Advertisement

Molecular Neurobiology

, Volume 52, Issue 3, pp 1180–1189 | Cite as

Bcl-2 Decreases the Affinity of SQSTM1/p62 to Poly-Ubiquitin Chains and Suppresses the Aggregation of Misfolded Protein in Neurodegenerative Disease

  • Liang Zhou
  • Hongfeng Wang
  • Haigang Ren
  • Qingsong Hu
  • Zheng YingEmail author
  • Guanghui WangEmail author
Article

Abstract

Poly-ubiquitinated protein aggregate formation is the most striking hallmark of various neurodegenerative diseases such as Alzheimer’s disease, Huntington’s disease, amyotrophic lateral sclerosis, and prion disease. Mutations of many ubiquitin-associated proteins involved in the regulation of protein aggregation, such as SQSTM1/p62 (p62), parkin, and VCP, are closely linked to neurodegeneration. B-cell lymphoma 2 (Bcl-2) is a key regulator in autophagy, apoptosis, and mitochondria quality control in many cell types including neurons, and it plays important roles in the pathogenesis of neurodegenerative diseases mentioned above. Our previous work showed that Bcl-2 can directly bind to p62, and here we report that Bcl-2 directly interacts with the N-terminus of p62, but not the C-terminus (UBA domain). Interestingly and importantly, Bcl-2 affects the affinity of p62 to poly-ubiquitin chains and suppresses the aggregation of poly-ubiquitinated proteins such as mutant huntingtin associated with Huntington’s disease. Our study reveals a role of Bcl-2 that involves in the regulation of misfolded proteins.

Keywords

Neurodegenerative disease Huntingtin Aggregate Bcl-2 SQSTM1/p62 

Notes

Acknowledgments

This work was supported in part by the National High-tech Research and Development program of China 973-projects (2011CB504102), the National Natural Sciences Foundation of China (Nos. 31200803 and 31371072), Natural Science Foundation of Jiangsu Province (BK2012181), a project funded by Jiangsu Key Laboratory of Translational Research and Therapy for Neuro-Psycho-Diseases (BM2013003), and a project funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions.

Supplementary material

12035_2014_8908_MOESM1_ESM.docx (811 kb)
Fig. S1 H1299 cells were transfected with EGFP-Bcl-2 for 48 h, and then the cells were fixed and subjected to immunofluorescent assay using anti-p62 (Enzo life) antibody to detect endogenous p62 (red) and DAPI to stain the nuclei (blue). The scale bar indicates 10 μm. (DOCX 810 kb)
12035_2014_8908_MOESM2_ESM.docx (1.4 mb)
Fig. S2 HEK293 cells were transfected with EGFP-Htt-60Q and HA-ubiquitin (Ub) for 12 h, then the cells were subjected to immunofluorescent assay using anti-HA antibody (blue). The arrows indicate that Htt-60Q and ubiquitin could co-aggregate together. The scale bar indicates 10 μm. (DOCX 1387 kb)

