A Novel SCA3 Knock-in Mouse Model Mimics the Human SCA3 Disease Phenotype Including Neuropathological, Behavioral, and Transcriptional Abnormalities Especially in Oligodendrocytes

Spinocerebellar ataxia type 3 is the most common autosomal dominant inherited ataxia worldwide, caused by a CAG repeat expansion in the Ataxin-3 gene resulting in a polyglutamine (polyQ)-expansion in the corresponding protein. The disease is characterized by neuropathological, phenotypical, and specific transcriptional changes in affected brain regions. So far, there is no mouse model available representing all the different aspects of the disease, yet highly needed for a better understanding of the disease pathomechanisms. Here, we characterized a novel Ataxin-3 knock-in mouse model, expressing a heterozygous or homozygous expansion of 304 CAACAGs in the murine Ataxin-3 locus using biochemical, behavioral, and transcriptomic approaches. We compared neuropathological, and behavioral features of the new knock-in model with the in SCA3 research mostly used YAC84Q mouse model. Further, we compared transcriptional changes found in cerebellar samples of the SCA3 knock-in mice and post-mortem human SCA3 patients. The novel knock-in mouse is characterized by the expression of a polyQ-expansion in the murine Ataxin-3 protein, leading to aggregate formation, especially in brain regions known to be vulnerable in SCA3 patients, and impairment of Purkinje cells. Along these neuropathological changes, the mice showed a reduction in body weight accompanied by gait and balance instability. Transcriptomic analysis of cerebellar tissue revealed age-dependent differential expression, enriched for genes attributed to myelinating oligodendrocytes. Comparing these changes with those found in cerebellar tissue of SCA3 patients, we discovered an overlap of differentially expressed genes pointing towards similar gene expression perturbances in several genes linked to myelin sheaths and myelinating oligodendrocytes.

A Novel SCA3 Knock-in Mouse Model Mimics the Human SCA3 Disease Phenotype Including Neuropathological, Behavioral, and Transcriptional Abnormalities Especially in Oligodendrocytes

Spinocerebellar ataxia type 3 is the most common autosomal dominant inherited ataxia worldwide, caused by a CAG repeat expansion in the Ataxin-3 gene resulting in a polyQ-expansion in the corresponding protein. The disease is characterized by neuropathological, phenotypical, and specific transcriptional changes in affected brain regions. So far, there is no mouse model available representing all the different aspects of the disease, yet highly needed for a better understanding of the disease pathomechanisms. Here, we characterized a novel Ataxin-3 knock-in mouse model, expressing a heterozygous or homozygous expansion of 304 CAACAGs in the murine Ataxin-3 locus using biochemical, behavioral, and transcriptomic approaches. We compared neuropathological, and behavioral features of the new knock-in model with the in SCA3 research mostly used YAC84Q mouse model. Further, we compared transcriptional changes found in cerebellar samples of SCA3 the knock-in mice and post-mortem human SCA3 patients. The novel knock-in mouse is characterized by the expression of a polyQ-expansion in the murine Ataxin-3 protein, leading to aggregate formation, especially in brain regions known to be vulnerable in SCA3 patients, and impairment of Purkinje cells. Along these neuropathological changes, the mice showed a reduction in body weight accompanied by gait and balance instability. Transcriptomic analysis of cerebellar tissue revealed age-dependent differential expression, enriched for genes attributed to myelinating oligodendrocytes. Comparing these changes with those found in cerebellar tissue of SCA3 patients, we discovered an overlap of differentially expressed genes pointing towards similar gene expression perturbances in several genes linked to myelin sheaths and myelinating oligodendrocytes.

PolyQ-expanded proteins impair cellular proteostasis of ataxin-3 through sequestering the co-chaperone HSJ1 into aggregates

Polyglutamine (polyQ) expansion of proteins can trigger protein misfolding and amyloid-like aggregation, which thus lead to severe cytotoxicities and even the respective neurodegenerative diseases. However, why polyQ aggregation is toxic to cells is not fully elucidated. Here, we took the fragments of polyQ-expanded (PQE) ataxin-7 (Atx7) and huntingtin (Htt) as models to investigate the effect of polyQ aggregates on the cellular proteostasis of endogenous ataxin-3 (Atx3), a protein that frequently appears in diverse inclusion bodies. We found that PQE Atx7 and Htt impair the cellular proteostasis of Atx3 by reducing its soluble as well as total Atx3 level but enhancing formation of the aggregates. Expression of these polyQ proteins promotes proteasomal degradation of endogenous Atx3 and accumulation of its aggregated form. Then we verified that the co-chaperone HSJ1 is an essential factor that orchestrates the balance of cellular proteostasis of Atx3; and further discovered that the polyQ proteins can sequester HSJ1 into aggregates or inclusions in a UIM domain-dependent manner. Thereby, the impairment of Atx3 proteostasis may be attributed to the sequestration and functional loss of cellular HSJ1. This study deciphers a potential mechanism underlying how PQE protein triggers proteinopathies, and also provides additional evidence in supporting the hijacking hypothesis that sequestration of cellular interacting partners by protein aggregates leads to cytotoxicity or neurodegeneration.

