Memantine suppresses the excitotoxicity but fails to rescue the ataxic phenotype in SCA1 model mice.

Spinocerebellar ataxia type 1 (SCA1) is a debilitating neurodegenerative disorder of the cerebellum and brainstem. Memantine has been proposed as a potential treatment for SCA1. It blocks N-methyl-D-aspartate (NMDA) receptors on neurons, reduces excitotoxicity and decreases neurodegeneration in Alzheimer models. However, in cerebellar neurodegenerative diseases, the potential value of memantine is still unclear. We investigated the effects of memantine on motor performance and synaptic transmission in the cerebellum in a mouse model where mutant ataxin 1 is specifically targeted to glia. Lentiviral vectors (LVV) were used to express mutant ataxin 1 selectively in Bergmann glia (BG). In mice transduced with the mutant ataxin 1, chronic treatment with memantine improved motor activity during initial tests, presumably due to preserved BG and Purkinje cell (PC) morphology and numbers. However, mice were unable to improve their rota rod scores during next days of training. Memantine also compromised improvement in the rota rod scores in control mice upon repetitive training. These effects may be due to the effects of memantine on plasticity (LTD suppression) and NMDA receptor modulation. Some effects of chronically administered memantine persisted even after its wash-out from brain slices. Chronic memantine reduced morphological signs of neurodegeneration in the cerebellum of SCA1 model mice. This resulted in an apparent initial reduction of ataxic phenotype, but memantine also affected cerebellar plasticity and ultimately compromised motor learning. We speculate that that clinical application of memantine in SCA1 might be hampered by its ability to suppress NMDA-dependent plasticity in cerebellar cortex.

Mapping SCA1 regional vulnerabilities reveals neural and skeletal muscle contributions to disease.

Spinocerebellar ataxia type 1 (SCA1) is a fatal neurodegenerative disease caused by an expanded polyglutamine tract in the widely expressed ataxin-1 (ATXN1) protein. To elucidate anatomical regions and cell types that underlie mutant ATXN1-induced disease phenotypes, we developed a floxed conditional knockin mouse (f-ATXN1146Q/2Q) with mouse Atxn1 coding exons replaced by human ATXN1 exons encoding 146 glutamines. f-ATXN1146Q/2Q mice manifested SCA1-like phenotypes including motor and cognitive deficits, wasting, and decreased survival. Central nervous system (CNS) contributions to disease were revealed using f-ATXN1146Q/2Q;Nestin-Cre mice, which showed improved rotarod, open field, and Barnes maze performance by 6-12 weeks of age. In contrast, striatal contributions to motor deficits using f-ATXN1146Q/2Q;Rgs9-Cre mice revealed that mice lacking ATXN1146Q/2Q in striatal medium-spiny neurons showed a trending improvement in rotarod performance at 30 weeks of age. Surprisingly, a prominent role for muscle contributions to disease was revealed in f-ATXN1146Q/2Q;ACTA1-Cre mice based on their recovery from kyphosis and absence of muscle pathology. Collectively, data from the targeted conditional deletion of the expanded allele demonstrated CNS and peripheral contributions to disease and highlighted the need to consider muscle in addition to the brain for optimal SCA1 therapeutics.

Dysregulation of alternative splicing in spinocerebellar ataxia type 1.

Spinocerebellar ataxia type 1 is caused by an expansion of the polyglutamine tract in ATAXIN-1. Ataxin-1 is broadly expressed throughout the brain and is involved in regulating gene expression. However, it is not yet known if mutant ataxin-1 can impact the regulation of alternative splicing events. We performed RNA sequencing in mouse models of spinocerebellar ataxia type 1 and identified that mutant ataxin-1 expression abnormally leads to diverse splicing events in the mouse cerebellum of spinocerebellar ataxia type 1. We found that the diverse splicing events occurred in a predominantly cell autonomous manner. A majority of the transcripts with misregulated alternative splicing events were previously unknown, thus allowing us to identify overall new biological pathways that are distinctive to those affected by differential gene expression in spinocerebellar ataxia type 1. We also provide evidence that the splicing factor Rbfox1 mediates the effect of mutant ataxin-1 on misregulated alternative splicing and that genetic manipulation of Rbfox1 expression modifies neurodegenerative phenotypes in a Drosophila model of spinocerebellar ataxia type 1 in vivo. Together, this study provides novel molecular mechanistic insight into the pathogenesis of spinocerebellar ataxia type 1 and identifies potential therapeutic strategies for spinocerebellar ataxia type 1.

