Druggable genome screen identifies new regulators of the abundance and toxicity of ATXN3, the Spinocerebellar Ataxia type 3 disease protein

Ashraf, Naila S.; Sutton, Joanna R.; Yang, Yemen; Ranxhi, Bedri; Libohova, Kozeta; Shaw, Emily D.; Barget, Anna J.; Todi, Sokol V.; Paulson, Henry L.; Costa, Maria do Carmo

doi:10.1016/j.nbd.2019.104697

Cited by 16 publications

(12 citation statements)

References 62 publications

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“…To expand on our observations, we shifted our focus to a SCA3 model specific to the fly eye. We previously utilized similar models to observe the phenotypic deterioration that occurs with pathogenic ataxin-3 expression, as well as to perform screens of various molecules and genetic modifications that may ameliorate SCA3 ( Ashraf et al, 2020 ; Johnson et al, 2020 ). Observation of the eye allows for the detection of more subtle changes among groups of flies that might go undetected in other expression patterns.…”

Section: Resultsmentioning

confidence: 99%

Targeting the VCP-binding motif of ataxin-3 improves phenotypes in Drosophila models of Spinocerebellar Ataxia Type 3

Johnson

Libohova

Blount

et al. 2021

Neurobiology of Disease

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Of the family of polyglutamine (polyQ) neurodegenerative diseases, Spinocerebellar Ataxia Type 3 (SCA3) is the most common. Like other polyQ diseases, SCA3 stems from abnormal expansions in the CAG triplet repeat of its disease gene resulting in elongated polyQ repeats within its protein, ataxin-3. Various ataxin-3 protein domains contribute to its toxicity, including the valosin-containing protein (VCP)-binding motif (VBM). We previously reported that VCP, a homo-hexameric protein, enhances pathogenic ataxin-3 aggregation and exacerbates its toxicity. These findings led us to explore the impact of targeting the SCA3 protein by utilizing a decoy protein comprising the N-terminus of VCP (N-VCP) that binds ataxin-3’s VBM. The notion was that N-VCP would reduce binding of ataxin-3 to VCP, decreasing its aggregation and toxicity. We found that expression of N-VCP in Drosophila melanogaster models of SCA3 ameliorated various phenotypes, coincident with reduced ataxin-3 aggregation. This protective effect was specific to pathogenic ataxin-3 and depended on its VBM. Increasing the amount of N-VCP resulted in further phenotype improvement. Our work highlights the protective potential of targeting the VCP-ataxin-3 interaction in SCA3, a key finding in the search for therapeutic opportunities for this incurable disorder.

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Section: Resultsmentioning

confidence: 99%

Targeting the VCP-binding motif of ataxin-3 improves phenotypes in Drosophila models of Spinocerebellar Ataxia Type 3

Johnson

Libohova

Blount

et al. 2021

Neurobiology of Disease

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“…On the other side, CHIP was shown to promote the activation of NF-κB signaling ( 30 ). Additionally, an siRNA-based drug screen in mammalian cells expressing expanded ataxin-3 identified 15 genes which are related to TNF/NF-κB and ERK1/2 pathways and concluded that expanded ataxin-3 can be regulated by these pro-inflammatory and cell death/survival pathways ( 31 ). Moreover, ataxin-3 may be involved in deubiquitination of several signaling molecules and it is hard to predict how expanded ataxin-3 may affect certain modifications and thus the behavior or fate of such proteins.…”

Section: Discussionmentioning

confidence: 99%

Ataxin-3, The Spinocerebellar Ataxia Type 3 Neurodegenerative Disorder Protein, Affects Mast Cell Functions

et al. 2022

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Spinocerebellar ataxia type 3 (SCA3), also known as Machado-Joseph Disease, is a progressive neurodegenerative disorder characterized by loss of neuronal matter due to the expansion of the CAG repeat in the ATXN3/MJD1 gene and subsequent ataxin-3 protein. Although the underlying pathogenic protein expansion has been known for more than 20 years, the complexity of its effects is still under exploration. The ataxin-3 protein in its expanded form is known to aggregate and disrupt cellular processes in neuronal tissue but the role of the protein on populations of immune cells is unknown. Recently, mast cells have emerged as potential key players in neuroinflammation and neurodegeneration. Here, we examined the mast cell-related effects of ataxin-3 expansion in the brain tissues of 304Q ataxin-3 knock-in mice and SCA3 patients. We also established cultures of mast cells from the 304Q knock-in mice and examined the effects of 304Q ataxin-3 knock-in on the immune responses of these cells and on markers involved in mast cell growth, development and function. Specifically, our results point to a role for expanded ataxin-3 in suppression of mast cell marker CD117/c-Kit, pro-inflammatory cytokine TNF-α and NF-κB inhibitor IκBα along with an increased expression of the granulocyte-attracting chemokine CXCL1. These results are the beginning of a more holistic understanding of ataxin-3 and could point to the development of novel therapeutic targets which act on inflammation to mitigate symptoms of SCA3.

