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.

  • Article
  • Published:

The deubiquitylating enzyme USP35 restricts regulated cell death to promote survival of renal clear cell carcinoma

Cell Death & Differentiation volume 30, pages 1757–1770 (2023)Cite this article

Subjects

Abstract

The ubiquitin-proteasome system governs a wide spectrum of cellular events and offers therapeutic opportunities for pharmacological intervention in cancer treatment. Renal clear cell carcinoma represents the predominant histological subtype and accounts for the majority of cancer death related to kidney malignancies. Through a systematic survey in the association of human ubiquitin-specific proteases with patient prognosis of renal clear cell carcinoma and subsequent phenotypic validation, we uncovered the tumor-promoting role of USP35. Biochemical characterizations confirmed the stabilizing effects of USP35 towards multiple members of the IAP family in an enzymatic activity-dependent manner. USP35 silencing led to reduced expression levels of IAP proteins, which were accompanied with increased cellular apoptosis. Further transcriptomic analysis revealed that USP35 knockdown affected the expression levels of NRF2 downstream transcripts, which were conferred by compromised NRF2 abundance. USP35 functions to maintain NRF2 levels by catalyzing its deubiquitylation and thus antagonizing degradation. NRF2 reduction imposed by USP35 silencing rendered renal clear cell carcinoma cells increased sensitivity to ferroptosis induction. Finally, induced USP35 knockdown markedly attenuated xenograft formation of renal clear cell carcinoma in nude mice. Hence, our findings reveal a number of USP35 substrates and uncover the protecting roles of USP35 against both apoptosis and ferroptosis in renal clear cell carcinoma.

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

Fig. 1: USP35 expression predicts adverse prognosis of patients with RCC.
Fig. 2: USP35 stabilizes and associates with multiple IAP proteins.
Fig. 3: USP35 maintains the stability and abates the ubiquitylation of IAP proteins.
Fig. 4: USP35 regulates the expression levels and turnover of endogenous IAP proteins.
Fig. 5: USP35 silencing leads to NRF2 downregulation in RCC cells.
Fig. 6: USP35 silencing renders RCC cells increased sensitivity to ferroptosis.
Fig. 7: The overexpression of NRF2 alleviates increased ferroptotic cell death imposed by USP35 silencing.
Fig. 8: The tumor-promoting role of USP35 in RCC.

Similar content being viewed by others

Data availability

The data supporting the findings of this study are included in the article and supplementary files. RNA-seq data have been deposited in Gene Expression Omnibus of NCBI (accession code GSE216154).

References

  1. Sung H, Ferlay J, Siegel RL, Laversanne M, Soerjomataram I, Jemal A, et al. Global Cancer Statistics 2020: GLOBOCAN Estimates of Incidence and Mortality Worldwide for 36 Cancers in 185 Countries. CA: a cancer J clinicians. 2021;71:209–49.

    Google Scholar 

  2. Motzer RJ, Bander NH, Nanus DM. Renal-cell carcinoma. N. Engl J Med. 1996;335:865–75.

    Article  CAS  PubMed  Google Scholar 

  3. Hsieh JJ, Purdue MP, Signoretti S, Swanton C, Albiges L, Schmidinger M, et al. Renal cell carcinoma. Nat Rev Dis Prim. 2017;3:17009.

    Article  PubMed  Google Scholar 

  4. Cohen P, Tcherpakov M. Will the ubiquitin system furnish as many drug targets as protein kinases? Cell 2010;143:686–93.

    Article  CAS  PubMed  Google Scholar 

  5. Huang X, Dixit VM. Drugging the undruggables: exploring the ubiquitin system for drug development. Cell Res. 2016;26:484–98.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Tokheim C, Wang X, Timms RT, Zhang B, Mena EL, Wang B, et al. Systematic characterization of mutations altering protein degradation in human cancers. Mol cell. 2021;81:1292–308.e11

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Kandoth C, McLellan MD, Vandin F, Ye K, Niu B, Lu C, et al. Mutational landscape and significance across 12 major cancer types. Nature 2013;502:333–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Clark DJ, Dhanasekaran SM, Petralia F, Pan J, Song X, Hu Y, et al. Integrated Proteogenomic Characterization of Clear Cell Renal Cell Carcinoma. Cell 2019;179:964–83.e31

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Dizman N, Philip EJ, Pal SK. Genomic profiling in renal cell carcinoma. Nat Rev Nephrol. 2020;16:435–51.

