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Abstract Background Spinocerebellar ataxia type 1, is a rare neurodegenerative disorder with autosomal dominant inheritance belonging to the polyglutamine diseases. The diagnosis of this disease requires genetic testing that may also include the search for CAT interruption of the CAG repeat tract. Case presentation One 23-years-old patient suffers from a severe ataxia, with early-onset and rapid progression of the disease. His father might have been affected, but no molecular confirmation has been performed. The genetic results were negative for the Friedreich’s ataxia, spinocerebellar ataxia type 2, 3, 6, 7 and 17. The numbers of CAG repeats in the ATXN1 gene was assessed by fluorescent PCR, tripled-primed PCR and enzymatic digestion for the search of sequence interruption in the CAG repeats. The patient carried one pathogenic allele of 61 CAG and one intermediate allele of 37 CAG in the ATXN1 gene. Both alleles were uninterrupted. Conclusions We report a rare case of spinocerebellar ataxia type 1 with an intermediate allele and a large SCA1 expansion. The determination of the absence of CAT interruption brought crucial information concerning this molecular diagnosis, the prediction of the disease and had practical consequences for genetic counseling.

Background: Spinocerebellar ataxia type 1, is a rare neurodegenerative disorder with autosomal dominant inheritance belonging to the polyglutamine diseases. The diagnosis of this disease requires genetic testing that may also include the search for CAT interruption of the CAG repeat tract. Case presentation: One 23-years-old patient suffers from a severe ataxia, with early-onset and rapid progression of the disease. His father might have been affected, but no molecular confirmation has been performed. The genetic results were negative for the Friedreich's ataxia, spinocerebellar ataxia type 2, 3, 6, 7 and 17. The numbers of CAG repeats in the ATXN1 gene was assessed by fluorescent PCR, tripled-primed PCR and enzymatic digestion for the search of sequence interruption in the CAG repeats. The patient carried one pathogenic allele of 61 CAG and one intermediate allele of 37 CAG in the ATXN1 gene. Both alleles were uninterrupted. Conclusions: We report a rare case of spinocerebellar ataxia type 1 with an intermediate allele and a large SCA1 expansion. The determination of the absence of CAT interruption brought crucial information concerning this molecular diagnosis, the prediction of the disease and had practical consequences for genetic counseling. [ABSTRACT FROM AUTHOR]

Machado–Joseph disease (MJD) is an autosomal dominant neurodegenerative spinocerebellar ataxia caused by a polyglutamine-coding CAG repeat expansion in the ATXN3 gene. While the CAG length correlates negatively with the age at onset, it accounts for approximately 50% of its variability only. Despite larger efforts in identifying contributing genetic factors, candidate genes with a robust and plausible impact on the molecular pathogenesis of MJD are scarce. Therefore, we analysed missense single nucleotide polymorphism variants in the PRKN gene encoding the Parkinson's disease-associated E3 ubiquitin ligase parkin, which is a well-described interaction partner of the MJD protein ataxin-3, a deubiquitinase. By performing a correlation analysis in the to-date largest MJD cohort of more than 900 individuals, we identified the V380L variant as a relevant factor, decreasing the age at onset by 3 years in homozygous carriers. Functional analysis in an MJD cell model demonstrated that parkin V380L did not modulate soluble or aggregate levels of ataxin-3 but reduced the interaction of the two proteins. Moreover, the presence of parkin V380L interfered with the execution of mitophagy—the autophagic removal of surplus or damaged mitochondria—thereby compromising cell viability. In summary, we identified the V380L variant in parkin as a genetic modifier of MJD, with negative repercussions on its molecular pathogenesis and disease age at onset. [ABSTRACT FROM AUTHOR]

Abstract Background Spinal and bulbar muscular atrophy (SBMA) is a hereditary neuromuscular disorder caused by the expansion of trinucleotide cytosine–adenine–guanine (CAG) repeats, which encodes a polyglutamine (polyQ) tract in the androgen receptor (AR) gene. Recent evidence suggests that, in addition to motor neuron degeneration, defective skeletal muscles are also the primary contributors to the pathogenesis in SBMA. While benefits of physical exercise have been suggested in SBMA, underlying mechanism remains elusive. Methods We investigated the effect of running exercise in a transgenic mouse model of SBMA carrying human AR with 97 expanded CAGs (AR97Q). We assigned AR97Q mice to exercise and sedentary control groups, and mice in the exercise group received 1‐h forced running wheel (5 m/min) 5 days a week for 4 weeks during the early stage of the disease. Motor function (grip strength and rotarod performance) and survival of each group were analysed, and histopathological and biological features in skeletal muscles and motor neurons were evaluated. Results AR97Q mice in the exercise group showed improvement in motor function (~40% and ~50% increase in grip strength and rotarod performance, respectively, P