Stressing the role of a short linear motif in ataxin-2 condensation.

Ataxin-2, an RNA-binding protein that is conserved across eukaryotes, is involved in stress granule assembly and age-associated neurodegenerative diseases. In this issue of Molecular Cell, Boeynaems et al.1 identify a short linear motif in ataxin-2 as a condensation switch, providing molecular insights into its essential role in cellular stress response.

Therapeutic Strategies for Spinocerebellar Ataxia Type 1.

Spinocerebellar ataxia type 1 (SCA1) is an autosomal dominant neurodegenerative disorder that affects one or two individuals per 100,000. The disease is caused by an extended CAG repeat in exon 8 of the ATXN1 gene and is characterized mostly by a profound loss of cerebellar Purkinje cells, leading to disturbances in coordination, balance, and gait. At present, no curative treatment is available for SCA1. However, increasing knowledge on the cellular and molecular mechanisms of SCA1 has led the way towards several therapeutic strategies that can potentially slow disease progression. SCA1 therapeutics can be classified as genetic, pharmacological, and cell replacement therapies. These different therapeutic strategies target either the (mutant) ATXN1 RNA or the ataxin-1 protein, pathways that play an important role in downstream SCA1 disease mechanisms or which help restore cells that are lost due to SCA1 pathology. In this review, we will provide a summary of the different therapeutic strategies that are currently being investigated for SCA1.

TR-FRET-Based Immunoassay to Measure Ataxin-2 as a Target Engagement Marker in Spinocerebellar Ataxia Type 2.

Spinocerebellar ataxia type 2 (SCA2) is an autosomal dominantly inherited neurodegenerative disease, which belongs to the trinucleotide repeat disease group with a CAG repeat expansion in exon 1 of the ATXN2 gene resulting in an ataxin-2 protein with an expanded polyglutamine (polyQ)-stretch. The disease is late manifesting leading to early death. Today, therapeutic interventions to cure the disease or even to decelerate disease progression are not available yet. Furthermore, primary readout parameter for disease progression and therapeutic intervention studies are limited. Thus, there is an urgent need for quantifiable molecular biomarkers such as ataxin-2 becoming even more important due to numerous potential protein-lowering therapeutic intervention strategies. The aim of this study was to establish a sensitive technique to measure the amount of soluble polyQ-expanded ataxin-2 in human biofluids to evaluate ataxin-2 protein levels as prognostic and/or therapeutic biomarker in SCA2. Time-resolved fluorescence energy transfer (TR-FRET) was used to establish a polyQ-expanded ataxin-2-specific immunoassay. Two different ataxin-2 antibodies and two different polyQ-binding antibodies were validated in three different concentrations and tested in cellular and animal tissue as well as in human cell lines, comparing different buffer conditions to evaluate the best assay conditions. We established a TR-FRET-based immunoassay for soluble polyQ-expanded ataxin-2 and validated measurements in human cell lines including iPSC-derived cortical neurons. Additionally, our immunoassay was sensitive enough to monitor small ataxin-2 expression changes by siRNA or starvation treatment. We successfully established the first sensitive ataxin-2 immunoassay to measure specifically soluble polyQ-expanded ataxin-2 in human biomaterials.

BDNF is altered in a brain-region specific manner and rescues deficits in Spinocerebellar Ataxia Type 1.

