Skip to main content
SpringerLink
Log in

Overexpression of p62/SQSTM1 promotes the degradations of abnormally accumulated PrP mutants in cytoplasm and relieves the associated cytotoxicities via autophagy–lysosome-dependent way

  • Original Investigation
  • Published:
Medical Microbiology and Immunology Aims and scope Submit manuscript

Abstract

The protein of p62/sequestosome 1 (SQSTM1), a key cargo adaptor protein involved in autophagy–lysosome degradation, exhibits inclusion bodies structure in cytoplasm and plays a protective role in some models of neurodegenerative diseases. Some PrP mutants, such as PrP-CYTO and PrP-PG14, also form cytosolic inclusion bodies and trigger neuronal apoptosis either in cultured cells or in transgenic mice. Here, we demonstrated that the cellular p62/SQSTM1 incorporated into the inclusion bodies formed by expressing the abnormal PrP mutants, PrP-CYTO and PrP-PG14, in human embryonic kidney 293 cells. Overexpression of p62/SQSTM1 efficiently relieved the cytosolic aggregations and cell apoptosis induced by the abnormal PrPs. Autophagy–lysosome inhibitors instead of proteasome inhibitor sufficiently blocked the p62/SQSTM1-mediated degradations of abnormal PrPs. Overexpression of p62/SQSTM1 did not alter the levels of light chain 3 (LC3) in the cells expressing various PrPs. However, more complexes of p62/SQSTM1 with LC3 were detected in the cells expressing the misfolded PrPs. These data imply that p62/SQSTM1 plays an important role in the homeostasis of abnormal PrPs via autophagy–lysosome-dependent way.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

References

  1. Wang F, Wang X, Yuan CG, Ma J (2010) Generating a prion with bacterially expressed recombinant prion protein. Science 327(5969):1132–1135. doi:10.1126/science.1183748

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  2. Shi Q, Jing YY, Wang SB, Chen C, Sun H, Xu Y, Gao C, Zhang J, Tian C, Guo Y, Ren K, Dong XP (2013) PrP octarepeats region determined the interaction with caveolin-1 and phosphorylation of caveolin-1 and Fyn. Med Microbiol Immunol 202(3):215–227. doi:10.1007/s00430-012-0284-8

    Article  PubMed  CAS  Google Scholar 

  3. Wisniewski T, Sigurdsson EM (2007) Therapeutic approaches for prion and Alzheimer’s diseases. FEBS J 274(15):3784–3798. doi:10.1111/j.1742-4658.2007.05919.x

    Article  PubMed  CAS  Google Scholar 

  4. Lasmezas CI, Deslys JP, Robain O, Jaegly A, Beringue V, Peyrin JM, Fournier JG, Hauw JJ, Rossier J, Dormont D (1997) Transmission of the BSE agent to mice in the absence of detectable abnormal prion protein. Science 275(5298):402–405

    Article  PubMed  CAS  Google Scholar 

  5. Xie WL, Shi Q, Xia SL, Zhang BY, Gong HS, Wang SB, Xu Y, Guo Y, Tian C, Zhang J, Xu BL, Liu Y, Dong XP (2013) Comparison of the pathologic and pathogenic features in six different regions of postmortem brains of three patients with fatal familial insomnia. Int J Mol Med 31(1):81–90. doi:10.3892/ijmm 2012.1194

    PubMed  CAS  Google Scholar 

  6. Bueler H, Raeber A, Sailer A, Fischer M, Aguzzi A, Weissmann C (1994) High prion and PrPSc levels but delayed onset of disease in scrapie-inoculated mice heterozygous for a disrupted PrP gene. Mol Med 1(1):19–30

    PubMed  CAS  Google Scholar 

  7. Sandberg MK, Al-Doujaily H, Sharps B, Clarke AR, Collinge J (2011) Prion propagation and toxicity in vivo occur in two distinct mechanistic phases. Nature 470(7335):540–542. doi:10.1038/nature09768

    Article  PubMed  CAS  Google Scholar 

  8. Wang SB, Shi Q, Xu Y, Xie WL, Zhang J, Tian C, Guo Y, Wang K, Zhang BY, Chen C, Gao C, Dong XP (2012) Protein disulfide isomerase regulates endoplasmic reticulum stress and the apoptotic process during prion infection and PrP mutant-induced cytotoxicity. PLoS ONE 7(6):e38221. doi:10.1371/journal.pone.0038221

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  9. Ma J, Wollmann R, Lindquist S (2002) Neurotoxicity and neurodegeneration when PrP accumulates in the cytosol. Science 298(5599):1781–1785. doi:10.1126/science.1073725

