulation and intracellular transport. Physiology. 2011;26:181–91.

  10. 10.

    Dougherty MK, Morrison DK. Unlocking the code of 14-3-3. J Cell Sci. 2004;117:1875–84.

  11. 11.

    Fraile JM, Quesada V, Rodriguez D, Freije JM, Lopez-Otin C. Deubiquitinases in cancer: new functions and therapeutic options. Oncogene. 2012;31:2373–88.

  12. 12.

    Xia R, Jia H, Fan J, Liu Y, Jia J. USP8 promotes smoothened signaling by preventing its ubiquitination and changing its subcellular localization. PLoS Biol. 2012;10:e1001238.

  13. 13.

    Mukai A, Yamamoto-Hino M, Awano W, Watanabe W, Komada M, Goto S. Balanced ubiquitylation and deubiquitylation of Frizzled regulate cellular responsiveness to Wg/Wnt. EMBO J. 2010;29:2114–25.

  14. 14.

    Wu X, Yen L, Irwin L, Sweeney C, Carraway KL 3rd. Stabilization of the E3 ubiquitin ligase Nrdp1 by the deubiquitinating enzyme USP8. Mol Cell Biol. 2004;24:7748–57.

  15. 15.

    Mizuno E, Iura T, Mukai A, Yoshimori T, Kitamura N, Komada M. Regulation of epidermal growth factor receptor down-regulation by UBPY-mediated deubiquitination at endosomes. Mol Biol Cell. 2005;16:5163–74.

  16. 16.

    Meijer IM, van Leeuwen JE. ERBB2 is a target for USP8-mediated deubiquitination. Cell Signal. 2011;23:458–67.

  17. 17.

    Smith GA, Fearnley GW, Abdul-Zani I, Wheatcroft SB, Tomlinson DC, Harrison MA, et al. VEGFR2 trafficking, signaling and proteolysis is regulated by the ubiquitin isopeptidase USP8. Traffic. 2016;17:53–65.

  18. 18.

    Niendorf S, Oksche A, Kisser A, Lohler J, Prinz M, Schorle H, et al. Essential role of ubiquitin-specific protease 8 for receptor tyrosine kinase stability and endocytic trafficking in vivo. Mol Cell Biol. 2007;27:5029–39.

  19. 19.

    Takeuchi K, Ito F. Receptor tyrosine kinases and targeted cancer therapeutics. Biol Pharm Bull. 2011;34:1774–80.

  20. 20.

    Kim Y, Shiba-Ishii A, Nakagawa T, Husni RE, Sakashita S, Takeuchi T, et al. Ubiquitin-specific protease 8 is a novel prognostic marker in early-stage lung adenocarcinoma. Pathol Int. 2017;67:292–301.

  21. 21.

    Reincke M, Sbiera S, Hayakawa A, Theodoropoulou M, Osswald A, Beuschlein F, et al. Mutations in the deubiquitinase gene USP8 cause Cushing’s disease. Nat Genet. 2015;47:31–8.

  22. 22.

    Ma ZY, Song ZJ, Chen JH, Wang YF, Li SQ, Zhou LF, et al. Recurrent gain-of-function USP8 mutations in Cushing’s disease. Cell Res. 2015;25:306–17.

  23. 23.

    Ballif BA, Cao Z, Schwartz D, Carraway KL 3rd, Gygi SP. Identification of 14-3-3epsilon substrates from embryonic murine brain. J Proteome Res. 2006;5:2372–9.

  24. 24.

    Dufner A, Kisser A, Niendorf S, Basters A, Reissig S, Schonle A, et al. The ubiquitin-specific protease USP8 is critical for the development and homeostasis of T cells. Nat Immunol. 2015;16:950–60.

  25. 25.

    Mizuno E, Kitamura N, Komada M. 14-3-3-dependent inhibition of the deubiquitinating activity of UBPY and its cancellation in the M phase. Exp Cell Res. 2007;313:3624–34.

  26. 26.

    Benzinger A, Muster N, Koch HB, Yates JR 3rd, Hermeking H. Targeted proteomic analysis of 14-3-3 sigma, a p53 effector commonly silenced in cancer. Mol Cell Proteom. 2005;4:785–95.

  27. 27.

    Shimada A, Kano J, Ishiyama T, Okubo C, Iijima T, Morishita Y, et al. Establishment of an immortalized cell line from a precancerous lesion of lung adenocarcinoma, and genes highly expressed in the early stages of lung adenocarcinoma development. Cancer Sci. 2005;96:668–75.