Spinocerebellar ataxia type 1 (SCA1) is an adult-onset, dominantly inherited neurodegenerative disease caused by the expanded polyQ tract in the protein ATAXIN1 (ATXN1) and characterized by progressive motor and cognitive impairments. There are no disease-modifying treatments or cures for SCA1. Brain-derived neurotrophic factor (BDNF) plays important role in cerebellar physiology and has shown therapeutic potential for cerebellar pathology in the transgenic mouse model of SCA1, ATXN1[82Q] line that overexpress mutant ATXN1 under a cerebellar Purkinje-cell-specific promoter. Here we demonstrate decreased expression of brain derived neurotrophic factor (BDNF) in the cerebellum and medulla of patients with SCA1. Early stages of disease seem most amenable to therapy. Thus, we next quantified Bdnf expression in Atxn1154Q/2Q mice, a knock-in mouse model of SCA1, during the early symptomatic disease stage in four clinically relevant brain regions: cerebellum, medulla, hippocampus and motor cortex. We found that during the early stages of disease, Bdnf mRNA expression is reduced in the hippocampus and cerebellum, while it is increased in the cortex and brainstem. Importantly, we observed that pharmacological delivery of recombinant BDNF improved motor and cognitive performance, and mitigated pathology in the cerebellum and hippocampus of Atxn1154Q/2Q mice. Our findings demonstrate brain-region specific deficiency of BDNF in SCA1 and show that reversal of low BDNF levels offers the potential for meaningful treatment of motor and cognitive deficits in SCA1.

On the identification of potential novel therapeutic targets for spinocerebellar ataxia type 1 (SCA1) neurodegenerative disease using EvoPPI3.

EvoPPI (http://evoppi.i3s.up.pt), a meta-database for protein-protein interactions (PPI), has been upgraded (EvoPPI3) to accept new types of data, namely, PPI from patients, cell lines, and animal models, as well as data from gene modifier experiments, for nine neurodegenerative polyglutamine (polyQ) diseases caused by an abnormal expansion of the polyQ tract. The integration of the different types of data allows users to easily compare them, as here shown for Ataxin-1, the polyQ protein involved in spinocerebellar ataxia type 1 (SCA1) disease. Using all available datasets and the data here obtained for Drosophila melanogaster wt and exp Ataxin-1 mutants (also available at EvoPPI3), we show that, in humans, the Ataxin-1 network is much larger than previously thought (380 interactors), with at least 909 interactors. The functional profiling of the newly identified interactors is similar to the ones already reported in the main PPI databases. 16 out of 909 interactors are putative novel SCA1 therapeutic targets, and all but one are already being studied in the context of this disease. The 16 proteins are mainly involved in binding and catalytic activity (mainly kinase activity), functional features already thought to be important in the SCA1 disease.

Decreasing mutant ATXN1 nuclear localization improves a spectrum of SCA1-like phenotypes and brain region transcriptomic profiles.

Spinocerebellar ataxia type 1 (SCA1) is a dominant trinucleotide repeat neurodegenerative disease characterized by motor dysfunction, cognitive impairment, and premature death. Degeneration of cerebellar Purkinje cells is a frequent and prominent pathological feature of SCA1. We previously showed that transport of ATXN1 to Purkinje cell nuclei is required for pathology, where mutant ATXN1 alters transcription. To examine the role of ATXN1 nuclear localization broadly in SCA1-like disease pathogenesis, CRISPR-Cas9 was used to develop a mouse with an amino acid alteration (K772T) in the nuclear localization sequence of the expanded ATXN1 protein. Characterization of these mice indicates that proper nuclear localization of mutant ATXN1 contributes to many disease-like phenotypes including motor dysfunction, cognitive deficits, and premature lethality. RNA sequencing analysis of genes with expression corrected to WT levels in Atxn1175QK772T/2Q mice indicates that transcriptomic aspects of SCA1 pathogenesis differ between the cerebellum, brainstem, cerebral cortex, hippocampus, and striatum.

Pre-ataxic loss of intrinsic plasticity and motor learning in a mouse model of SCA1.

