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Acta Neuropathologica

, Volume 120, Issue 2, pp 145–154 | Cite as

Proteinase K-resistant α-synuclein is deposited in presynapses in human Lewy body disease and A53T α-synuclein transgenic mice

  • Kunikazu TanjiEmail author
  • Fumiaki Mori
  • Junsei Mimura
  • Ken Itoh
  • Akiyoshi Kakita
  • Hitoshi Takahashi
  • Koichi Wakabayashi
Original Paper

Abstract

Abnormally modified α-synuclein is a pathological hallmark of Parkinson’s disease and the other α-synucleinopathies. Since proteinase K (PK) treatment is known to enhance the immunoreactivity of abnormal α-synuclein, we immunohistochemically examined the brain of transgenic (Tg) mice expressing human mutant A53T α-synuclein using this retrieval method. PK treatment abolished the immunoreactivity of α-synuclein in abnormal inclusions as well as of endogenous α-synuclein in Tg mice, whereas PK-resistant α-synuclein was found in the presynaptic nerve terminals, especially in the hippocampus and temporal cortex. In human Lewy body disease, PK-resistant α-synuclein was deposited in Lewy bodies and Lewy neurites, as well as in the presynapses in distinct brain regions, including the hippocampus, temporal cortex and substantia nigra. Biochemical analysis revealed that PK-resistant α-synuclein was detected in the presynaptic fraction in Tg mice and human Lewy body disease. Although PK-resistant α-synuclein was found in the presynapse in Tg mice even at 1 week of age, it was not phosphorylated until at least 8 months of age. Moreover, PK-resistant α-synuclein in the presynapse was not phosphorylated in human Lewy body disease. These findings suggest that phosphorylation is not necessary to cause the conversion of soluble form to PK-resistant α-synuclein. Considering that native α-synuclein is a soluble protein localized to the presynaptic terminals, our findings suggest that PK-resistant α-synuclein may disturb the neurotransmission in α-synucleinopathies.

Keywords

α-Synuclein Dementia with Lewy bodies Parkinson’s disease Presynapse Proteinase K Transgenic mice 

Notes

Acknowledgments

This work was supported by Grants-in-Aid 20590335 (K.T.), 20591361 (F.M.) and 20300123 (K.W.) for Scientific Research from the Ministry of Education, Culture, Sports, Science, and Technology, Japan, a Grant for Hirosaki University Institutional Research (K.W.), a Hirosaki University Grant for Exploratory Research by Young Scientists (K.T.), and a Grant-in-Aid for Studies on the Development of Diagnostic Technique and Therapies for Lewy Body Disease, the Ministry of Health, Labour and Welfare, Japan (K.W.). The authors wish to express their gratitude to M. Nakata for her technical assistance.

