Spinocerebellar ataxia type 3 (SCA3), also known as Machado-Joseph disease, is a neurodegenerative disease caused by expanded polyglutamine repeats in exon 10 of the ataxin-3 gene, ATXN3. The accumulation of mutant ATXN3 protein leads to severe clinical manifestations and premature death. Clinically, SCA3 pathology is characterized by progressive ataxia leading to motor incoordination that may affect balance, gait and speech, and neuropathologically by a progressive degeneration of the spinal cord and cerebellum, as well as the cerebral cortex and basal ganglia. Although SCA3 is a rare disease, it is the most common autosomal dominant spinocerebellar ataxia worldwide. Its geographical distribution varies worldwide, with peak prevalence in certain regions of Brazil, Portugal and China. In 1994, the identification of the polyglutamine expansion in the ATXN3 gene made it possible not only to diagnose this pathology but also to dissect the mechanisms leading to cellular degeneration. As a monogenic disease for which only symptomatic treatment is available, the ATXN3 gene represents an attractive therapeutic target for gene editing strategies.

Machado-Joseph disease (MJD) is a devastating and incurable neurodegenerative disease characterised by progressive ataxia, difficulty speaking and swallowing. Consequently, affected individuals ultimately become wheelchair dependent, require constant care, and face a shortened life expectancy. The monogenic cause of MJD is expansion of a trinucleotide (CAG) repeat region within the ATXN3 gene, which results in polyglutamine (polyQ) expansion within the resultant ataxin-3 protein. While it is well established that the ataxin-3 protein functions as a deubiquitinating (DUB) enzyme and is therefore critically involved in proteostasis, several unanswered questions remain regarding the impact of polyQ expansion in ataxin-3 on its DUB function. Here we review the current literature surrounding ataxin-3\'s DUB function, its DUB targets, and what is known regarding the impact of polyQ expansion on ataxin-3\'s DUB function. We also consider the potential neuroprotective effects of ataxin-3\'s DUB function, and the intersection of ataxin-3\'s role as a DUB enzyme and regulator of gene transcription. Ataxin-3 is the principal pathogenic protein in MJD and also appears to be involved in cancer. As aberrant deubiquitination has been linked to both neurodegeneration and cancer, a comprehensive understanding of ataxin-3\'s DUB function is important for elucidating potential therapeutic targets in these complex conditions. In this review, we aim to consolidate knowledge of ataxin-3 as a DUB and unveil areas for future research to aid therapeutic targeting of ataxin-3\'s DUB function for the treatment of MJD and other diseases.

The accumulation of mutant ataxin-3 (Atx3) in neuronal nuclear inclusions is a pathological hallmark of Machado-Joseph disease (MJD), also known as Spinocerebellar Ataxia Type 3. Decreasing the protein aggregation burden is a possible disease-modifying strategy to tackle MJD and other neurodegenerative disorders for which only symptomatic treatments are currently available. We performed a drug repurposing screening to identify inhibitors of Atx3 aggregation with known toxicological and pharmacokinetic profiles. Interestingly, dopamine hydrochloride and other catecholamines are among the most potent inhibitors of Atx3 aggregation in vitro. Our results indicate that low micromolar concentrations of dopamine markedly delay the formation of mature amyloid fibrils of mutant Atx3 through the inhibition of the earlier oligomerization steps. Although dopamine itself does not cross the blood-brain barrier, dopamine levels in the brain can be increased by low doses of dopamine precursors and dopamine agonists commonly used to treat Parkinsonian symptoms. In agreement, treatment with levodopa ameliorated motor symptoms in a C. elegans model of MJD. These findings suggest a possible application of dopaminergic drugs to halt or reduce Atx3 accumulation in the brains of MJD patients.