Spinocerebellar ataxias are neurodegenerative diseases, the hallmark symptom of which is the development of ataxia due to cerebellar dysfunction. Purkinje cells, the principal neurons of the cerebellar cortex, are the main cells affected in these disorders, but the sequence of pathological events leading to their dysfunction is poorly understood. Understanding the origins of Purkinje cells dysfunction before it manifests is imperative to interpret the functional and behavioural consequences of cerebellar-related disorders, providing an optimal timeline for therapeutic interventions. Here, we report the cascade of events leading to Purkinje cells dysfunction before the onset of ataxia in a mouse model of spinocerebellar ataxia 1 (SCA1). Spatiotemporal characterization of the ATXN1[82Q] SCA1 mouse model revealed high levels of the mutant ATXN1[82Q] weeks before the onset of ataxia. The expression of the toxic protein first caused a reduction of Purkinje cells intrinsic excitability, which was followed by atrophy of Purkinje cells dendrite arborization and aberrant glutamatergic signalling, finally leading to disruption of Purkinje cells innervation of climbing fibres and loss of intrinsic plasticity of Purkinje cells. Functionally, we found that deficits in eyeblink conditioning, a form of cerebellum-dependent motor learning, precede the onset of ataxia, matching the timeline of climbing fibre degeneration and reduced intrinsic plasticity. Together, our results suggest that abnormal synaptic signalling and intrinsic plasticity during the pre-ataxia stage of spinocerebellar ataxias underlie an aberrant cerebellar circuitry that anticipates the full extent of the disease severity. Furthermore, our work indicates the potential for eyeblink conditioning to be used as a sensitive tool to detect early cerebellar dysfunction as a sign of future disease.

Generation of induced pluripotent stem cell(iPSC)line CJUHi001-A derived peripheral blood mononuclear cells of spinocerebellar ataxia type 1(SCA1) the CAG repeat mutation in ATXN1 gene.

Spinocerebellar ataxia type 1 (SCA1) is an autosomal dominant neurodegenerative disorder caused by CAG repeat mutations in the ATXN1 gene. In this study, we generated an induced pluripotent stem cell line (iPSC) by using non-integrating Sendai virus (SeV) from peripheral blood mononuclear cells(PBMCs)of SCA1 patient harboring a CAG repeat mutation in the ATXN1 gene. The induced patient-specific iPSC line with a normal karyotype and expresses pluripotent markers, it also shows differentiation totipotency and tridermogenesis in vitro. It may be an excellent model for studying spinocerebellar ataxia type 1 (SCA1) in vitro and will be beneficial for studying SCA1 pathogenesis and therapeutic intervention strategies.

Governing dynamics and preferential binding of the AXH domain influence the aggregation pathway of Ataxin-1.

The present state of understanding the mechanism of Spinocerebellar Ataxia-1, a fatal neurodegenerative disease linked to the protein Ataxin-1 (ATXN1), is baffled by a set of self-contradictory, and hence, inconclusive observations. This fallacy poses a bottleneck to the effective designing of curable drugs as the field is currently missing the specific druggable site. To understand the fundamentals of pathogenesis, we tried to decipher the intricacies of the extremely complicated landscape by targeting the relevant species that supposedly dictate the structure-function paradigm. The atomic-level description and characterization of the dynamism of the systems reveal the existence of structural polymorphism in all the leading stakeholders of the overall system. The very existence of conformational heterogeneity in every species creates numerous possible combinations of favorable interactions because of the variability in segmental cross-talks and hence claims its role in the choice of routes between functional activity and dysfunctional disease-causing aggregation. Despite this emergent configurational diversity, there is a common mode of operative intermolecular forces that dictates the extent of stability of all the multimeric complexes due to the localized population of a specific type of residue. The present research proposes a dynamic switch mechanism between aggregability and functional activity, based on the logical interpretation of the estimated variables, which is practically dictated by the effective concentration of the interacting species involved in the cell.

Suppressing gain-of-function proteins via CRISPR/Cas9 system in SCA1 cells.