PolyQ expansion does not alter the Huntingtin-HAP40 complex

SummaryThe abnormal amplification of a CAG repeat in the gene coding for huntingtin (HTT) leads to Huntington disease (HD). At the protein level, this translates into the expansion of a poly-glutamine (polyQ) stretch located at the HTT N-terminus, which renders it aggregation-prone by unknown mechanisms. Here we investigated the effects of polyQ expansion on HTT in a complex with its stabilizing interaction partner huntingtin-associated protein 40 (HAP40). Surprisingly, our comprehensive biophysical, crosslinking mass spectrometry and cryo-EM experiments revealed no major differences in the conformation of HTT-HAP40 complexes of various polyQ length, including 17QHTT-HAP40 (wild type), 46QHTT-HAP40 (typical polyQ length in HD patients) and 128QHTT-HAP40 (extreme polyQ length). Thus, HTT polyQ expansion does not alter the global structure of HTT when associated with HAP40.

Determining the Fate of Neurons in SCA3: ATX3, a Rising Decision Maker in Response to DNA Stresses and Beyond

DNA damage response (DDR) and apoptosis are reported to be involved in the pathogenesis of many neurodegenerative diseases including polyglutamine (polyQ) disorders, such as Spinocerebellar ataxia type 3 (SCA3) and Huntington's disease (HD). Consistently, an increasing body of studies provide compelling evidence for the crucial roles of ATX3, whose polyQ expansion is defined as the cause of SCA3, in the maintenance of genome integrity and regulation of apoptosis. The polyQ expansion in ATX3 seems to affect its physiological functions in these distinct pathways. These advances have expanded our understanding of the relationship between ATX3's cellular functions and the underlying molecular mechanism of SCA3. Interestingly, dysregulated DDR pathways also contribute to the pathogenesis of other neurodegenerative disorder such as HD, which presents a common molecular mechanism yet distinct in detail among different diseases. In this review, we provide a comprehensive overview of the current studies about the physiological roles of ATX3 in DDR and related apoptosis, highlighting the crosslinks between these impaired pathways and the pathogenesis of SCA3. Moreover, whether these mechanisms are shared in other neurodegenerative diseases are analyzed. Finally, the preclinical studies targeting DDR and related apoptosis for treatment of polyQ disorders including SCA3 and HD are also summarized and discussed.

SCA7 mouse cerebellar pathology reveals preferential downregulation of key Purkinje cell-identity genes and shared disease signature with SCA1 and SCA2.

Spinocerebellar ataxia type 7 (SCA7) is an inherited neurodegenerative disease mainly characterized by motor incoordination and visual impairment due to progressive cerebellar and retinal degeneration. Alteration of other nervous tissues also contributes to symptoms. The mechanisms underlying motor incoordination of SCA7 remain to be characterized. SCA7 is caused by a polyglutamine (polyQ) expansion in ATXN7, a member of the transcriptional coactivator SAGA complex, which harbors histone modification activities. PolyQ expansion in other proteins is responsible for 5 other SCAs (SCA1-3, 6 and 17). However, the converging and diverging pathophysiological points remain poorly understood. Using a new SCA7 knock-in model carrying 140 glutamines in ATXN7, we analyzed cell-type specific gene expression in the cerebellum. We show that gene deregulation affects all cerebellar cell types, although at variable degree, and correlates with alterations of SAGA-dependent epigenetic marks histone H3 acetylation and H2B ubiquitination. Our results further show that Purkinje cells (PCs) are far the most affected neurons: unlike other cerebellar cell types, PCs show reduced expression of 83 cell-type identity genes, critical for their spontaneous firing activity and synaptic functions. PC gene downregulation precedes morphological alterations, pacemaker dysfunction and motor incoordination. Strikingly, most PC identity genes downregulated in SCA7 mice are also decreased in early symptomatic SCA1 and SCA2 mice, revealing a common signature of early PC pathology involving cGMP-PKG and phosphatidylinositol signaling pathways and long-term depression. Our study thus points out molecular targets for therapeutic development which may prove beneficial for several SCAs. Finally, we show that unlike previous SCA7 mouse models, SCA7140Q/5Q mice exhibit the major disease features observed in patients, including cerebellar damage, cerebral atrophy, peripheral nerves pathology and photoreceptor dystrophy, which account for progressive impairment of behavior, motor and vision functions. Therefore, SCA7140Q/5Q mice represent an accurate model for the investigation of different aspects of SCA7 pathogenesis.

