The two-stage pathway of ataxin-3 fibrillogenesis involves a polyglutamine-independent step.
The aggregation of ataxin-3 is associated with spinocerebellar ataxia type 3, which is characterized by the formation of intraneuronal aggregates. However, the mechanism of aggregation is currently not well understood. Ataxin-3 consists of a folded Josephin domain followed by two ubiquitin-interacting motifs and a C-terminal polyglutamine tract, which in the non-pathological form is less than 45 residues in length. We demonstrate that ataxin-3 with 64 glutamines (at(Q64)) undergoes a two-stage aggregation. The first stage involves formation of SDS-soluble aggregates, and the second stage results in formation of SDS-insoluble aggregates via the poly(Q) region. Both these first and second stage aggregates display typical amyloid-like characteristics. Under the same conditions at(Q15) and at(QHQ) undergo a single step aggregation event resulting in SDS-soluble aggregates, which does not involve the polyglutamine tract. These aggregates do not convert to the SDS-insoluble form. These observations demonstrate that ataxin-3 has an inherent capacity to aggregate through its non-polyglutamine domains. However, the presence of a pathological length polyglutamine tract introduces an additional step resulting in formation of a highly stable amyloid-like aggregate.
Ellisdon
Andrew M
AM
Department of Biochemistry and Molecular Biology, Monash University, P. O. Box 13D, Wellington Road, Clayton, Victoria 3800, Australia.
Thomas
Bronwen
B
Department of Biochemistry and Molecular Biology, Monash University, P. O. Box 13D, Wellington Road, Clayton, Victoria 3800, Australia.
Bottomley
Stephen P
SP
Department of Biochemistry and Molecular Biology, Monash University, P. O. Box 13D, Wellington Road, Clayton, Victoria 3800, Australia. Electronic address: steve.bottomley@med.monash.edu.au.
D000067699:Ataxin-3; D052160:Benzothiazoles; D001671:Biochemistry / Q000379:methods; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D046529:Microscopy, Electron, Transmission; D009419:Nerve Tissue Proteins / Q000737:chemistry*; D009687:Nuclear Proteins / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry*; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry; D012097:Repressor Proteins / Q000737:chemistry*; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D013844:Thiazoles / Q000737:chemistry; D014364:Tryptophan / Q000737:chemistry; D025801:Ubiquitin / Q000737:chemistry
D052160:Benzothiazoles; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins; D010455:Peptides; D011994:Recombinant Proteins; D012097:Repressor Proteins; D013844:Thiazoles; D025801:Ubiquitin; C009462:thioflavin T; C097188:polyglutamine; D012967:Sodium Dodecyl Sulfate; D014364:Tryptophan; C092341:ATXN3 protein, human; D000067699:Ataxin-3