    Article  PubMed  Google Scholar 

  10. Clague MJ, Barsukov I, Coulson JM, Liu H, Rigden DJ, Urbe S. Deubiquitylases from genes to organism. Physiological Rev. 2013;93:1289–315.

    Article  CAS  Google Scholar 

  11. Duns G, Hofstra RM, Sietzema JG, Hollema H, van Duivenbode I, Kuik A, et al. Targeted exome sequencing in clear cell renal cell carcinoma tumors suggests aberrant chromatin regulation as a crucial step in ccRCC development. Hum Mutat. 2012;33:1059–62.

    Article  CAS  PubMed  Google Scholar 

  12. Pena-Llopis S, Vega-Rubin-de-Celis S, Liao A, Leng N, Pavia-Jimenez A, Wang S, et al. BAP1 loss defines a new class of renal cell carcinoma. Nat Genet. 2012;44:751–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Hong K, Hu L, Liu X, Simon JM, Ptacek TS, Zheng X, et al. USP37 promotes deubiquitination of HIF2alpha in kidney cancer. Proc Natl Acad Sci USA. 2020;117:13023–32.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Clague MJ, Urbe S, Komander D. Breaking the chains: deubiquitylating enzyme specificity begets function. Nat Rev Mol cell Biol. 2019;20:338–52.

    Article  CAS  PubMed  Google Scholar 

  15. Chen D, Ning Z, Chen H, Lu C, Liu X, Xia T, et al. An integrative pan-cancer analysis of biological and clinical impacts underlying ubiquitin-specific-processing proteases. Oncogene 2020;39:587–602.

    Article  CAS  PubMed  Google Scholar 

  16. Liu S, Wang T, Shi Y, Bai L, Wang S, Guo D, et al. USP42 drives nuclear speckle mRNA splicing via directing dynamic phase separation to promote tumorigenesis. Cell death Differ. 2021;28:2482–98.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Wu Y, Zhang Y, Wang D, Zhang Y, Zhang J, Zhang Y, et al. USP29 enhances chemotherapy-induced stemness in non-small cell lung cancer via stabilizing Snail1 in response to oxidative stress. Cell death Dis. 2020;11:796.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Sun Q, Zhang J, Li X, Yang G, Cheng S, Guo D, et al. The ubiquitin-specific protease 8 antagonizes melatonin-induced endocytic degradation of MT(1) receptor to promote lung adenocarcinoma growth. J Adv Res. 2022;41:1–12.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Zhang J, Liu S, Li Q, Shi Y, Wu Y, Liu F, et al. The deubiquitylase USP2 maintains ErbB2 abundance via counteracting endocytic degradation and represents a therapeutic target in ErbB2-positive breast cancer. Cell death Differ. 2020;27:2710–25.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Nagy A, Munkacsy G, Gyorffy B. Pancancer survival analysis of cancer hallmark genes. Sci Rep. 2021;11:6047.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Leznicki P, Natarajan J, Bader G, Spevak W, Schlattl A, Abdul Rehman SA, et al. Expansion of DUB functionality generated by alternative isoforms - USP35, a case study. J Cell Sci. 2018;131:jcs212753.

  22. Yang CS, Sinenko SA, Thomenius MJ, Robeson AC, Freel CD, Horn SR, et al. The deubiquitinating enzyme DUBAI stabilizes DIAP1 to suppress Drosophila apoptosis. Cell death Differ. 2014;21:604–11.