A novel Ataxin-3 knock-in mouse model mimics the human SCA3 disease phenotype including neuropathological, behavioral, and transcriptional abnormalities

BackgroundSpinocerebellar ataxia type 3 is the most common autosomal dominant inherited ataxia worldwide and is caused by a CAG repeat expansion in the Ataxin-3 gene resulting in a polyQ expansion in the corresponding protein. The disease is characterized by neuropathological (aggregate formation, cell loss), phenotypical (gait instability, body weight reduction), and specific transcriptional changes in affected brain regions. So far, there is no mouse model available representing all the different aspects of the disease, yet highly needed to gain a better understanding of the disease pathomechanism.MethodsHere, we characterized a novel Ataxin-3 knock-in mouse model, expressing either a heterozygous or homozygous expansion of 304 CAG/CAAs in the murine Ataxin-3 locus using biochemical, behavioral, and transcriptomic approaches. Further, we compared the transcriptional changes of the knock-in mice to those found in human SCA3 patients, to evaluate the comparability of our model.ResultsThe novel Ataxin-3 knock-in mouse is characterized by the expression of a polyQ-expansion in the murine Ataxin-3 protein, leading to massive aggregate formation, especially in brain regions known to be vulnerable in SCA3 patients, and impairment of Purkinje cells. Along these neuropathological changes, mice showed a reduction in body weight accompanied by gait and balance instability. Transcriptomic analysis of cerebellar tissue revealed age-dependent differential expression, enriched for genes attributed to myelinating oligodendrocytes. Comparing these transcriptional changes with those found in cerebellar tissue of SCA3 patients, we discovered an overlap of differentially expressed genes pointing towards similar gene expression perturbances in several genes linked to myelin sheaths and myelinating oligodendrocytes.ConclusionThe novel Ataxin-3 knock-in model shares neuropathological, behavioral, and transcriptomic features with human SCA3 patients and, therefore, represents an ideal model to investigate early-onset developments, therapy studies, or longitudinal biomarker alterations.

ATXN1 N-terminal region explains the binding differences of wild-type and expanded forms

Background Wild-type (wt) polyglutamine (polyQ) regions are implicated in stabilization of protein-protein interactions (PPI). Pathological polyQ expansion, such as that in human Ataxin-1 (ATXN1), that causes spinocerebellar ataxia type 1 (SCA1), results in abnormal PPI. For ATXN1 a larger number of interactors has been reported for the expanded (82Q) than the wt (29Q) protein. Methods To understand how the expanded polyQ affects PPI, protein structures were predicted for wt and expanded ATXN1, as well as, for 71 ATXN1 interactors. Then, the binding surfaces of wt and expanded ATXN1 with the reported interactors were inferred. Results Our data supports that the polyQ expansion alters the ATXN1 conformation and that it enhances the strength of interaction with ATXN1 partners. For both ATXN1 variants, the number of residues at the predicted binding interface are greater after the polyQ, mainly due to the AXH domain. Moreover, the difference in the interaction strength of the ATXN1 variants was due to an increase in the number of interactions at the N-terminal region, before the polyQ, for the expanded form. Conclusions There are three regions at the AXH domain that are essential for ATXN1 PPI. The N-terminal region is responsible for the strength of the PPI with the ATXN1 variants. How the predicted motifs in this region affect PPI is discussed, in the context of ATXN1 post-transcriptional modifications.

Juvenile Huntington’s Disease Skin Fibroblasts Respond with Elevated Parkin Level and Increased Proteasome Activity as a Potential Mechanism to Counterbalance the Pathological Consequences of Mutant Huntingtin Protein

Huntington’s disease (HD) is an inherited neurodegenerative disorder, caused by an abnormal polyglutamine (polyQ) expansion in the huntingtin protein (Htt). Mitochondrial dysfunction and impairment of the ubiquitin-proteasome system (UPS) are hallmarks of HD neurons. The extraneural manifestations of HD are still unclear. We investigated the crosstalk between mitochondria and proteolytic function in skin fibroblasts from juvenile HD patients. We found reduced mitosis, increased cell size, elevated ROS and increased mitochondrial membrane potential in juvenile HD fibroblasts, while cellular viability was maintained. Mitochondrial OXPHOS analysis did not reveal significant differences compared to control. However, the level of mitochondrial fusion and fission proteins was significantly lower and branching in the mitochondria network was reduced. We hypothesized that juvenile HD fibroblasts counterbalance cellular damage and mitochondrial network deficit with altered proteasome activity to promote cell survival. Our data reveal that juvenile HD fibroblasts exhibit higher proteasome activity, which was associated with elevated gene and protein expression of parkin. Moreover, we demonstrate elevated proteasomal degradation of the mitochondrial fusion protein Mfn1 in diseased cells compared to control cells. Our data suggest that juvenile HD fibroblasts respond to mutant polyQ expansion of Htt with enhanced proteasome activity and faster turnover of specific UPS substrates to protect cells.