Neuronal intranuclear inclusions (NIIs) are common key structures in polyglutamine (polyQ) diseases such as Huntington disease (HD), spinocerebellar ataxia type 1 (SCA1), and SCA3. Marinesco bodies (MBs) of dopaminergic neurons in the substantia nigra are also intranuclear structures and are frequently seen in normal elderly people. Ribosomal dysfunction is closely related to two differential processes; therefore, we aimed to identify the pathological characteristics of ribosomal protein SA (RPSA), a ribosomal protein, in both states. To this end, we evaluated the autopsy findings in four patients with HD, two SCA3, and five normal elderly cases (NCs). Immunohistochemical studies demonstrated that both NIIs and MBs contain RPSA. In polyQ diseases, RPSA was co‐localized with polyQ aggregations, and 3D‐reconstructed images revealed their mosaic‐like distribution. Assessments of the organization of RPSA and p62 in NIIs showed that RPSA was more localized toward the center than p62 and that this unique organization was more evident in the MBs. Immunoblotting of the temporal cortices revealed that the nuclear fraction of HD patients contained more RPSA than that of NCs. In conclusion, our study revealed that RPSA is a common component of both NIIs and MBs, indicating that a similar mechanism contributes to the formation of polyQ NIIs and MBs. [ABSTRACT FROM AUTHOR]

Incidence of cancer is markedly reduced in patients with the hereditary neurodegenerative polyglutamine (polyQ) diseases. We have very poor knowledge of the underlying molecular mechanisms, but the expanded polyQ sequence is assumed to play a central role, because it is common to the respective disease related proteins. The inhibition seems to take place in all kinds of cells, because the lower cancer frequency applies to nearly all types of tumors and is not related with the characteristic pathological changes in specific brain tissues. Further, the cancer repressing mechanisms appear to be active early in life including in pre-symptomatic and early phase polyQ patients. Autophagy plays a central role in clearing proteins with expanded polyQ tracts, and autophagy modulation has been demonstrated and particularly investigated in Huntington's disease (HD). Macroautophagy may be dysfunctional due to defects in several steps of the process, whereas increased chaperone-mediated autophagy (CMA) has been shown in HD patients, cell and animal models. Recently, CMA is assumed to play a key role in prevention of cellular transformation of normal cells into cancer cells. Investigations of normal cells from HD and other polyQ carriers could therefore add further insight into the protective mechanisms of CMA in tumorigenesis, and be important for development of autophagy based strategies to prevent malignant processes leading to cancer and neurodegeneration. [ABSTRACT FROM AUTHOR]

Polyglutamine (polyQ) diseases are rare autosomal-dominant neurodegenerative diseases associated with the expansion of glutamine-encoding triplet repeats in certain genes. To investigate the functional influence of repeat expansion on disease mechanisms, we applied a biallelic genome-engineering platform that we recently established, called Universal Knock-in System or UKiS, to develop a human cell trio, a set of three isogenic cell lines that are homozygous for two different numbers of repeats (first and second lines) or heterozygous for the two repeat numbers (third line). As an example of a polyQ disease, we chose spinocerebellar ataxia type 2 (SCA2). In a pseudodiploid human cell line, both alleles of the glutamine-encoding triplet repeat in the SCA2-causing gene, ataxin 2 or ATXN2, were first knocked in with a donor sequence encoding both thymidine kinase and either puromycin or blasticidin resistance proteins under dual drug selection. The knocked-in donor alleles were then substituted with a payload having either 22 or 76 triplet repeats in ATXN2 by ganciclovir negative selection. The two-step substitution and subsequent SNP typing and genomic sequencing confirmed that the SCA2-modeling isogenic cell trio was obtained: three clones of 22-repeat homozygotes, two clones of 22/76-repeat heterozygotes and two clones of 76-repeat homozygotes. Finally, RT-PCR and immunoblotting using the obtained clones showed that, consistent with previous observations, glutamine tract expansion reduced transcriptional and translational expression of ATXN2. The cell clones with homozygous longrepeat alleles, which are rarely obtained from patients with SCA2, showed more drastic reduction of ATXN2 expression than the heterozygous clones. This study thus demonstrates the potential of UKiS, which is a beneficial platform for the efficient development of cell models not only for polyQ diseases but also for any other genetic diseases, which may accelerate our deeper understanding of disease mechanisms and cell-based screening for therapeutic drugs. [ABSTRACT FROM AUTHOR]