References

  1. 1.
    Abeliovich A, Schmitz Y, Farinas I et al (2000) Mice lacking alpha-synuclein display functional deficits in the nigrostriatal dopamine system. Neuron 25:239–252CrossRefPubMedGoogle Scholar
  2. 2.
    Beach TG, White CL, Hamilton RL et al (2008) Evaluation of alpha-synuclein immunohistochemical methods used by invited experts. Acta Neuropathol 116:277–288CrossRefPubMedGoogle Scholar
  3. 3.
    Braak H, Del Tredici K, Rub U et al (2003) Staging of brain pathology related to sporadic Parkinson’s disease. Neurobiol Aging 24:197–211CrossRefPubMedGoogle Scholar
  4. 4.
    Duffy PE, Tennyson VM (1965) Phase and electron microscopic observations of Lewy bodies and melanin granules in the substantia nigra and locus caeruleus in Parkinson’s disease. J Neuropathol Exp Neurol 24:398–414CrossRefGoogle Scholar
  5. 5.
    Fernagut PO, Hutson CB, Fleming SM et al (2007) Behavioral and histopathological consequences of paraquat intoxication in mice: effects of alpha-synuclein over-expression. Synapse 61:991–1001CrossRefPubMedGoogle Scholar
  6. 6.
    Fujiwara H, Hasegawa M, Dohmae N et al (2002) Alpha-synuclein is phosphorylated in synucleinopathy lesions. Nat Cell Biol 4:160–164CrossRefPubMedGoogle Scholar
  7. 7.
    Galvin JE, Uryu K, Lee VM, Trojanowski JQ (1999) Axon pathology in Parkinson’s disease and Lewy body dementia hippocampus contains alpha-, beta-, and gamma-synuclein. Proc Natl Acad Sci USA 96:13450–13455CrossRefPubMedGoogle Scholar
  8. 8.
    Giasson BI, Duda JE, Murray IV et al (2000) Oxidative damage linked to neurodegeneration by selective alpha-synuclein nitration in synucleinopathy lesions. Science 290:985–989CrossRefPubMedGoogle Scholar
  9. 9.
    Giasson BI, Duda JE, Quinn SM et al (2002) Neuronal alpha-synucleinopathy with severe movement disorder in mice expressing A53T human alpha-synuclein. Neuron 34:521–533CrossRefPubMedGoogle Scholar
  10. 10.
    Hasegawa M, Fujiwara H, Nonaka T et al (2002) Phosphorylated alpha-synuclein is ubiquitinated in alpha-synucleinopathy lesions. J Biol Chem 277:49071–49076CrossRefPubMedGoogle Scholar
  11. 11.
    Iwai A, Masliah E, Yoshimoto M et al (1995) The precursor protein of non-Abeta component of Alzheimer’s disease amyloid is a presynaptic protein of the central nervous system. Neuron 14:467–475CrossRefPubMedGoogle Scholar
  12. 12.
    Iwatsubo T, Yamaguchi H, Fujimuro M et al (1996) Purification and characterization of Lewy bodies from the brains of patients with diffuse Lewy body disease. Am J Pathol 148:1517–1529PubMedGoogle Scholar
  13. 13.
    Jakes R, Crowther RA, Lee VM et al (1999) Epitope mapping of LB509, a monoclonal antibody directed against human alpha-synuclein. Neurosci Lett 269:13–16CrossRefPubMedGoogle Scholar
  14. 14.
    Kahle PJ (2008) Alpha-synucleinopathy models and human neuropathology: similarities and differences. Acta Neuropathol 115:87–95CrossRefPubMedGoogle Scholar
  15. 15.
    Kahle PJ, Neumann M, Ozmen L et al (2000) Subcellular localization of wild-type and Parkinson’s disease-associated mutant alpha-synuclein in human and transgenic mouse brain. J Neurosci 20:6365–6373PubMedGoogle Scholar
  16. 16.
    Kramer ML, Schulz-Schaeffer WJ (2007) Presynaptic alpha-synuclein aggregates, not Lewy bodies, cause neurodegeneration in dementia with Lewy bodies. J Neurosci 27:1405–1410CrossRefPubMedGoogle Scholar
  17. 17.
    Kruger R, Kuhn W, Muller T et al (1998) Ala30Pro mutation in the gene encoding alpha-synuclein in Parkinson’s disease. Nat Genet 18:106–108CrossRefPubMedGoogle Scholar
  18. 18.
    Kuzuhara S, Mori H, Izumiyama N, Yoshimura M, Ihara Y (1988) Lewy bodies are ubiquitinated. A light and electron microscopic immunocytochemical study. Acta Neuropathol 75:345–353CrossRefPubMedGoogle Scholar
  19. 19.
    Lee MK, Stirling W, Xu Y et al (2002) Human alpha-synuclein-harboring familial Parkinson’s disease-linked Ala-53→Thr mutation causes neurodegenerative disease with alpha-synuclein aggregation in transgenic mice. Proc Natl Acad Sci USA 99:8968–8973CrossRefPubMedGoogle Scholar
  20. 20.
    Liu S, Ninan I, Antonova I et al (2004) Alpha-synuclein produces a long-lasting increase in neurotransmitter release. EMBO J 23:4506–4516CrossRefPubMedGoogle Scholar
  21. 21.
    Lu XH, Fleming SM, Meurers B et al (2009) Bacterial artificial chromosome transgenic mice expressing a truncated mutant parkin exhibit age-dependent hypokinetic motor deficits, dopaminergic neuron degeneration, and accumulation of proteinase K-resistant alpha-synuclein. J Neurosci 29:1962–1976CrossRefPubMedGoogle Scholar
  22. 22.
    Maroteaux L, Scheller RH (1991) The rat brain synucleins; family of proteins transiently associated with neuronal membrane. Brain Res Mol Brain Res 11:335–343CrossRefPubMedGoogle Scholar
  23. 23.