SCAs are autosomal dominant neurodegenerative disorders caused by a gain-of-function protein with toxic activities, containing an expanded polyQ tract in the coding region. There are no treatments available to delay the onset, stop or slow down the progression of these pathologies. In this work we focus our attention on SCA1 which is one of the most common genotypes circulating in Italy. Here, we develop a CRISPR/Cas9-based approach to reduce both forms of the ATXN1 protein, normal and mutated with expanded polyQ. We started with the screening of 10 different sgRNAs able to target Exon 8 of the ATXN1 gene. The two most promising sgRNAs were validated in fibroblasts isolated from SCA1 patients, following the identification of the best transfection method for this type of cell. Our silencing approach significantly downregulated the expression of ataxin1, due to large deletions and the introduction of small changes in the ATXN1 gene, evidenced by NGS analysis, without major effects on cell viability. Furthermore, very few significant guide RNA-dependent off-target effects were observed. These preliminary results not only allowed us to identify the best transfection method for SCA1 fibroblasts, but strongly support CRISPR/Cas9 as a promising approach for the treatment of expanded polyQ diseases. Further investigations will be needed to verify the efficacy of our silencing system in SCA1 neurons and animal models.

Spatial and Temporal Diversity of Astrocyte Phenotypes in Spinocerebellar Ataxia Type 1 Mice.

While astrocyte heterogeneity is an important feature of the healthy brain, less is understood about spatiotemporal heterogeneity of astrocytes in brain disease. Spinocerebellar ataxia type 1 (SCA1) is a progressive neurodegenerative disease caused by a CAG repeat expansion in the gene Ataxin1 (ATXN1). We characterized astrocytes across disease progression in the four clinically relevant brain regions, cerebellum, brainstem, hippocampus, and motor cortex, of Atxn1154Q/2Q mice, a knock-in mouse model of SCA1. We found brain region-specific changes in astrocyte density and GFAP expression and area, early in the disease and prior to neuronal loss. Expression of astrocytic core homeostatic genes was also altered in a brain region-specific manner and correlated with neuronal activity, indicating that astrocytes may compensate or exacerbate neuronal dysfunction. Late in disease, expression of astrocytic homeostatic genes was reduced in all four brain regions, indicating loss of astrocyte functions. We observed no obvious correlation between spatiotemporal changes in microglia and spatiotemporal astrocyte alterations, indicating a complex orchestration of glial phenotypes in disease. These results support spatiotemporal diversity of glial phenotypes as an important feature of the brain disease that may contribute to SCA1 pathogenesis in a brain region and disease stage-specific manner.

Identifying Disease Signatures in the Spinocerebellar Ataxia Type 1 Mouse Cortex.

The neurodegenerative disease spinocerebellar ataxia type 1 (SCA1) is known to lead to the progressive degeneration of specific neuronal populations, including cerebellar Purkinje cells (PCs), brainstem cranial nerve nuclei and inferior olive nuclei, and spinocerebellar tracts. The disease-causing protein ataxin-1 is fairly ubiquitously expressed throughout the brain and spinal cord, but most studies have primarily focused on the role of ataxin-1 in the cerebellum and brainstem. Therefore, the functions of ataxin-1 and the effects of SCA1 mutations in other brain regions including the cortex are not well-known. Here, we characterized pathology in the motor cortex of a SCA1 mouse model and performed RNA sequencing in this brain region to investigate the impact of mutant ataxin-1 towards transcriptomic alterations. We identified progressive cortical pathology and significant transcriptomic changes in the motor cortex of a SCA1 mouse model. We also identified progressive, region-specific, colocalization of p62 protein with mutant ataxin-1 aggregates in broad brain regions, but not the cerebellum or brainstem. A cross-regional comparison of the SCA1 cortical and cerebellar transcriptomic changes identified both common and unique gene expression changes between the two regions, including shared synaptic dysfunction and region-specific kinase regulation. These findings suggest that the cortex is progressively impacted via both shared and region-specific mechanisms in SCA1.

