Ubiquitination of a New Form of α-Synuclein by Parkin from Human Brain: Implications for Parkinson's Disease

Parkinson's disease (PD) is a common neurodegenerative disorder characterized by the progressive accumulation in selected neurons of protein inclusions containing α-synuclein and ubiquitin. Rare inherited forms of PD are caused by autosomal dominant mutations in α-synuclein or by autosomal recessive mutations in parkin, an E3 ubiquitin ligase. We hypothesized that these two gene products interact functionally, namely, that parkin ubiquitinates α-synuclein normally and that this process is altered in autosomal recessive PD. We have now identified a protein complex in normal human brain that includes parkin as the E3 ubiquitin ligase, UbcH7 as its associated E2 ubiquitin conjugating enzyme, and a new 22-kilodalton glycosylated form of α-synuclein (αSp22) as its substrate. In contrast to normal parkin, mutant parkin associated with autosomal recessive PD failed to bind αSp22. In an in vitro ubiquitination assay, αSp22 was modified by normal but not mutant parkin into polyubiquitinated, high molecular weight species. Accordingly, αSp22 accumulated in a non-ubiquitinated form in parkin-deficient PD brains. We conclude that αSp22 is a substrate for parkin's ubiquitin ligase activity in normal human brain and that loss of parkin function causes pathological αSp22 accumulation. These findings demonstrate a critical biochemical reaction between the two PD-linked gene products and suggest that this reaction underlies the accumulation of ubiquitinated α-synuclein in conventional PD.

[1]  N. Hattori,et al.  Genetics of Parkinson's disease. , 1999, Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie.

[2]  A. Shevchenko,et al.  Mass spectrometric sequencing of proteins silver-stained polyacrylamide gels. , 1996, Analytical chemistry.

[3]  R. Kopito,et al.  Impairment of the ubiquitin-proteasome system by protein aggregation. , 2001, Science.

[4]  Gerald W. Hart,et al.  Glycosylation of Nucleocytoplasmic Proteins: Signal Transduction and O-GlcNAc , 2001, Science.

[5]  A. Lang,et al.  Parkinson's disease. First of two parts. , 1998, The New England journal of medicine.

[6]  A. Brookes,et al.  Point Mutations (Thr240Arg and Ala311Stop) in theParkinGene , 1998 .

[7]  C. Mariani,et al.  New mutation (R42P) of the parkin gene in the ubiquitinlike domain associated with parkinsonism , 2001, Neurology.

[8]  A. Weissman,et al.  RING Finger Proteins Mediators of Ubiquitin Ligase Activity , 2000, Cell.

[9]  S. Pulst,et al.  Parkin is associated with actin filaments in neuronal and nonneural cells , 2000, Annals of neurology.

[10]  P. Howley,et al.  The Role of E6AP in the Regulation of p53 Protein Levels in Human Papillomavirus (HPV)-positive and HPV-negative Cells* , 1998, The Journal of Biological Chemistry.

[11]  P. Lansbury,et al.  Is there a cause-and-effect relationship between α-synuclein fibrillization and Parkinson’s disease? , 2000, Nature Cell Biology.

[12]  M. Scheffner,et al.  The HPV-16 E6 and E6-AP complex functions as a ubiquitin-protein ligase in the ubiquitination of p53 , 1993, Cell.

[13]  J. Bressler,et al.  Genetics of Angelman syndrome. , 1999, American journal of human genetics.

[14]  P. Lansbury,et al.  Inhibition of fibrillization and accumulation of prefibrillar oligomers in mixtures of human and mouse alpha-synuclein. , 2000, Biochemistry.

[15]  Y. Imai,et al.  Parkin Suppresses Unfolded Protein Stress-induced Cell Death through Its E3 Ubiquitin-protein Ligase Activity* , 2000, The Journal of Biological Chemistry.