    Masliah E, Rockenstein E, Veinbergs I et al (2000) Dopaminergic loss and inclusion body formation in alpha-synuclein mice: implications for neurodegenerative disorders. Science 287:1265–1269CrossRefPubMedGoogle Scholar
  24. 24.
    Matsuoka Y, Vila M, Lincoln S et al (2001) Lack of nigral pathology in transgenic mice expressing human alpha-synuclein driven by the tyrosine hydroxylase promoter. Neurobiol Dis 8:535–539CrossRefPubMedGoogle Scholar
  25. 25.
    Mori F, Piao YS, Hayashi S et al (2003) Alpha-synuclein accumulates in Purkinje cells in Lewy body disease but not in multiple system atrophy. J Neuropathol Exp Neurol 62:812–819PubMedGoogle Scholar
  26. 26.
    Mori F, Tanji K, Yoshimoto M, Takahashi H, Wakabayashi K (2002) Demonstration of alpha-synuclein immunoreactivity in neuronal and glial cytoplasm in normal human brain tissue using proteinase K and formic acid pretreatment. Exp Neurol 176:98–104CrossRefPubMedGoogle Scholar
  27. 27.
    Neumann M, Kahle PJ, Giasson BI et al (2002) Misfolded proteinase K-resistant hyperphosphorylated alpha-synuclein in aged transgenic mice with locomotor deterioration and in human alpha-synucleinopathies. J Clin Invest 110:1429–1439PubMedGoogle Scholar
  28. 28.
    Neumann M, Muller V, Kretzschmar HA, Haass C, Kahle PJ (2004) Regional distribution of proteinase K-resistant alpha-synuclein correlates with Lewy body disease stage. J Neuropathol Exp Neurol 63:1225–1235PubMedGoogle Scholar
  29. 29.
    Piao YS, Mori F, Hayashi S et al (2003) Alpha-synuclein pathology affecting Bergmann glia of the cerebellum in patients with alpha-synucleinopathies. Acta Neuropathol 105:403–409PubMedGoogle Scholar
  30. 30.
    Polymeropoulos MH, Lavedan C, Leroy E et al (1997) Mutation in the alpha-synuclein gene identified in families with Parkinson’s disease. Science 276:2045–2047CrossRefPubMedGoogle Scholar
  31. 31.
    Richfield EK, Thiruchelvam MJ, Cory-Slechta DA et al (2002) Behavioral and neurochemical effects of wild-type and mutated human alpha-synuclein in transgenic mice. Exp Neurol 175:35–48CrossRefPubMedGoogle Scholar
  32. 32.
    Rockenstein E, Mallory M, Hashimoto M et al (2002) Differential neuropathological alterations in transgenic mice expressing alpha-synuclein from the platelet-derived growth factor and Thy-1 promoters. J Neurosci Res 68:568–578CrossRefPubMedGoogle Scholar
  33. 33.
    Takeda A, Hashimoto M, Mallory M et al (2000) C-terminal alpha-synuclein immunoreactivity in structures other than Lewy bodies in neurodegenerative disorders. Acta Neuropathol 99:296–304CrossRefPubMedGoogle Scholar
  34. 34.
    Takeda A, Hashimoto M, Mallory M et al (1998) Abnormal distribution of the non-Abeta component of Alzheimer’s disease amyloid precursor/alpha-synuclein in Lewy body disease as revealed by proteinase K and formic acid pretreatment. Lab Invest 78:1169–1177PubMedGoogle Scholar
  35. 35.
    Tanji K, Tanaka T, Mori F et al (2006) NUB1 suppresses the formation of Lewy body-like inclusions by proteasomal degradation of synphilin-1. Am J Pathol 169:553–565CrossRefPubMedGoogle Scholar
  36. 36.
    van der Putten H, Wiederhold KH, Probst A et al (2000) Neuropathology in mice expressing human alpha-synuclein. J Neurosci 20:6021–6029PubMedGoogle Scholar
  37. 37.
    Wakabayashi K, Yoshimoto M, Tsuji S, Takahashi H (1998) Alpha-synuclein immunoreactivity in glial cytoplasmic inclusions in multiple system atrophy. Neurosci Lett 249:180–182CrossRefPubMedGoogle Scholar
  38. 38.
    Yavich L, Oksman M, Tanila H et al (2005) Locomotor activity and evoked dopamine release are reduced in mice overexpressing A30P-mutated human alpha-synuclein. Neurobiol Dis 20:303–313CrossRefPubMedGoogle Scholar
  39. 39.
    Yavich L, Tanila H, Vepsalainen S, Jakala P (2004) Role of alpha-synuclein in presynaptic dopamine recruitment. J Neurosci 24:11165–11170CrossRefPubMedGoogle Scholar
  40. 40.
    Zarranz JJ, Alegre J, Gomez-Esteban JC et al (2004) The new mutation, E46K, of alpha-synuclein causes Parkinson and Lewy body dementia. Ann Neurol 55:164–173CrossRefPubMedGoogle Scholar
  41. 41.
    Zhang HX, Tanji K, Mori F, Wakabayashi K (2008) Epitope mapping of 2E2-D3, a monoclonal antibody directed against human TDP-43. Neurosci Lett 434:170–174CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Kunikazu Tanji
    • 1
    Email author
  • Fumiaki Mori
    • 1
  • Junsei Mimura
    • 2
  • Ken Itoh
    • 2
  • Akiyoshi Kakita
    • 3
  • Hitoshi Takahashi
    • 4
  • Koichi Wakabayashi
    • 1
  1. 1.Department of Neuropathology, Institute of Brain ScienceHirosaki University Graduate School of MedicineHirosakiJapan
  2. 2.Department of Stress Response ScienceHirosaki University Graduate School of MedicineHirosakiJapan
  3. 3.Department of Pathological Neuroscience, Center for Bioresource-based ResearchesUniversity of NiigataNiigataJapan
  4. 4.Department of Pathology, Brain Research InstituteUniversity of NiigataNiigataJapan

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