Aggregated forms of α-synuclein play a crucial role in the pathogenesis of synucleinopathies such as Parkinson's disease (PD). However, the molecular mechanisms underlying the pathogenic effects of α-synuclein are not completely understood. Here we show that asparagine endopeptidase (AEP) cleaves human α-synuclein, triggers its aggregation and escalates its neurotoxicity, thus leading to dopaminergic neuronal loss and motor impairments in a mouse model. AEP is activated and cleaves human α-synuclein at N103 in an age-dependent manner. AEP is highly activated in human brains with PD, and it fragments α-synuclein, which is found aggregated in Lewy bodies. Overexpression of the AEP-cleaved α-synuclein1–103 fragment in the substantia nigra induces both dopaminergic neuronal loss and movement defects in mice. In contrast, inhibition of AEP-mediated cleavage of α-synuclein (wild type and A53T mutant) diminishes α-synuclein's pathologic effects. Together, these findings support AEP's role as a key mediator of α-synuclein-related etiopathological effects in PD.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.


  1. 1.

    , , , & The role of alpha-synuclein in Parkinson's disease: insights from animal models. Nat. Rev. Neurosci. 4, 727–738 (2003).

  2. 2.

    et al. Mutation in the alpha-synuclein gene identified in families with Parkinson's disease. Science 276, 2045–2047 (1997).

  3. 3.

    et al. Ala30Pro mutation in the gene encoding α-synuclein in Parkinson's disease. Nat. Genet. 18, 106–108 (1998).

  4. 4.

    , & Intron-exon structure of ubiquitin c-terminal hydrolase-L1. DNA Res. 5, 397–400 (1998).

  5. 5.

    et al. Mutations in the parkin gene cause autosomal recessive juvenile parkinsonism. Nature 392, 605–608 (1998).

  6. 6.

    et al. α-Synuclein in Lewy bodies. Nature 388, 839–840 (1997).

  7. 7.

    & Parkinson's disease and alpha synuclein: is Parkinson's disease a prion-like disorder? Mov. Disord. 28, 31–40 (2013).

  8. 8.

    et al. Dopaminergic loss and inclusion body formation in alpha-synuclein mice: implications for neurodegenerative disorders. Science 287, 1265–1269 (2000).

  9. 9.

    , , & Physiology and pathophysiology of alpha-synuclein: cell culture and transgenic animal models based on a Parkinson's disease-associated protein. Ann. NY Acad. Sci. 920, 33–41 (2000).

  10. 10.

    & Drosophila model of Parkinson's disease. Nature 404, 394–398 (2000).

  11. 11.

    et al. Properties of native brain α-synuclein. Nature 498, E4–E6 (2013).

  12. 12.

    , , & The many faces of α-synuclein: from structure and toxicity to therapeutic target. Nat. Rev. Neurosci. 14, 38–48 (2013).

  13. 13.

    et al. Aggregation promoting C-terminal truncation of alpha-synuclein is a normal cellular process and is enhanced by the familial Parkinson's disease-linked mutations. Proc. Natl. Acad. Sci. USA 102, 2162–2167 (2005).

  14. 14.

    et al. A precipitating role for truncated alpha-synuclein and the proteasome in alpha-synuclein aggregation: implications for pathogenesis of Parkinson disease. J. Biol. Chem. 280, 22670–22678 (2005).

  15. 15.

    , , & Impact of the acidic C-terminal region comprising amino acids 109-140 on alpha-synuclein aggregation in vitro. Biochemistry 43, 16233–16242 (2004).

  16. 16.

    et al. Role of alpha-synuclein carboxy-terminus on fibril formation in vitro. Biochemistry 42, 8530–8540 (2003).

  17. 17.

    et al. Distinct cleavage patterns of normal and pathologic forms of alpha-synuclein by calpain I in vitro. J. Neurochem. 86, 836–847 (2003).

  18. 18.

    et al. Cleavage of alpha-synuclein by calpain: potential role in degradation of fibrillized and nitrated species of alpha-synuclein. Biochemistry 44, 7818–7829 (2005).

  19. 19.

    , & Cathepsin D is the main lysosomal enzyme involved in the degradation of alpha-synuclein and generation of its carboxy-terminally truncated species. Biochemistry 47, 9678–9687 (2008).