References

  1. 1.
    Rubinsztein DC (2006) The roles of intracellular protein-degradation pathways in neurodegeneration. Nature 443(7113):780–786. doi: 10.1038/nature05291 CrossRefPubMedGoogle Scholar
  2. 2.
    Ross CA, Poirier MA (2005) Opinion: what is the role of protein aggregation in neurodegeneration? Nat Rev Mol Cell Biol 6(11):891–898. doi: 10.1038/nrm1742 CrossRefPubMedGoogle Scholar
  3. 3.
    Ross CA, Poirier MA (2004) Protein aggregation and neurodegenerative disease. Nat Med 10:S10–S17. doi: 10.1038/nm1066 CrossRefPubMedGoogle Scholar
  4. 4.
    Kim SH, Shi Y, Hanson KA, Williams LM, Sakasai R, Bowler MJ, Tibbetts RS (2009) Potentiation of amyotrophic lateral sclerosis (ALS)-associated TDP-43 aggregation by the proteasome-targeting factor, ubiquilin 1. J Biol Chem 284(12):8083–8092. doi: 10.1074/jbc.M808064200 CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Olzmann JA, Li L, Chudaev MV, Chen J, Perez FA, Palmiter RD, Chin LS (2007) Parkin-mediated K63-linked polyubiquitination targets misfolded DJ-1 to aggresomes via binding to HDAC6. J Cell Biol 178(6):1025–1038CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Sha Y, Pandit L, Zeng S, Eissa NT (2009) A critical role for CHIP in the aggresome pathway. Mol Cell Biol 29(1):116–128CrossRefPubMedGoogle Scholar
  7. 7.
    Bruijn LI, Houseweart MK, Kato S, Anderson KL, Anderson SD, Ohama E, Reaume AG, Scott RW, Cleveland DW (1998) Aggregation and motor neuron toxicity of an ALS-Linked SOD1 mutant independent from wild-type SOD1. Science 281(5384):1851–1854. doi: 10.1126/science.281.5384.1851 CrossRefPubMedGoogle Scholar
  8. 8.
    Kalchman MA, Graham RK, Xia G, Koide HB, Hodgson JG, Graham KC, Goldberg YP, Gietz RD, Pickart CM, Hayden MR (1996) Huntingtin is ubiquitinated and interacts with a specific ubiquitin-conjugating enzyme. J Biol Chem 271(32):19385–19394CrossRefPubMedGoogle Scholar
  9. 9.
    Paulson HL, Perez MK, Trottier Y, Trojanowski JQ, Subramony SH, Das SS, Vig P, Mandel JL, Fischbeck KH, Pittman RN (1997) Intranuclear inclusions of expanded polyglutamine protein in spinocerebellar ataxia type 3. Neuron 19(2):333–344CrossRefPubMedGoogle Scholar
  10. 10.
    Ying Z, Wang H, Wang G (2013) The ubiquitin proteasome system as a potential target for the treatment of neurodegenerative diseases. Curr Pharm Des 19(18):3305–3314CrossRefPubMedGoogle Scholar
  11. 11.
    Kuusisto E, Parkkinen L, Alafuzoff I (2003) Morphogenesis of Lewy bodies: dissimilar incorporation of alpha-synuclein, ubiquitin, and p62. J Neuropathol Exp Neurol 62(12):1241–1253CrossRefPubMedGoogle Scholar
  12. 12.
    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 CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Gal J, Strom AL, Kilty R, Zhang F, Zhu H (2007) p62 accumulates and enhances aggregate formation in model systems of familial amyotrophic lateral sclerosis. J Biol Chem 282(15):11068–11077. doi: 10.1074/jbc.M608787200 CrossRefPubMedGoogle Scholar
  14. 14.
    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):228–237CrossRefPubMedGoogle Scholar
  15. 15.
    Nagaoka U, Kim K, Jana NR, Doi H, Maruyama M, Mitsui K, Oyama F, Nukina N (2004) Increased expression of p62 in expanded polyglutamine-expressing cells and its association with polyglutamine inclusions. J Neurochem 91(1):57–68. doi: 10.1111/j.1471-4159.2004.02692.x CrossRefPubMedGoogle Scholar