  28. 28.

    Mei Y, Hahn AA, Hu S, Yang X. The USP19 deubiquitinase regulates the stability of c-IAP1 and c-IAP2. J Biol Chem. 2011;286:35380–7.

  29. 29.

    Huang WG, Cheng AL, Chen ZC, Peng F, Zhang PF, Li MY, et al. Targeted proteomic analysis of 14-3-3sigma in nasopharyngeal carcinoma. Int J Biochem Cell Biol. 2010;42:137–47.

  30. 30.

    Byun S, Lee SY, Lee J, Jeong CH, Farrand L, Lim S, et al. USP8 is a novel target for overcoming gefitinib resistance in lung cancer. Clin Cancer Res. 2013;19:3894–904.

  31. 31.

    Li Z, Liu JY, Zhang JT. 14-3-3sigma, the double-edged sword of human cancers. Am J Transl Res. 2009;1:326–40.

  32. 32.

    Sbiera S, Deutschbein T, Weigand I, Reincke M, Fassnacht M, Allolio B. The new molecular landscape of Cushing’s disease. Trends Endocrinol Metab. 2015;26:573–83.

  33. 33.

    Panner A, Crane CA, Weng C, Feletti A, Fang S, Parsa AT, et al. Ubiquitin-specific protease 8 links the PTEN-Akt-AIP4 pathway to the control of FLIPS stability and TRAIL sensitivity in glioblastoma multiforme. Cancer Res. 2010;70:5046–53.

  34. 34.

    Wilker EW, Grant RA, Artim SC, Yaffe MB. A structural basis for 14-3-3sigma functional specificity. J Biol Chem. 2005;280:18891–8.

  35. 35.

    Shiba-Ishii A, Noguchi M. Aberrant stratifin overexpression is regulated by tumor-associated CpG demethylation in lung adenocarcinoma. Am J Pathol. 2012;180:1653–62.

  36. 36.

    Kim JO, Kim SR, Lim KH, Kim JH, Ajjappala B, Lee HJ, et al. Deubiquitinating enzyme USP37 regulating oncogenic function of 14-3-3gamma. Oncotarget. 2015;6:36551–76.

  37. 37.

    Alwan HA, van Leeuwen JE. UBPY-mediated epidermal growth factor receptor (EGFR) de-ubiquitination promotes EGFR degradation. J Biol Chem. 2007;282:1658–69.

  38. 38.

    Savio MG, Wollscheid N, Cavallaro E, Algisi V, Di Fiore PP, Sigismund S, et al. USP9X controls EGFR fate by deubiquitinating the endocytic adaptor Eps15. Curr Biol. 2016;26:173–83.

  39. 39.

    Pareja F, Ferraro DA, Rubin C, Cohen-Dvashi H, Zhang F, Aulmann S, et al. Deubiquitination of EGFR by Cezanne-1 contributes to cancer progression. Oncogene. 2012;31:4599–608.

  40. 40.

    Clague MJ, Urbe S. Endocytosis: the DUB version. Trends Cell Biol. 2006;16:551–9.

  41. 41.

    Row PE, Prior IA, McCullough J, Clague MJ, Urbe S. The ubiquitin isopeptidase UBPY regulates endosomal ubiquitin dynamics and is essential for receptor down-regulation. J Biol Chem. 2006;281:12618–24.

  42. 42.

    Komada M. Controlling receptor downregulation by ubiquitination and deubiquitination. Curr Drug Discov Technol. 2008;5:78–84.

  43. 43.

    Ledda F, Paratcha G. Negative regulation of receptor tyrosine kinase (RTK) signaling: a developing field. Biomark Insights. 2007;2:45–58.

  44. 44.

    Guo L, Kozlosky CJ, Ericsson LH, Daniel TO, Cerretti DP, Johnson RS. Studies of ligand-induced site-specific phosphorylation of epidermal growth factor receptor. J Am Soc Mass Spectrom. 2003;14:1022–31.

  45. 45.

    Perez-Rivas LG, Theodoropoulou M, Ferrau F, Nusser C, Kawaguchi K, Stratakis CA, et al. The gene of the ubiquitin-specific protease 8 is frequently mutated in adenomas causing Cushing’s disease. J Clin Endocrinol Metab. 2015;100:E997–1004.

  46. 46.