    Article  CAS  PubMed  Google Scholar 

  23. Kumar S, Fairmichael C, Longley DB, Turkington RC. The Multiple Roles of the IAP Super-family in cancer. Pharmacol therapeutics. 2020;214:107610.

    Article  CAS  Google Scholar 

  24. Hayes JD, Dinkova-Kostova AT, Tew KD. Oxidative Stress in Cancer. Cancer cell. 2020;38:167–97.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Hayes JD, Chowdhry S, Dinkova-Kostova AT, Sutherland C. Dual regulation of transcription factor Nrf2 by Keap1 and by the combined actions of beta-TrCP and GSK-3. Biochemical Soc Trans. 2015;43:611–20.

    Article  CAS  Google Scholar 

  26. Baird L, Yamamoto M. The Molecular Mechanisms Regulating the KEAP1-NRF2 Pathway. Mol Cell Biol. 2020;40:e00099-20.

  27. McMahon M, Itoh K, Yamamoto M, Hayes JD. Keap1-dependent proteasomal degradation of transcription factor Nrf2 contributes to the negative regulation of antioxidant response element-driven gene expression. The. J Biol Chem. 2003;278:21592–600.

    Article  CAS  PubMed  Google Scholar 

  28. Itoh K, Wakabayashi N, Katoh Y, Ishii T, O’Connor T, Yamamoto M. Keap1 regulates both cytoplasmic-nuclear shuttling and degradation of Nrf2 in response to electrophiles. Genes Cells: devoted Mol Cell mechanisms. 2003;8:379–91.

    Article  CAS  Google Scholar 

  29. Kobayashi A, Kang MI, Okawa H, Ohtsuji M, Zenke Y, Chiba T, et al. Oxidative stress sensor Keap1 functions as an adaptor for Cul3-based E3 ligase to regulate proteasomal degradation of Nrf2. Mol Cell Biol. 2004;24:7130–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Cullinan SB, Gordan JD, Jin J, Harper JW, Diehl JA. The Keap1-BTB protein is an adaptor that bridges Nrf2 to a Cul3-based E3 ligase: oxidative stress sensing by a Cul3-Keap1 ligase. Mol Cell Biol. 2004;24:8477–86.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Zhang DD, Lo SC, Cross JV, Templeton DJ, Hannink M. Keap1 is a redox-regulated substrate adaptor protein for a Cul3-dependent ubiquitin ligase complex. Mol Cell Biol. 2004;24:10941–53.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Dixon SJ, Lemberg KM, Lamprecht MR, Skouta R, Zaitsev EM, Gleason CE, et al. Ferroptosis: an iron-dependent form of nonapoptotic cell death. Cell 2012;149:1060–72.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Stockwell BR, Jiang X. The Chemistry and Biology of Ferroptosis. Cell Chem Biol. 2020;27:365–75.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Tang D, Chen X, Kang R, Kroemer G. Ferroptosis: molecular mechanisms and health implications. Cell Res. 2021;31:107–25.

    Article  CAS  PubMed  Google Scholar 

  35. Chen X, Kang R, Kroemer G, Tang D. Broadening horizons: the role of ferroptosis in cancer. Nat Rev Clin Oncol. 2021;18:280–96.

    Article  CAS  PubMed  Google Scholar 

  36. Anandhan A, Dodson M, Schmidlin CJ, Liu P, Zhang DD. Breakdown of an Ironclad Defense System: The Critical Role of NRF2 in Mediating Ferroptosis. Cell Chem Biol. 2020;27:436–47.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Yang WS, SriRamaratnam R, Welsch ME, Shimada K, Skouta R, Viswanathan VS, et al. Regulation of ferroptotic cancer cell death by GPX4. Cell 2014;156:317–31.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Tang Z, Jiang W, Mao M, Zhao J, Chen J, Cheng N. Deubiquitinase USP35 modulates ferroptosis in lung cancer via targeting ferroportin. Clin Transl Med. 2021;11:e390.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Hanahan D, Weinberg RA. Hallmarks of cancer: the next generation. Cell 2011;144:646–74.