  16. 16.
    Seidel K, den Dunnen WF, Schultz C, Paulson H, Frank S, de Vos RA, Brunt ER, Deller T, Kampinga HH, Rub U (2010) Axonal inclusions in spinocerebellar ataxia type 3. Acta Neuropathol 120(4):449–460. doi: 10.1007/s00401-010-0717-7 CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Kirkin V, McEwan DG, Novak I, Dikic I (2009) A role for ubiquitin in selective autophagy. Mol Cell 34(3):259–269. doi: 10.1016/j.molcel.2009.04.026 CrossRefPubMedGoogle Scholar
  18. 18.
    Moscat J, Diazmeco M, Wooten M (2007) Signal integration and diversification through the p62 scaffold protein. Trends Biochem Sci 32(2):95–100. doi: 10.1016/j.tibs.2006.12.002 CrossRefPubMedGoogle Scholar
  19. 19.
    Saio T, Yokochi M, Inagaki F (2009) The NMR structure of the p62 PB1 domain, a key protein in autophagy and NF-κB signaling pathway. J Biomol NMR 45(3):335–341. doi: 10.1007/s10858-009-9370-7 CrossRefPubMedGoogle Scholar
  20. 20.
    Nakamura K, Kimple AJ, Siderovski DP, Johnson GL (2010) PB1 domain interaction of p62/sequestosome 1 and MEKK3 regulates NF-κB activation. J Biol Chem 285(3):2077–2089. doi: 10.1074/jbc.M109.065102 CrossRefPubMedGoogle Scholar
  21. 21.
    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 CrossRefPubMedGoogle Scholar
  22. 22.
    Tung YT, Hsu WM, Lee H, Huang WP, Liao YF (2010) The evolutionarily conserved interaction between LC3 and p62 selectively mediates autophagy-dependent degradation of mutant huntingtin. Cell Mol Neurobiol 30(5):795–806. doi: 10.1007/s10571-010-9507-y CrossRefPubMedGoogle Scholar
  23. 23.
    Seibenhener ML, Babu JR, Geetha T, Wong HC, Krishna NR, Wooten MW (2004) Sequestosome 1/p62 is a polyubiquitin chain binding protein involved in ubiquitin proteasome degradation. Mol Cell Biol 24(18):8055–8068. doi: 10.1128/mcb.24.18.8055-8068.2004 CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Teyssou E, Takeda T, Lebon V, Boillée S, Doukouré B, Bataillon G, Sazdovitch V, Cazeneuve C, Meininger V, LeGuern E, Salachas F, Seilhean D, Millecamps S (2013) Mutations in SQSTM1 encoding p62 in amyotrophic lateral sclerosis: genetics and neuropathology. Acta Neuropathol 125(4):511–522. doi: 10.1007/s00401-013-1090-0 CrossRefPubMedGoogle Scholar
  25. 25.
    Hiruma Y, Kurihara N, Subler MA, Zhou H, Boykin CS, Zhang H, Ishizuka S, Dempster DW, Roodman GD, Windle JJ (2008) A SQSTM1/p62 mutation linked to Paget’s disease increases the osteoclastogenic potential of the bone microenvironment. Hum Mol Genet 17(23):3708–3719. doi: 10.1093/hmg/ddn266 CrossRefPubMedPubMedCentralGoogle Scholar
  26. 26.
    Cavey JR, Ralston SH, Hocking LJ, Sheppard PW, Ciani B, Searle MS, Layfield R (2005) Loss of ubiquitin-binding associated with Paget’s disease of bone p62 (SQSTM1) mutations. J Bone Min Res Off J Am Soc Bone Min Res 20(4):619–624. doi: 10.1359/jbmr.041205 CrossRefGoogle Scholar
  27. 27.
    Green JC, Reed DR (1998) Mitochondria and apoptosis. Science 281(5381):1309–1312. doi: 10.1126/science.281.5381.1309 CrossRefPubMedGoogle Scholar
  28. 28.
    Yang J (1997) Prevention of apoptosis by Bcl-2: release of cytochrome c from mitochondria blocked. Science 275(5303):1129–1132CrossRefPubMedGoogle Scholar
  29. 29.