Differential effects of Wnt-ß-catenin signaling in Purkinje cells and Bergmann glia in spinocerebellar ataxia type 1.

Spinocerebellar ataxia type 1 (SCA1) is a dominantly inherited neurodegenerative disease characterized by progressive ataxia and degeneration of specific neuronal populations, including Purkinje cells (PCs) in the cerebellum. Previous studies have demonstrated a critical role for various evolutionarily conserved signaling pathways in cerebellar patterning, such as the Wnt-ß-catenin pathway; however, the roles of these pathways in adult cerebellar function and cerebellar neurodegeneration are largely unknown. In this study, we found that Wnt-ß-catenin signaling activity was progressively enhanced in multiple cell types in the adult SCA1 mouse cerebellum, and that activation of this signaling occurs in an ataxin-1 polyglutamine (polyQ) expansion-dependent manner. Genetic manipulation of the Wnt-ß-catenin signaling pathway in specific cerebellar cell populations revealed that activation of Wnt-ß-catenin signaling in PCs alone was not sufficient to induce SCA1-like phenotypes, while its activation in astrocytes, including Bergmann glia (BG), resulted in gliosis and disrupted BG localization, which was replicated in SCA1 mouse models. Our studies identify a mechanism in which polyQ-expanded ataxin-1 positively regulates Wnt-ß-catenin signaling and demonstrate that different cell types have distinct responses to the enhanced Wnt-ß-catenin signaling in the SCA1 cerebellum, underscoring an important role of BG in SCA1 pathogenesis.

Indirect Negative Effect of Mutant Ataxin-1 on Short- and Long-Term Synaptic Plasticity in Mouse Models of Spinocerebellar Ataxia Type 1.

Spinocerebellar ataxia type 1 (SCA1) is an intractable progressive neurodegenerative disease that leads to a range of movement and motor defects and is eventually lethal. Purkinje cells (PC) are typically the first to show signs of degeneration. SCA1 is caused by an expansion of the polyglutamine tract in the ATXN1 gene and the subsequent buildup of mutant Ataxin-1 protein. In addition to its toxicity, mutant Ataxin-1 protein interferes with gene expression and signal transduction in cells. Recently, it is evident that ATXN1 is not only expressed in neurons but also in glia, however, it is unclear the extent to which either contributes to the overall pathology of SCA1. There are various ways to model SCA1 in mice. Here, functional deficits at cerebellar synapses were investigated in two mouse models of SCA1 in which mutant ATXN1 is either nonspecifically expressed in all cell types of the cerebellum (SCA1 knock-in (KI)), or specifically in Bergmann glia with lentiviral vectors expressing mutant ATXN1 under the control of the astrocyte-specific GFAP promoter. We report impairment of motor performance in both SCA1 models. In both cases, prominent signs of astrocytosis were found using immunohistochemistry. Electrophysiological experiments revealed alteration of presynaptic plasticity at synapses between parallel fibers and PCs, and climbing fibers and PCs in SCA1 KI mice, which is not observed in animals expressing mutant ATXN1 solely in Bergmann glia. In contrast, short- and long-term synaptic plasticity was affected in both SCA1 KI mice and glia-targeted SCA1 mice. Thus, non-neuronal mechanisms may underlie some aspects of SCA1 pathology in the cerebellum. By combining the outcomes of our current work with our previous data from the B05 SCA1 model, we further our understanding of the mechanisms of SCA1.

Epigenetic control of ataxin-1 in multiple sclerosis.