    Article  PubMed  CAS  Google Scholar 

  10. Zhang J, Wang K, Guo Y, Shi Q, Tian C, Chen C, Gao C, Zhang BY, Dong XP (2012) Heat shock protein 70 selectively mediates the degradation of cytosolic PrPs and restores the cytosolic PrP-induced cytotoxicity via a molecular interaction. Virol J 9:303. doi:10.1186/1743-422X-9-303

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  11. Peethumnongsin E, Yang L, Kallhoff-Munoz V, Hu L, Takashima A, Pautler RG, Zheng H (2010) Convergence of presenilin- and tau-mediated pathways on axonal trafficking and neuronal function. J Neurosci 30(40):13409–13418. doi:10.1523/JNEUROSCI.1964-10.2010

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  12. Watanabe Y, Tatebe H, Taguchi K, Endo Y, Tokuda T, Mizuno T, Nakagawa M, Tanaka M (2012) p62/SQSTM1-dependent autophagy of Lewy body-like alpha-synuclein inclusions. PLoS ONE 7(12):e52868. doi:10.1371/journal.pone.0052868

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  13. Bjorkoy G, Lamark T, Brech A, Outzen H, Perander M, Overvatn A, Stenmark H, Johansen T (2005) p62/SQSTM1 forms protein aggregates degraded by autophagy and has a protective effect on huntingtin-induced cell death. J Cell Biol 171(4):603–614. doi:10.1083/jcb.200507002

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  14. Komatsu M, Waguri S, Koike M, Sou YS, Ueno T, Hara T, Mizushima N, Iwata J, Ezaki J, Murata S, Hamazaki J, Nishito Y, Iemura S, Natsume T, Yanagawa T, Uwayama J, Warabi E, Yoshida H, Ishii T, Kobayashi A, Yamamoto M, Yue Z, Uchiyama Y, Kominami E, Tanaka K (2007) Homeostatic levels of p62 control cytoplasmic inclusion body formation in autophagy-deficient mice. Cell 131(6):1149–1163. doi:10.1016/j.cell.2007.10.035

    Article  PubMed  CAS  Google Scholar 

  15. Moscat J, Diaz-Meco MT (2009) p62 at the crossroads of autophagy, apoptosis, and cancer. Cell 137(6):1001–1004. doi:10.1016/j.cell.2009.05.023

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  16. Kuusisto E, Salminen A, Alafuzoff I (2002) Early accumulation of p62 in neurofibrillary tangles in Alzheimer’s disease: possible role in tangle formation. Neuropathol Appl Neurobiol 28(3):228–237

    Article  PubMed  CAS  Google Scholar 

  17. Zatloukal K, Stumptner C, Fuchsbichler A, Heid H, Schnoelzer M, Kenner L, Kleinert R, Prinz M, Aguzzi A, Denk H (2002) p62 Is a common component of cytoplasmic inclusions in protein aggregation diseases. Am J Pathol 160(1):255–263. doi:10.1016/S0002-9440(10)64369-6

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  18. Stumptner C, Fuchsbichler A, Zatloukal K, Denk H (2007) In vitro production of Mallory bodies and intracellular hyaline bodies: the central role of sequestosome 1/p62. Hepatology 46(3):851–860. doi:10.1002/hep.21744

    Article  PubMed  CAS  Google Scholar 

  19. Pankiv S, Clausen TH, Lamark T, Brech A, Bruun JA, Outzen H, Overvatn A, Bjorkoy G, Johansen T (2007) p62/SQSTM1 binds directly to Atg8/LC3 to facilitate degradation of ubiquitinated protein aggregates by autophagy. J Biol Chem 282(33):24131–24145. doi:10.1074/jbc.M702824200

    Article  PubMed  CAS  Google Scholar 

  20. Zhao Y, Yang J, Liao W, Liu X, Zhang H, Wang S, Wang D, Feng J, Yu L, Zhu WG (2010) Cytosolic FoxO1 is essential for the induction of autophagy and tumour suppressor activity. Nat Cell Biol 12(7):665–675. doi:10.1038/ncb2069

    Article  PubMed  CAS  Google Scholar 

  21. Shibata M, Lu T, Furuya T, Degterev A, Mizushima N, Yoshimori T, MacDonald M, Yankner B, Yuan J (2006) Regulation of intracellular accumulation of mutant Huntingtin by Beclin 1. J Biol Chem 281(20):14474–14485. doi:10.1074/jbc.M600364200