    Wei Y, Pattingre S, Sinha S, Bassik M, Levine B (2008) JNK1-mediated phosphorylation of Bcl-2 regulates starvation-induced autophagy. Mol Cell 30(6):678–688. doi: 10.1016/j.molcel.2008.06.001 CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Pattingre S, Tassa A, Qu X, Garuti R, Liang XH, Mizushima N, Packer M, Schneider MD, Levine B (2005) Bcl-2 antiapoptotic proteins inhibit Beclin 1-dependent autophagy. Cell 122(6):927–939. doi: 10.1016/j.cell.2005.07.002 CrossRefPubMedGoogle Scholar
  31. 31.
    Sassone J, Maraschi A, Sassone F, Silani V, Ciammola A (2013) Defining the role of the Bcl-2 family proteins in Huntington’s disease. Cell Death Dis 4:e772. doi: 10.1038/cddis.2013.300 CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Jana NR, Zemskov EA, Wang G, Nukina N (2001) Altered proteasomal function due to the expression of polyglutamine-expanded truncated N-terminal huntingtin induces apoptosis by caspase activation through mitochondrial cytochrome c release. Hum Mol Genet 10(10):1049–1059CrossRefPubMedGoogle Scholar
  33. 33.
    Wong E, Cuervo AM (2010) Autophagy gone awry in neurodegenerative diseases. Nat Neurosci 13(7):805–811. doi: 10.1038/nn.2575 CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    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 CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Shvets E, Fass E, Scherz-Shouval R, Elazar Z (2008) The N-terminus and Phe52 residue of LC3 recruit p62/SQSTM1 into autophagosomes. J Cell Sci 121(16):2685–2695. doi: 10.1242/jcs.026005 CrossRefPubMedGoogle Scholar
  36. 36.
    Zhou L, Wang H-f, Ren H-g, Chen D, Gao F, Hu Q-s, Fu C, Xu R-j, Ying Z, Wang G-h (2013) Bcl-2-dependent upregulation of autophagy by sequestosome 1/p62 in vitro. Acta pharmacologica SinicaGoogle Scholar
  37. 37.
    Chen D, Gao F, Li B, Wang H, Xu Y, Zhu C, Wang G (2010) Parkin mono-ubiquitinates Bcl-2 and regulates autophagy. J Biol Chem 285(49):38214–38223. doi: 10.1074/jbc.M110.101469 CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Korolchuk VI, Mansilla A, Menzies FM, Rubinsztein DC (2009) Autophagy inhibition compromises degradation of ubiquitin-proteasome pathway substrates. Mol Cell 33(4):517–527. doi: 10.1016/j.molcel.2009.01.021 CrossRefPubMedPubMedCentralGoogle Scholar
  39. 39.
    Itakura E, Mizushima N (2011) p62 targeting to the autophagosome formation site requires self-oligomerization but not LC3 binding. J Cell Biol 192(1):17–27. doi: 10.1083/jcb.201009067 CrossRefPubMedPubMedCentralGoogle Scholar
  40. 40.
    Seibenhener ML, Babu JR, Geetha T, Wong HC, Krishna NR, Wooten MW (2004) Sequestosome 1/p62 is a polyubiquitin chain binding protein involved in ubiquitin proteasome degradation. Mol Cell Biol 24(18):8055–8068. doi: 10.1128/MCB.24.18.8055-8068.2004 CrossRefPubMedPubMedCentralGoogle Scholar
  41. 41.
    Lamark T, Perander M, Outzen H, Kristiansen K, Overvatn A, Michaelsen E, Bjorkoy G, Johansen T (2003) Interaction codes within the family of mammalian Phox and Bem1p domain-containing proteins. J Biol Chem 278(36):34568–34581. doi: 10.1074/jbc.M303221200 CrossRefPubMedGoogle Scholar
  42. 42.
    Matsumoto G, Wada K, Okuno M, Kurosawa M, Nukina N (2011) Serine 403 phosphorylation of p62/SQSTM1 regulates selective autophagic clearance of ubiquitinated proteins. Mol Cell 44(2):279–289. doi: 10.1016/j.molcel.2011.07.039 CrossRefPubMedGoogle Scholar
  43. 43.