Spinocerebellar ataxia type 3 (SCA3) is a dominantly inherited neurodegenerative disease caused by CAG (encoding glutamine) repeat expansion in the Ataxin-3 (ATXN3) gene. We have shown previously that ATXN3-depleted or pathogenic ATXN3-expressing cells abrogate polynucleotide kinase 3\'-phosphatase (PNKP) activity. Here, we report that ATXN3 associates with RNA polymerase II (RNAP II) and the classical nonhomologous end-joining (C-NHEJ) proteins, including PNKP, along with nascent RNAs under physiological conditions. Notably, ATXN3 depletion significantly decreased global transcription, repair of transcribed genes, and error-free double-strand break repair of a 3\'-phosphate-containing terminally gapped, linearized reporter plasmid. The missing sequence at the terminal break site was restored in the recircularized plasmid in control cells by using the endogenous homologous transcript as a template, indicating ATXN3\'s role in PNKP-mediated error-free C-NHEJ. Furthermore, brain extracts from SCA3 patients and mice show significantly lower PNKP activity, elevated p53BP1 level, more abundant strand-breaks in the transcribed genes, and degradation of RNAP II relative to controls. A similar RNAP II degradation is also evident in mutant ATXN3-expressing Drosophila larval brains and eyes. Importantly, SCA3 phenotype in Drosophila was completely amenable to PNKP complementation. Hence, salvaging PNKP\'s activity can be a promising therapeutic strategy for SCA3.

We describe a protocol for culturing neurons from transgenic zebrafish embryos to investigate the subcellular distribution and protein aggregation status of neurodegenerative disease-causing proteins. The utility of the protocol was demonstrated on cell cultures from zebrafish that transgenically express disease-causing variants of human fused in sarcoma (FUS) and ataxin-3 proteins, in order to study amyotrophic lateral sclerosis (ALS) and spinocerebellar ataxia type-3 (SCA3), respectively. A mixture of neuronal subtypes, including motor neurons, exhibited differentiation and neurite outgrowth in the cultures. As reported previously, mutant human FUS was found to be mislocalized from nuclei to the cytosol, mimicking the pathology seen in human ALS and the zebrafish FUS model. In contrast, neurons cultured from zebrafish expressing human ataxin-3 with disease-associated expanded polyQ repeats did not accumulate within nuclei in a manner often reported to occur in SCA3. Despite this, the subcellular localization of the human ataxin-3 protein seen in cell cultures was similar to that found in the SCA3 zebrafish themselves. The finding of similar protein localization and aggregation status in the neuronal cultures and corresponding transgenic zebrafish models confirms that this cell culture model is a useful tool for investigating the cell biology and proteinopathy signatures of mutant proteins for the study of neurodegenerative disease.

Bcl2-associated athanogene 2 (BAG2) shares a similar molecular structure and function with other BAG family members. Functioning as a co-chaperone, it interacts with the ATPase domain of the heat shock protein 70 (dHsp70) through its BAG domain. It also interacts with many other molecules and regulates various cellular functions. An increasing number of studies have indicated that BAG2 is involved in the pathogenesis of various diseases, including cancers and neurodegenerative diseases. This paper is a comprehensive review of the structure, functions, and protein interactions of BAG2. We also discuss its roles in diseases, including cancer, Alzheimer\'s disease, Parkinson\'s disease and spinocerebellar ataxia type-3. Further research on BAG2 could lead to an understanding of the pathogenesis of these disorders or even to novel therapeutic approaches.

Polyglutamine expansion is a hallmark of nine neurodegenerative diseases, with protein aggregation intrinsically linked to disease progression. Although polyglutamine expansion accelerates protein aggregation, the misfolding process is frequently instigated by flanking domains. For example, polyglutamine expansion in ataxin-3 allosterically triggers the aggregation of the catalytic Josephin domain. The molecular mechanism that underpins this allosteric aggregation trigger remains to be determined. Here, we establish that polyglutamine expansion increases the molecular mobility of two juxtaposed helices critical to ataxin-3 deubiquitinase activity. Within one of these helices, we identified a highly amyloidogenic sequence motif that instigates aggregation and forms the core of the growing fibril. Critically, by mutating residues within this key region, we decrease local structural fluctuations to slow ataxin-3 aggregation. This provides significant insight, down to the molecular level, into how polyglutamine expansion drives aggregation and explains the positive correlation between polyglutamine tract length, protein aggregation, and disease severity.

The expansion of a polyglutamine domain in the protein ataxin3 causes spinocerebellar ataxia type-3 (SCA3). However, there is little information to date about the upstream proteins in the ubiquitin-proteasome system of pathogenic ataxin3-80Q. Here, we report that BAG2 (Bcl-2 associated athanogene family protein 2) and BAG5 (Bcl-2-associated athanogene family protein 5) stabilise pathogenic ataxin3-80Q by inhibiting its ubiquitination as determined based on western blotting and co-immunofluorescence experiments. The association of the BAG2 and BAG5 proteins with pathogenic ataxin3-80Q strengthens the important roles of the BAG family in neurodegenerative diseases.