    Meijer IM, Kerperien J, Sotoca AM, van Zoelen EJ, van Leeuwen JE. The Usp8 deubiquitination enzyme is post-translationally modified by tyrosine and serine phosphorylation. Cell Signal. 2013;25:919–30.

  47. 47.

    Devarakonda S, Morgensztern D, Govindan R. Genomic alterations in lung adenocarcinoma. Lancet Oncol. 2015;16:e342–51.

  48. 48.

    Bansal P, Osman D, Gan GN, Simon GR, Boumber Y. Recent advances in targetable therapeutics in metastatic non-squamous NSCLC. Front Oncol. 2016;6:112.

  49. 49.

    Naviglio S, Mattecucci C, Matoskova B, Nagase T, Nomura N, Di Fiore PP, et al. UBPY: a growth-regulated human ubiquitin isopeptidase. EMBO J. 1998;17:3241–50.

  50. 50.

    Sato T, Shiba-Ishii A, Kim Y, Dai T, Husni RE, Hong J, et al. miR-3941: a novel microRNA that controls IGBP1 expression and is associated with malignant progression of lung adenocarcinoma. Cancer Sci. 2017;108:536–42.

  51. 51.

    Travis WD, Brambilla E, Noguchi M, Nicholson AG, Geisinger KR, Yatabe Y, et al. International association for the study of lung cancer/american thoracic society/european respiratory society international multidisciplinary classification of lung adenocarcinoma. J Thorac Oncol. 2011;6:244–85.

  52. 52.

    Travis WD, Brambilla E, Nicholson AG, Yatabe Y, Austin JH, Beasley MB, et al. The2015 world health organization classification of lung tumors: impact of genetic, clinical and radiologic advances since the 2004 classification. J Thorac Oncol. 2015;10:1243–60.

Download references

Acknowledgements

We express our appreciation to Professor Yasunori Kanaho and Dr. Yuji Funakoshi for research support and kindly providing the plasmids and reagents. We also thank Professor Mitsuyasu Kato and Dr. Hiroyuki Suzuki (Faculty of Medicine, University of Tsukuba) for research support and Professor Flaminia Miyamasu (Medical English Communications Center, University of Tsukuba) for critical review of this manuscript.

Author information

Author notes

  1. These authors contributed equally: Yunjung Kim, Aya Shiba-Ishii

Affiliations

  1. Department of Pathology, Faculty of Medicine, University of Tsukuba, Ibaraki, Japan

    • Yunjung Kim
    • , Aya Shiba-Ishii
    • , Noriyuki Nakano
    • , Ryota Matsuoka
    • , Shingo Sakashita
    •  & Masayuki Noguchi
  2. Doctoral Program in Biomedical Sciences, Graduate School of Comprehensive Human Sciences, University of Tsukuba, Ibaraki, Japan

    • Tomoki Nakagawa
  3. Translational Research Center, Fukushima Medical University, Fukushima, Japan

    • Shun-ichiro Iemura
  4. Molecular Profiling Research Center for Drug Discovery, National Institutes of Advanced Industrial Science and Technology, Tokyo, Japan

    • Tohru Natsume
  5. Ph.D. Program in Human Biology, School of Integrative and Global Majors, University of Tsukuba, Ibaraki, Japan

    • SangJoon Lee
  6. Department of Infection Biology, Faculty of Medicine, University of Tsukuba, Ibaraki, Japan

    • Atsushi Kawaguchi
  7. Department of Thoracic Surgery, Faculty of Medicine, University of Tsukuba, Ibaraki, Japan

    • Yukio Sato

Authors

  1. Search for Yunjung Kim in:

  2. Search for Aya Shiba-Ishii in:

  3. Search for Tomoki Nakagawa in:

  4. Search for Shun-ichiro Iemura in:

  5. Search for Tohru Natsume in:

  6. Search for Noriyuki Nakano in:

  7. Search for Ryota Matsuoka in:

  8. Search for Shingo Sakashita in:

  9. Search for SangJoon Lee in:

  10. Search for Atsushi Kawaguchi in:

  11. Search for Yukio Sato in:

  12. Search for Masayuki Noguchi in:

Conflict of interest

The authors declare that they have no conflict of interest.

Corresponding authors

Correspondence to Aya Shiba-Ishii or Masayuki Noguchi.

Electronic supplementary material

About this article

Publication history

Received

Revised

Accepted

Published

DOI

https://doi.org/10.1038/s41388-018-0342-9