OBJECTIVE: ATXN1 encodes the polyglutamine protein ataxin-1, which we have demonstrated exerting an immunomodulatory function in the context of central nervous system (CNS) autoimmunity, in addition to its classical role in the neurodegenerative disorder spinocerebellar ataxia type 1 (SCA1). In this study, we dissected the contribution of DNA methylation to the regulation of ATXN1 in multiple sclerosis (MS). METHODS: We interrogated a DNA methylation dataset previously generated via bisulfate DNA sequencing (BS-seq) in sorted peripheral immune cytotypes (CD4+ and CD8+ T cells, CD19+ B cells, and CD14+ monocytes) isolated from untreated MS patients at symptoms onset. RESULTS: Here, we report that ATXN1 undergoes hypo-methylation at four distinct regions upon MS, exclusively in B cells. We also highlight how these differentially methylated sites overlap with other regulatory epigenetic marks and MS risk variants. Lastly, we employ luciferase assays to assess the functionality of these regions, showing that the loss of methylation leads to an increase in ATXN1 expression. INTERPRETATION: Altogether, these findings provide biological insights into ataxin-1 regulation in the immune system as well as into the molecular mechanisms underlying MS risk.

Identification of the ataxin-1 interaction network and its impact on spinocerebellar ataxia type 1.

BACKGROUND: Spinocerebellar ataxia type 1 (SCA1) is a neurodegenerative disease caused by a polyglutamine expansion in the ataxin-1 protein. The pathogenic mechanism resulting in SCA1 is still unclear. Protein-protein interactions affect the function and stability of ataxin-1. METHODS: Wild-type and mutant ataxin-1 were expressed in HEK-293T cells. The levels of expression were assessed using real-time polymerase chain reaction (PCR) and Western blots. Co-immunoprecipitation was done in HEK-293T cells expressing exogenous wild-type and mutant ataxin-1 using anti-Flag antibody following by tandem affinity purification in order to study protein-protein interactions. The candidate interacting proteins were validated by immunoprecipitation. Chromatin immunoprecipitation and high-throughput sequencing and RNA immunoprecipitation and high-throughput sequencing were performed using HEK-293T cells expressing wild-type or mutant ataxin-1. RESULTS: In this study using HEK-293T cells, we found that wild-type ataxin-1 interacted with MCM2, GNAS, and TMEM206, while mutant ataxin-1 lost its interaction with MCM2, GNAS, and TMEM206. Two ataxin-1 binding targets containing the core GGAG or AAAT were identified in HEK-293T cells using ChIP-seq. Gene Ontology analysis of the top ataxin-1 binding genes identified SLC6A15, NTF3, KCNC3, and DNAJC6 as functional genes in neurons in vitro. Ataxin-1 also was identified as an RNA-binding protein in HEK-293T cells using RIP-seq, but the polyglutamine expansion in the ataxin-1 had no direct effects on the RNA-binding activity of ataxin-1. CONCLUSIONS: An expanded polyglutamine tract in ataxin-1 might interfere with protein-protein or protein-DNA interactions but had little effect on protein-RNA interactions. This study suggested that the dysfunction of protein-protein or protein-DNA interactions is involved in the pathogenesis of SCA1.

Cross-species genetic screens identify transglutaminase 5 as a regulator of polyglutamine-expanded ataxin-1.

Many neurodegenerative disorders are caused by abnormal accumulation of misfolded proteins. In spinocerebellar ataxia type 1 (SCA1), accumulation of polyglutamine-expanded (polyQ-expanded) ataxin-1 (ATXN1) causes neuronal toxicity. Lowering total ATXN1, especially the polyQ-expanded form, alleviates disease phenotypes in mice, but the molecular mechanism by which the mutant ATXN1 is specifically modulated is not understood. Here, we identified 22 mutant ATXN1 regulators by performing a cross-species screen of 7787 and 2144 genes in human cells and Drosophila eyes, respectively. Among them, transglutaminase 5 (TG5) preferentially regulated mutant ATXN1 over the WT protein. TG enzymes catalyzed cross-linking of ATXN1 in a polyQ-length-dependent manner, thereby preferentially modulating mutant ATXN1 stability and oligomerization. Perturbing Tg in Drosophila SCA1 models modulated mutant ATXN1 toxicity. Moreover, TG5 was enriched in the nuclei of SCA1-affected neurons and colocalized with nuclear ATXN1 inclusions in brain tissue from patients with SCA1. Our work provides a molecular insight into SCA1 pathogenesis and an opportunity for allele-specific targeting for neurodegenerative disorders.