    Isogai S, Morimoto D, Arita K, Unzai S, Tenno T, Hasegawa J, Sou YS, Komatsu M, Tanaka K, Shirakawa M, Tochio H (2011) Crystal structure of the ubiquitin-associated (UBA) domain of p62 and its interaction with ubiquitin. J Biol Chem 286(36):31864–31874. doi: 10.1074/jbc.M111.259630 CrossRefPubMedPubMedCentralGoogle Scholar
  44. 44.
    Tien CL, Wen FC, Hsieh M (2008) The polyglutamine-expanded protein ataxin-3 decreases bcl-2 mRNA stability. Biochem Biophys Res Commun 365(2):232–238. doi: 10.1016/j.bbrc.2007.10.162 CrossRefPubMedGoogle Scholar
  45. 45.
    Tsai HF, Tsai HJ, Hsieh M (2004) Full-length expanded ataxin-3 enhances mitochondrial-mediated cell death and decreases Bcl-2 expression in human neuroblastoma cells. Biochem Biophys Res Commun 324(4):1274–1282. doi: 10.1016/j.bbrc.2004.09.192 CrossRefPubMedGoogle Scholar
  46. 46.
    Williams AJ, Paulson HL (2008) Polyglutamine neurodegeneration: protein misfolding revisited. Trends Neurosci 31(10):521–528. doi: 10.1016/j.tins.2008.07.004 CrossRefPubMedPubMedCentralGoogle Scholar
  47. 47.
    Ilieva H, Polymenidou M, Cleveland DW (2009) Non-cell autonomous toxicity in neurodegenerative disorders: ALS and beyond. J Cell Biol 187(6):761–772. doi: 10.1083/jcb.200908164 CrossRefPubMedPubMedCentralGoogle Scholar
  48. 48.
    Taylor JP, Hardy J, Fischbeck KH (2002) Toxic proteins in neurodegenerative disease. Science 296(5575):1991–1995. doi: 10.1126/science.1067122 CrossRefPubMedGoogle Scholar
  49. 49.
    Temussi PA, Masino L, Pastore A (2003) From Alzheimer to Huntington: why is a structural understanding so difficult? EMBO J 22(3):355–361. doi: 10.1093/emboj/cdg044 CrossRefPubMedPubMedCentralGoogle Scholar
  50. 50.
    Brady OA, Meng P, Zheng Y, Mao Y, Hu F (2011) Regulation of TDP-43 aggregation by phosphorylation and p62/SQSTM1. J Neurochem 116(2):248–259. doi: 10.1111/j.1471-4159.2010.07098.x CrossRefPubMedGoogle Scholar
  51. 51.
    Jana NR, Tanaka M, Wang G-h, Nukina N (2000) Polyglutamine length-dependent interaction of Hsp40 and Hsp70 family chaperones with truncated N-terminal huntingtin: their role in suppression of aggregation and cellular toxicity. Hum Mol Genet 9(13):2009–2018. doi: 10.1093/hmg/9.13.2009 CrossRefPubMedGoogle Scholar
  52. 52.
    Ying Z, Wang H, Fan H, Zhu X, Zhou J, Fei E, Wang G (2009) Gp78, an ER associated E3, promotes SOD1 and ataxin-3 degradation. Hum Mol Genet 18(22):4268–4281. doi: 10.1093/hmg/ddp380 CrossRefPubMedGoogle Scholar
  53. 53.
    Wang H, Ying Z, Wang G (2012) Ataxin-3 regulates aggresome formation of copper-zinc superoxide dismutase (SOD1) by editing K63-linked polyubiquitin chains. J Biol Chem 287(34):28576–28585. doi: 10.1074/jbc.M111.299990 CrossRefPubMedPubMedCentralGoogle Scholar
  54. 54.
    Ying Z, Wang H, Fan H, Wang G (2011) The endoplasmic reticulum (ER)-associated degradation system regulates aggregation and degradation of mutant neuroserpin. J Biol Chem 286(23):20835–20844. doi: 10.1074/jbc.M110.200808 CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  1. 1.Laboratory of Molecular Neuropathology, Jiangsu Key Laboratory of Translational Research and Therapy for Neuro-Psycho-Diseases and College of Pharmaceutical SciencesSoochow UniversitySuzhouChina
  2. 2.Key Laboratory of Brain Function and Disease, School of Life SciencesUniversity of Science and Technology of China, Chinese Academy of SciencesHefeiChina

Personalised recommendations