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“…The SCA3 mouse lines used above overexpress wild-type or polyQ-expanded human ATXN3. To probe whether physiological levels of expanded ATXN3 affect the abundance of Ub species in human neuronal cells, we assessed the levels of Ub species in neuronal progenitor cells (NPCs) derived from a hESC line harboring one disease and one healthy allele (SCA3, NIH registry # 0286) (Moore et al, 2019; Ashraf et al, 2020). For comparison, we also assessed NPCs derived from a hESC line harboring two nonpathogenic ATXN3 alleles (CTRL, NIH registry #0147).…”

Section: Resultsmentioning

confidence: 99%

“…Mouse embryonic fibroblasts (MEFs) from both Atxn3 KO mice and WT littermates (Reina et al, 2010) were maintained in DMEM with 10% FBS, 1% Non-essential Amino Acids solution and 1% penicillin/streptomycin at 37□ and 5% CO2. Control and SCA3 neuronal progenitor cells (NPCs) were generated from the respective human embryonic stem cell (hESC) lines (CTRL, NIH registry #0147; SCA3, NIH registry # 0286), and were maintained in STEMdiff Neural Progenitor Medium (NPM), as previously described (Ashraf et al, 2020). Where indicated, cells were treated with lactacystin (15μM; Enzo Life Sciences), Chloroquine (100μM; Sigma-Aldrich), or DMSO (Sigma-Aldrich) for 12 hours.…”

Section: Methodsmentioning

confidence: 99%

Regional and age-dependent changes in ubiquitination in cellular and mouse models of Spinocerebellar ataxia type 3

Luo¹,

Todi²,

Paulson³

et al. 2023

Preprint

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Spinocerebellar ataxia type 3 (SCA3), also known as Machado–Joseph disease, is the most common dominantly inherited ataxia. SCA3 is caused by a CAG repeat expansion in theATXN3gene that encodes an expanded tract of polyglutamine (polyQ) in the disease protein ataxin–3 (ATXN3). As a deubiquitinating enzyme, ATXN3 regulates numerous cellular processes including proteasome– and autophagy–mediated protein degradation. In SCA3 disease brain, polyQ–expanded ATXN3 accumulates with other cellular constituents, including ubiquitin (Ub)–modified proteins, in select areas like the cerebellum and the brainstem, but whether pathogenic ATXN3 affects the abundance of ubiquitinated species is unknown. Here, in mouse and cellular models of SCA3, we investigated whether elimination of murine Atxn3 or expression of wild-type or polyQ-expanded human ATXN3 alters soluble levels of overall ubiquitination, as well as K48–linked (K48–Ub) and K63–linked (K63–Ub) chains. Levels of ubiquitination were assessed in the cerebellum and brainstem of 7– and 47–week–oldAtxn3knockout and SCA3 transgenic mice, and also in relevant mouse and human cell lines. In older mice, we observed that wild–type ATXN3 impacts the cerebellar levels of K48 –Ub proteins. In contrast, pathogenic ATXN3 leads to decreased brainstem abundance of K48–Ub species in younger mice and changes in both cerebellar and brainstem K63–Ub levels in an age-dependent manner: younger SCA3 mice have higher levels of K63–Ub while older mice have lower levels of K63–Ub compared to controls. Human SCA3 neuronal progenitor cells also show a relative increase in K63–Ub proteins upon autophagy inhibition. We conclude that wild-type and mutant ATXN3 differentially impact K48–Ub– and K63–Ub–modified proteins in the brain in a region– and age–dependent manner.

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Druggable genome screen identifies new regulators of the abundance and toxicity of ATXN3, the Spinocerebellar Ataxia type 3 disease protein

Cited by 16 publications

References 62 publications

Targeting the VCP-binding motif of ataxin-3 improves phenotypes in Drosophila models of Spinocerebellar Ataxia Type 3

Targeting the VCP-binding motif of ataxin-3 improves phenotypes in Drosophila models of Spinocerebellar Ataxia Type 3

Ataxin-3, The Spinocerebellar Ataxia Type 3 Neurodegenerative Disorder Protein, Affects Mast Cell Functions

Regional and age-dependent changes in ubiquitination in cellular and mouse models of Spinocerebellar ataxia type 3

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