  20. 20.

    et al. Oxidative stress-induced phosphorylation, degradation and aggregation of alpha-synuclein are linked to upregulated CK2 and cathepsin D. Eur. J. Neurosci. 26, 863–874 (2007).

  21. 21.

    , , , & Autocatalytic activation of human legumain at aspartic acid residues. FEBS Lett. 438, 114–118 (1998).

  22. 22.

    et al. Caspase cleavage of tau: linking amyloid and neurofibrillary tangles in Alzheimer's disease. Proc. Natl. Acad. Sci. USA 100, 10032–10037 (2003).

  23. 23.

    et al. Inhibition of mammalian legumain by some cystatins is due to a novel second reactive site. J. Biol. Chem. 274, 19195–19203 (1999).

  24. 24.

    et al. Neuroprotective actions of PIKE-L by inhibition of SET proteolytic degradation by asparagine endopeptidase. Mol. Cell 29, 665–678 (2008).

  25. 25.

    , , & SET protein (TAF1beta, I2PP2A) is involved in neuronal apoptosis induced by an amyloid precursor protein cytoplasmic subdomain. FASEB J. 19, 1905–1907 (2005).

  26. 26.

    , , , & Activation of asparaginyl endopeptidase leads to Tau hyperphosphorylation in Alzheimer disease. J. Biol. Chem. 288, 17495–17507 (2013).

  27. 27.

    et al. Asparaginyl endopeptidase cleaves TDP-43 in brain. Proteomics 12, 2455–2463 (2012).

  28. 28.

    et al. Cleavage of tau by asparagine endopeptidase mediates the neurofibrillary pathology in Alzheimer's disease. Nat. Med. 20, 1254–1262 (2014).

  29. 29.

    et al. Delta-secretase cleaves amyloid precursor protein and regulates the pathogenesis in Alzheimer's disease. Nat. Commun. 6, 8762 (2015).

  30. 30.

    , , , & Multistep autoactivation of asparaginyl endopeptidase in vitro and in vivo. J. Biol. Chem. 278, 38980–38990 (2003).

  31. 31.

    et al. Tyrosine and serine phosphorylation of alpha-synuclein have opposing effects on neurotoxicity and soluble oligomer formation. J. Clin. Invest. 119, 3257–3265 (2009).

  32. 32.

    , , , & α-Synuclein is phosphorylated by members of the Src family of protein-tyrosine kinases. J. Biol. Chem. 276, 3879–3884 (2001).

  33. 33.

    & α-Synuclein posttranslational modification and alternative splicing as a trigger for neurodegeneration. Mol. Neurobiol. 47, 509–524 (2013).

  34. 34.

    et al. Neuronal alpha-synucleinopathy with severe movement disorder in mice expressing A53T human alpha-synuclein. Neuron 34, 521–533 (2002).

  35. 35.

    et al. 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, 8968–8973 (2002).

  36. 36.

    et al. Age-related changes in brain energetics and phospholipid metabolism. NMR Biomed. 23, 242–250 (2010).

  37. 37.

    , , , & Protective mechanisms by cystatin C in neurodegenerative diseases. Front. Biosci. (Schol. Ed.) 3, 541–554 (2011).

  38. 38.

    & Structure and function of legumain in health and disease. Biochimie 122, 126–150 (2016).

  39. 39.

    , & Role of post-translational modifications in modulating the structure, function and toxicity of alpha-synuclein: implications for Parkinson's disease pathogenesis and therapies. Prog. Brain Res. 183, 115–145 (2010).

  40. 40.

    et al. Caspase-1 causes truncation and aggregation of the Parkinson's disease-associated protein α-synuclein. Proc. Natl. Acad. Sci. USA 113, 9587–9592 (2016).

  41. 41.

    et al. Biosynthetic processing of cathepsins and lysosomal degradation are abolished in asparaginyl endopeptidase-deficient mice. J. Biol. Chem. 278, 33194–33199 (2003).

  42. 42.

    , & Addition of exogenous α-synuclein preformed fibrils to primary neuronal cultures to seed recruitment of endogenous α-synuclein to Lewy body and Lewy neurite-like aggregates. Nat. Protoc. 9, 2135–2146 (2014).

  43. 43.

    et al. 7,8-dihydroxyflavone prevents synaptic loss and memory deficits in a mouse model of Alzheimer's disease. Neuropsychopharmacology 39, 638–650 (2014).