    Article  PubMed  CAS  Google Scholar 

  22. Ravikumar B, Vacher C, Berger Z, Davies JE, Luo S, Oroz LG, Scaravilli F, Easton DF, Duden R, O’Kane CJ, Rubinsztein DC (2004) Inhibition of mTOR induces autophagy and reduces toxicity of polyglutamine expansions in fly and mouse models of Huntington disease. Nat Genet 36(6):585–595. doi:10.1038/ng1362

    Article  PubMed  CAS  Google Scholar 

  23. Aguib Y, Heiseke A, Gilch S, Riemer C, Baier M, Schatzl HM, Ertmer A (2009) Autophagy induction by trehalose counteracts cellular prion infection. Autophagy 5(3):361–369

    Article  PubMed  CAS  Google Scholar 

  24. Klionsky DJ, Abdalla FC, Abeliovich H et al (2012) Guidelines for the use and interpretation of assays for monitoring autophagy. Autophagy 8(4):445–544

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  25. Chen R, Jin R, Wu L, Ye X, Yang Y, Luo K, Wang W, Wu D, Huang L, Huang T, Xiao G (2011) Reticulon 3 attenuates the clearance of cytosolic prion aggregates via inhibiting autophagy. Autophagy 7(2):205–216

    Article  PubMed  CAS  Google Scholar 

  26. Rubinsztein DC (2006) The roles of intracellular protein-degradation pathways in neurodegeneration. Nature 443(7113):780–786. doi:10.1038/nature05291

    Article  PubMed  CAS  Google Scholar 

  27. Xu Y, Tian C, Wang SB, Xie WL, Guo Y, Zhang J, Shi Q, Chen C, Dong XP (2012) Activation of the macroautophagic system in scrapie-infected experimental animals and human genetic prion diseases. Autophagy 8(11):1604–1620. doi:10.4161/auto.21482

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  28. Schaeffer V, Lavenir I, Ozcelik S, Tolnay M, Winkler DT, Goedert M (2012) Stimulation of autophagy reduces neurodegeneration in a mouse model of human tauopathy. Brain J Neurol 135(Pt 7):2169–2177. doi:10.1093/brain/aws143

    Article  Google Scholar 

  29. Pickford F, Masliah E, Britschgi M, Lucin K, Narasimhan R, Jaeger PA, Small S, Spencer B, Rockenstein E, Levine B, Wyss-Coray T (2008) The autophagy-related protein beclin 1 shows reduced expression in early Alzheimer disease and regulates amyloid beta accumulation in mice. J Clin Invest 118(6):2190–2199. doi:10.1172/JCI33585

    PubMed Central  PubMed  CAS  Google Scholar 

  30. Webb JL, Ravikumar B, Rubinsztein DC (2004) Microtubule disruption inhibits autophagosome-lysosome fusion: implications for studying the roles of aggresomes in polyglutamine diseases. Int J Biochem Cell Biol 36(12):2541–2550. doi:10.1016/j.biocel.2004.02.003

    Article  PubMed  Google Scholar 

  31. Lee JH, Yu WH, Kumar A, Lee S, Mohan PS, Peterhoff CM, Wolfe DM, Martinez-Vicente M, Massey AC, Sovak G, Uchiyama Y, Westaway D, Cuervo AM, Nixon RA (2010) Lysosomal proteolysis and autophagy require presenilin 1 and are disrupted by Alzheimer-related PS1 mutations. Cell 141(7):1146–1158. doi:10.1016/j.cell.2010.05.008

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  32. Platt FM, Boland B, van der Spoel AC (2012) The cell biology of disease: lysosomal storage disorders: the cellular impact of lysosomal dysfunction. J Cell Biol 199(5):723–734. doi:10.1083/jcb.201208152

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  33. Martinez-Vicente M, Talloczy Z, Wong E, Tang G, Koga H, Kaushik S, de Vries R, Arias E, Harris S, Sulzer D, Cuervo AM (2010) Cargo recognition failure is responsible for inefficient autophagy in Huntington’s disease. Nat Neurosci 13(5):567–576. doi:10.1038/nn.2528

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  34. Boellaard JW, Kao M, Schlote W, Diringer H (1991) Neuronal autophagy in experimental scrapie. Acta Neuropathol 82(3):225–228

    Article  PubMed  CAS  Google Scholar 

  35. Sikorska B, Liberski PP, Giraud P, Kopp N, Brown P (2004) Autophagy is a part of ultrastructural synaptic pathology in Creutzfeldt–Jakob disease: a brain biopsy study. Int J Biochem Cell Biol 36(12):2563–2573. doi:10.1016/j.biocel.2004.04.014