    Article  CAS  PubMed  Google Scholar 

  40. Galluzzi L, Vitale I, Aaronson SA, Abrams JM, Adam D, Agostinis P, et al. Molecular mechanisms of cell death: recommendations of the Nomenclature Committee on Cell Death 2018. Cell death Differ. 2018;25:486–541.

    Article  PubMed  PubMed Central  Google Scholar 

  41. Diepstraten ST, Anderson MA, Czabotar PE, Lessene G, Strasser A, Kelly GL. The manipulation of apoptosis for cancer therapy using BH3-mimetic drugs. Nature Rev Cancer. 2021.

  42. Zhou Z, Luo A, Shrivastava I, He M, Huang Y, Bahar I, et al. Regulation of XIAP Turnover Reveals a Role for USP11 in Promotion of Tumorigenesis. EBioMedicine 2017;15:48–61.

    Article  PubMed  Google Scholar 

  43. Koschel J, Nishanth G, Just S, Harit K, Kroger A, Deckert M, et al. OTUB1 prevents lethal hepatocyte necroptosis through stabilization of c-IAP1 during murine liver inflammation. Cell Death Differ. 2021;28:2257–75.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Wang W, Lin H, Zheng E, Hou Z, Liu Y, Huang W, et al. Regulation of survivin protein stability by USP35 is evolutionarily conserved. Biochemical Biophys Res Commun. 2021;574:48–55.

    Article  CAS  Google Scholar 

  46. Pal A, Young MA, Donato NJ. Emerging potential of therapeutic targeting of ubiquitin-specific proteases in the treatment of cancer. Cancer Res. 2014;74:4955–66.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

Authors thank Prof. Yang Wang (Dalian Medical University) for generously providing reagents.

Funding

This work was supported by the National Natural Science Foundation of China (HL, 82273054) and the LiaoNing Revitalization Talents Program (HL, XLYC1807079).

Author information

Author notes

  1. These authors contributed equally: Shanshan Wang, Taishu Wang.

Authors and Affiliations

  1. Institute of Cancer Stem Cell, Dalian Medical University, Dalian, China

    Shanshan Wang, Taishu Wang, Xuehong Zhang, Shaoxuan Cheng, Chaoqun Chen, Guoheng Yang, Fuqiang Wang, Ruilin Wang, Dian Yang, Yingqiu Zhang, Shuyan Liu, Quentin Liu & Han Liu

  2. National Institute of Biological Sciences, Beijing, China

    Taishu Wang

  3. Department of Pathology, Dalian Medical University, Dalian, China

    Qingqing Zhang

  4. CAS Key Laboratory of Separation Science for Analytical Chemistry, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, China

    Hongqiang Qin

Authors
  1. Shanshan Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Taishu Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Xuehong Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Shaoxuan Cheng
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Chaoqun Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Guoheng Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Fuqiang Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Ruilin Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Qingqing Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Dian Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Yingqiu Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Shuyan Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. Hongqiang Qin
    View author publications

    You can also search for this author in PubMed Google Scholar

  14. Quentin Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  15. Han Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

HL conceived and designed the project. SW, TW, SC performed experiments and generated figures. XZ and CC carried out bioinformatic analysis. SW, TW, GY, FW, RW and QZ collected the data. SW, TW, DY, YZ, SL, HQ, and QL analyzed and interpreted the data. HL supervised the project and wrote the paper.

Corresponding author

Correspondence to Han Liu.

Ethics declarations

Competing interests

The authors declare no competing interest.

Ethics

All procedures followed were in accordance with the ethical standards approved by the Institutional Animal Care and Use Committee at Dalian Medical University.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Wang, S., Wang, T., Zhang, X. et al. The deubiquitylating enzyme USP35 restricts regulated cell death to promote survival of renal clear cell carcinoma. Cell Death Differ 30, 1757–1770 (2023). https://doi.org/10.1038/s41418-023-01176-3

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41418-023-01176-3

Search

Advanced search

Quick links