Human Molecular Genetics Advance Access originally published online on February 10, 2009
Human Molecular Genetics 2009 18(9):1556-1565; doi:10.1093/hmg/ddp067
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The ataxia protein sacsin is a functional co-chaperone that protects against polyglutamine-expanded ataxin-1
1 William Harvey Research Institute and 2 Institute of Cell and Molecular Science, Barts and the London School of Medicine and Dentistry, Queen Mary University of London, London, UK 3 MRC Centre for Neurodegeneration Research, King's College London, Institute of Psychiatry, London, UK 4 Division of Molecular and Cellular Neuroscience, UCL Institute of Ophthalmology, London, UK and 5 Biomedical Biotechnology Research Unit, Department of Biochemistry, Microbiology and Biotechnology, Rhodes University, Grahamstown, South Africa
* To whom correspondence should be addressed at: Centre for Endocrinology, William Harvey Research Institute, Barts and the London School of Medicine and Dentistry, Charterhouse Square, London EC1M 6BQ, UK. Tel: +44 20 7882 6242; Fax: +44 20 7882 6197; Email: j.p.chapple{at}qmul.ac.uk
Received November 14, 2008; Revised January 19, 2009; Accepted February 5, 2009
An extensive protein–protein interaction network has been identified between proteins implicated in inherited ataxias. The protein sacsin, which is mutated in the early-onset neurodegenerative disease autosomal recessive spastic ataxia of Charlevoix-Saguenay, is a node in this interactome. Here, we have established the neuronal expression of sacsin and functionally characterized domains of the 4579 amino acid protein. Sacsin is most highly expressed in large neurons, particularly within brain motor systems, including cerebellar Purkinje cells. Its subcellular localization in SH-SY5Y neuroblastoma cells was predominantly cytoplasmic with a mitochondrial component. We identified a putative ubiquitin-like (UbL) domain at the N-terminus of sacsin and demonstrated an interaction with the proteasome. Furthermore, sacsin contains a predicted J-domain, the defining feature of DnaJ/Hsp40 proteins. Using a bacterial complementation assay, the sacsin J-domain was demonstrated to be functional. The presence of both UbL and J-domains in sacsin suggests that it may integrate the ubiquitin–proteasome system and Hsp70 function to a specific cellular role. The Hsp70 chaperone machinery is an important component of the cellular response towards aggregation prone mutant proteins that are associated with neurodegenerative diseases. We therefore investigated the effects of siRNA-mediated sacsin knockdown on polyglutamine-expanded ataxin-1. Importantly, SACS siRNA did not affect cell viability with GFP-ataxin-1[30Q], but enhanced the toxicity of GFP-ataxin-1[82Q], suggesting that sacsin is protective against mutant ataxin-1. Thus, sacsin is an ataxia protein and a regulator of the Hsp70 chaperone machinery that is implicated in the processing of other ataxia-linked proteins.
The authors wish it to be known that, in their opinion, the first two authors should be regarded as joint First Authors.