    Article  PubMed  CAS  Google Scholar 

  36. Larsen KB, Lamark T, Overvatn A, Harneshaug I, Johansen T, Bjorkoy G (2010) A reporter cell system to monitor autophagy based on p62/SQSTM1. Autophagy 6(6):784–793

    Article  PubMed  CAS  Google Scholar 

  37. Heo SR, Han AM, Kwon YK, Joung I (2009) p62 protects SH-SY5Y neuroblastoma cells against H2O2-induced injury through the PDK1/Akt pathway. Neurosci Lett 450(1):45–50. doi:10.1016/j.neulet.2008.11.011

    Article  PubMed  CAS  Google Scholar 

  38. Zou X, Feng Z, Li Y, Wang Y, Wertz K, Weber P, Fu Y, Liu J (2012) Stimulation of GSH synthesis to prevent oxidative stress-induced apoptosis by hydroxytyrosol in human retinal pigment epithelial cells: activation of Nrf2 and JNK-p62/SQSTM1 pathways. J Nutr Biochem 23(8):994–1006. doi:10.1016/j.jnutbio.2011.05.006

    Article  PubMed  CAS  Google Scholar 

  39. Zuber C, Mitteregger G, Schuhmann N, Rey C, Knackmuss S, Rupprecht W, Reusch U, Pace C, Little M, Kretzschmar HA, Hallek M, Buning H, Weiss S (2008) Delivery of single-chain antibodies (scFvs) directed against the 37/67 kDa laminin receptor into mice via recombinant adeno-associated viral vectors for prion disease gene therapy. J Gen Virol 89(Pt 8):2055–2061. doi:10.1099/vir.0.83670-0

    Article  PubMed  CAS  Google Scholar 

  40. Pfeifer A, Eigenbrod S, Al-Khadra S, Hofmann A, Mitteregger G, Moser M, Bertsch U, Kretzschmar H (2006) Lentivector-mediated RNAi efficiently suppresses prion protein and prolongs survival of scrapie-infected mice. J Clin Invest 116(12):3204–3210. doi:10.1172/JCI29236

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  41. Friant S, Meier KD, Riezman H (2003) Increased ubiquitin-dependent degradation can replace the essential requirement for heat shock protein induction. EMBO J 22(15):3783–3791. doi:10.1093/emboj/cdg375

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  42. Lanneau D, Wettstein G, Bonniaud P, Garrido C (2010) Heat shock proteins: cell protection through protein triage. Sci World J 10:1543–1552. doi:10.1100/tsw.2010.152

    Article  CAS  Google Scholar 

  43. Marzo L, Marijanovic Z, Browman D, Chamoun Z, Caputo A, Zurzolo C (2013) 4-Hydroxytamoxifen leads to PrPSc clearance by conveying both PrPC and PrPSc to lysosomes independently of autophagy. J Cell Sci 126(Pt 6):1345–1354. doi:10.1242/jcs.114801

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

This work was supported by Chinese National Natural Science Foundation Grant (81100980, 81101302 and 31100117), China Mega-Project for Infectious Disease (2011ZX10004-101, 2012ZX10004215) and SKLID Development Grant (2012SKLID102, 2011SKLID302, 2011SKLID204, 2011SKLID211).

Author information

Authors and Affiliations

  1. State Key Laboratory for Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases (Zhejiang University), National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Chang-Bai Rd 155, Beijing, 102206, People’s Republic of China

    Yin Xu, Jin Zhang, Chan Tian, Ke Ren, Yu-E Yan, Ke Wang, Hui Wang, Cao Chen, Jing Wang, Qi Shi & Xiao-Ping Dong

  2. Chinese Academy of Sciences Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing, 100101, People’s Republic of China

    Xiao-Ping Dong

  3. Shandong International Travel Health Care Center, Shandong Entry-Exit Inspection and Quarantine Bureau, Qingdao, 266001, People’s Republic of China

    Jin Zhang

Authors
  1. Yin Xu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jin Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Chan Tian
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ke Ren
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Yu-E Yan
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Ke Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Hui Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Cao Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Jing Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Qi Shi
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Xiao-Ping Dong
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Xiao-Ping Dong.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Xu, Y., Zhang, J., Tian, C. et al. Overexpression of p62/SQSTM1 promotes the degradations of abnormally accumulated PrP mutants in cytoplasm and relieves the associated cytotoxicities via autophagy–lysosome-dependent way. Med Microbiol Immunol 203, 73–84 (2014). https://doi.org/10.1007/s00430-013-0316-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00430-013-0316-z

Keywords

Search

Navigation