  44. 44.

    et al. Dopamine modulates diurnal and circadian rhythms of protein phosphorylation in photoreceptor cells of mouse retina. Eur. J. Neurosci. 27, 2691–2700 (2008).

  45. 45.

    , , & Protein measurement with the Folin phenol reagent. J. Biol. Chem. 193, 265–275 (1951).

Download references


This work was supported by grants from the Michael J. Fox Foundation (grant ID 11137) to K.Y.; a grant from the National Natural Science Foundation (NSFC) of China (no. 81571249) to Zhentao Zhang; NSFC grant (no. 81528007) to K.Y. and J.-Z.W.; a National Key Basic Research Program of China grant (2010CB945202) to Y.E.S.; an NSFC grant (81330030) to Y.E.S.; and grants from the US Public Health Service (P30EY006360 and R01EY004864) to P.M.I. We thank the ADRC at Emory University for providing human PD, LBD and healthy-control samples, and C. Watts (University of Cambridge) for providing anti-AEP.

Author information


  1. Department of Neurology, Renmin Hospital of Wuhan University, Wuhan, China.

    • Zhentao Zhang
    •  & Zhaohui Zhang
  2. Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, Georgia, USA.

    • Zhentao Zhang
    • , Seong Su Kang
    • , Xia Liu
    • , Eun Hee Ahn
    •  & Keqiang Ye
  3. Department of Ophthalmology, Emory University School of Medicine, Atlanta, Georgia, USA.

    • Li He
    •  & P Michael Iuvone
  4. Department of Pharmacology, Emory University School of Medicine, Atlanta, Georgia, USA.

    • P Michael Iuvone
  5. Department of Biochemistry, Emory University School of Medicine, Atlanta, Georgia, USA.

    • Duc M Duong
    •  & Nicholas T Seyfried
  6. Center for Neurodegenerative Diseases, Emory University School of Medicine, Atlanta, Georgia, USA.

    • Duc M Duong
    •  & Nicholas T Seyfried
  7. Translational Science and Molecular Medicine, Michigan State University, College of Human Medicine, and Hauenstein Neuroscience Center, Mercy Health Saint Mary's, Grand Rapids, Michigan, USA.

    • Matthew J Benskey
    •  & Fredric P Manfredsson
  8. Translational Center for Stem Cell Research, Tongji Hospital, Department of Regenerative Medicine, Tongji University School of Medicine, Shanghai, China.

    • Lingjing Jin
    •  & Yi E Sun
  9. Pathophysiology Department, School of Basic Medicine and the Collaborative Innovation Center for Brain Science, Key Laboratory of the Ministry of Education of China for Neurological Disorders, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.

    • Jian-Zhi Wang


  1. Search for Zhentao Zhang in:

  2. Search for Seong Su Kang in:

  3. Search for Xia Liu in:

  4. Search for Eun Hee Ahn in:

  5. Search for Zhaohui Zhang in:

  6. Search for Li He in:

  7. Search for P Michael Iuvone in:

  8. Search for Duc M Duong in:

  9. Search for Nicholas T Seyfried in:

  10. Search for Matthew J Benskey in:

  11. Search for Fredric P Manfredsson in:

  12. Search for Lingjing Jin in:

  13. Search for Yi E Sun in:

  14. Search for Jian-Zhi Wang in:

  15. Search for Keqiang Ye in:


K.Y. conceived the project, designed the experiments and wrote the manuscript. Zhentao Zhang designed and performed most of the experiments. S.S.K. and X.L. prepared primary neurons and assisted with animal experiments. M.J.B. and F.P.M. provided clones and packaged viral vectors. D.M.D. and N.T.S. performed the mass spectrometry analysis. L.H. and P.M.I. performed the HPLC experiments and critically read and edited the manuscript. Zhaohui Zhang, E.H.A., L.J., Y.E.S., F.P.M. and J.-Z.W. designed the experiments, assisted with data analysis and interpretation and critically read the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Lingjing Jin or Jian-Zhi Wang or Keqiang Ye.

Integrated supplementary information

Supplementary information

PDF files

  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–8.

  2. 2.

    Supplementary Data Set 1

    Uncropped images of gels.

  3. 3.

    Life Sciences Reporting Summary

About this article

Publication history