The Machado–Joseph disease-associated mutant form of ataxin-3 regulates parkin ubiquitination and stability

Abstract

Machado-Joseph disease (MJD), the most common dominantly inherited ataxia worldwide, is caused by a polyglutamine (polyQ) expansion in the deubiquitinating (DUB) enzyme ataxin-3. Interestingly, MJD can present clinically with features of Parkinsonism. In this study, we identify parkin, an E3 ubiquitin-ligase responsible for a common familial form of Parkinson's disease, as a novel ataxin-3 binding partner. The interaction between ataxin-3 and parkin is direct, involves multiple domains and is greatly enhanced by parkin self-ubiquitination. Moreover, ataxin-3 deubiquitinates parkin directly in vitro and in cells. Compared with wild-type ataxin-3, MJD-linked polyQ-expanded mutant ataxin-3 is more active, possibly owing to its greater efficiency at DUB K27- and K29-linked Ub conjugates on parkin. Remarkably, mutant but not wild-type ataxin-3 promotes the clearance of parkin via the autophagy pathway. The finding is consistent with the reduction in parkin levels observed in the brains of transgenic mice over-expressing polyQ-expanded but not wild-type ataxin-3, raising the intriguing possibility that increased turnover of parkin may contribute to the pathogenesis of MJD and help explain some of its parkinsonian features.

Ataxin-3 Deubiquitination Is Coupled to Parkin Ubiquitination via E2 Ubiquitin-conjugating Enzyme

“…Goat anti-mouse light chain-specific antibodies (Jackson ImmunoResearch Laboratories) were used to detect ataxin-3 or parkin from immunoprecipitated SH-SY5 lysates or mouse brain fractions. All constructs used were described previously (19), with the exception of pCMV-Myc-Ubc7 (Addgene). Where indicated Ub-aldehyde (UBAL) (Boston Biochem) was added to a final concentration of 100 nM.…”

“…In Vitro Ubiquitination, Deubiquitination, and Charging Assays-In vitro ubiquitination and charging assays were carried out as described previously (19) with the reducing agent dithiothreitol (DTT) present at a final concentration of 1 mM. Charging reactions with UbcH7 (Fig.…”

“…We recently characterized an interaction between ataxin-3, an MJD class of DUB, and parkin, a RING-type E3 Ub-ligase (19). Mutations in SCA3, the gene encoding ataxin-3, cause MJD, also known as spinocerebellar ataxia-3 (SCA3) (20), the most common autosomal dominant inherited ataxia worldwide.…”

The E3 Ubiquitin Ligase Parkin Is Recruited to the 26 S Proteasome via the Proteasomal Ubiquitin Receptor Rpn13

“…Mutations within the Ubl domain render parkin insoluble and unstable (28,29). We have previously shown that the Ubl functions as a key interaction module allowing parkin to interact with proteins containing ubiquitin-binding domains, such as Eps15, Ataxin-3, and endophilin-A (8,10,30). Interestingly, Tsai et al (31) showed that the Ubl domain of parkin is necessary for its direct interaction with purified 26 S proteasomes in vitro.…”

Mouse Models of Spinocerebellar Ataxia Type 3 (Machado-Joseph Disease)

“…1) Model 1 (and predictably model 4) has peripheral neuropathy [32], metabolic abnormalities in cerebellum [49], deranged calcium signaling [40,50], reduced parkin levels possibly causing parkinsonism [51]. 2) Model 4 (and predictable model 1) has transcriptional dysregulation in the cerebellum [35], which is reversible with sodium butyrate [52].…”