Dorfin localizes to Lewy bodies and ubiquitylates synphilin-1.
Parkinson's disease (PD) is a neurodegenerative disease characterized by loss of nigra dopaminergic neurons. Lewy bodies (LBs) are a characteristic neuronal inclusion in PD brains. In this study, we report that Dorfin, a RING finger-type ubiquityl ligase for mutant superoxide dismutase-1, was localized with ubiquitin in LBs. Recently, synphilin-1 was identified to associate with alpha-synuclein and to be a major component of LBs. We found that overexpression of synphilin-1 in cultured cells led to the formation of large juxtanuclear inclusions, but showed no cytotoxicity. Dorfin colocalized in these large inclusions with ubiquitin and proteasomal components. In contrast to full-length synphilin-1, overexpression of the central portion of synphilin-1, including ankyrin-like repeats, a coiled-coil domain, and an ATP/GTP-binding domain, predominantly led to the formation of small punctate aggregates scattered throughout the cytoplasm and showed cytotoxic effects. Dorfin and ubiquitin did not localize in these small aggregates. Overexpression of the N or C terminus of synphilin-1 did not lead to the formation of any aggregates. Dorfin physically bound and ubiquitylated synphilin-1 through its central portion, but did not ubiquitylate wild-type or mutant alpha-synuclein. These results suggest that the central domain of synphilin-1 has an important role in the formation of aggregates and cytotoxicity and that Dorfin may be involved in the pathogenic process of PD and LB formation by ubiquitylation of synphilin-1.
Department of Neurology, Nagoya University Graduate School of Medicine, Showa-ku, Nagoya 466-8550, Japan.
Takashi
T
Ito
Jun-Ichi
J
Niwa
Nozomi
N
Hishikawa
Shinsuke
S
Ishigaki
Manabu
M
Doyu
Gen
G
Sobue
D000368:Aged; D000818:Animals; D001921:Brain / Q000648:ultrastructure; D019556:COS Cells; D002352:Carrier Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D002470:Cell Survival; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000032:analysis* / Q000378:metabolism*; D005260:Female; D015870:Gene Expression; D006801:Humans; D007150:Immunohistochemistry; D016631:Lewy Bodies / Q000737:chemistry*; D008297:Male; D008875:Middle Aged; D016296:Mutagenesis; D009419:Nerve Tissue Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D009474:Neurons / Q000648:ultrastructure; D010300:Parkinson Disease / Q000378:metabolism*; D011993:Recombinant Fusion Proteins; D013482:Superoxide Dismutase / Q000235:genetics / Q000378:metabolism; D000072105:Superoxide Dismutase-1; D051843:Synucleins; D014162:Transfection; D025801:Ubiquitin / Q000032:analysis / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases; D051844:alpha-Synuclein
D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D009419:Nerve Tissue Proteins; D011993:Recombinant Fusion Proteins; C497604:SNCA protein, human; C118958:SNCAIP protein, human; C000606290:SOD1 protein, human; D051843:Synucleins; D025801:Ubiquitin; D051844:alpha-Synuclein; D013482:Superoxide Dismutase; D000072105:Superoxide Dismutase-1; C423571:RNF19A protein, human; D044767:Ubiquitin-Protein Ligases