Machado-Joseph disease (MJD) is a dominant neurodegenerative disease caused by an expanded CAG repeat in the ATXN3 gene encoding the ataxin-3 protein. Several cellular processes, including transcription and apoptosis, are disrupted in MJD. To gain further insights into the extent of dysregulation of mitochondrial apoptosis in MJD and to evaluate if expression alterations of specific apoptosis genes/proteins can be used as transcriptional biomarkers of disease, the expression levels of BCL2, BAX and TP53 and the BCL2/BAX ratio (an indicator of susceptibility to apoptosis) were assessed in blood and post-mortem brain samples from MJD subjects and MJD transgenic mice and controls. While patients show reduced levels of blood BCL2 transcripts, this measurement displays low accuracy to discriminate patients from matched controls. However, increased levels of blood BAX transcripts and decreased BCL2/BAX ratio are associated with earlier onset of disease, indicating a possible association with MJD pathogenesis. Post-mortem MJD brains show increased BCL2/BAX transcript ratio in the dentate cerebellar nucleus (DCN) and increased BCL2/BAX insoluble protein ratio in the DCN and pons, suggesting that in these regions, severely affected by degeneration in MJD, cells show signs of apoptosis resistance. Interestingly, a follow-up study of 18 patients further shows that blood BCL2 and TP53 transcript levels increase over time in MJD patients. Furthermore, while the similar levels of blood BCL2, BAX, and TP53 transcripts observed in preclinical subjects and controls is mimicked by pre-symptomatic MJD mice, the expression profile of these genes in patient brains is partially replicated by symptomatic MJD mice. Globally, our findings indicate that there is tissue-specific vulnerability to apoptosis in MJD subjects and that this tissue-dependent behavior is partially replicated in a MJD mouse model. [ABSTRACT FROM AUTHOR]

Untranslated regions are involved in the regulation of transcriptional and post-transcriptional processes. Characterization of these regions remains poorly explored for ATXN3, the causative gene of Machado-Joseph disease (MJD). Although a few genetic modifiers have been identified for MJD age at onset (AO), they only explain a small fraction of the AO variance. Our aim was to analyse variation at the 3′UTR of ATXN3 in MJD patients, analyse its impact on AO and attempt to build haplotypes that might discriminate between normal and expanded alleles. After assessing ATXN3 3′UTR variants in molecularly confirmed MJD patients, an in silico analysis was conducted to predict their functional impact (e.g. their effect on miRNA-binding sites). Alleles in cis with the expanded (CAG)n were inferred from family data, and haplotypes were built. The effect of the alternative alleles on the AO and on SARA and NESSCA ataxia scales was tested. Nine variants, all previously described, were found. For eight variants, in silico analyses predicted (a) deleterious effects (rs10151135; rs55966267); (b) changes on miRNA-binding sites (rs11628764; rs55966267; rs709930) and (c) alterations of RNA-binding protein (RBP)-binding sites (rs1055996; rs910369; rs709930; rs10151135; rs3092822; rs7158733). Patients harbouring the alternative allele at rs10151135 had significantly higher SARA Axial subscores (p = 0.023), comparatively with those homozygous for the reference allele. Ten different haplotypes were obtained, one of which was exclusively found in cis with the expanded and four with the normal allele. These findings, which are relevant for the design of allele-specific therapies, warrant further investigation in independent MJD cohorts. [ABSTRACT FROM AUTHOR]

PQBP3 is a protein binding to polyglutamine tract sequences that are expanded in a group of neurodegenerative diseases called polyglutamine diseases. The function of PQBP3 was revealed recently as an inhibitor protein of proteasome-dependent degradation of Lamin B1 that is shifted from nucleolus to peripheral region of nucleus to keep nuclear membrane stability. Here, we address whether PQBP3 is an intrinsically disordered protein (IDP) like other polyglutamine binding proteins including PQBP1, PQBP5 and VCP. Multiple bioinformatics analyses predict that N -terminal region of PQBP3 is unstructured. High-speed atomic force microscopy (HS-AFM) reveals that N -terminal region of PQBP3 is dynamically changed in the structure consistently with the predictions of the bioinformatics analyses. These data support that PQBP3 is also an IDP. • This study addresses whether PQBP3/NOL7 is an intrinsically disordered protein. • IUPred2A predicts N -terminal region of PQBP3/NOL7 unstructured. • AlfaFold2 also predicts N -terminal region of PQBP3/NOL7 unstructured. • HS-AFM reveals N -terminal region of PQBP3 to change dynamically in the structure. [ABSTRACT FROM AUTHOR]

Synaptojanin 2 binding protein (SYNJ2BP) is an outer mitochondrial membrane protein with a cytosolic PDZ domain that functions as a cellular signaling hub. Few studies have evaluated its role in disease. Here we use induced pluripotent stem cell (iPSC)-derived motor neurons and post-mortem tissue from patients with two hereditary motor neuron diseases, spinal and bulbar muscular atrophy (SBMA) and amyotrophic lateral sclerosis type 4 (ALS4), and show that SYNJ2BP expression is increased in diseased motor neurons. Similarly, we show that SYNJ2BP expression increases in iPSC-derived motor neurons undergoing stress. Using proteomic analysis, we found that elevated SYNJ2BP alters the cellular distribution of mitochondria and increases mitochondrial-ER membrane contact sites. Furthermore, decreasing SYNJ2BP levels improves mitochondrial oxidative function in the diseased motor neurons. Together, our observations offer new insight into the molecular pathology of motor neuron disease and the role of SYNJ2BP in mitochondrial dysfunction.

Background: Dentatorubral‐pallidoluysian atrophy (DRPLA) is an autosomal dominant neurodegenerative disorder caused by the unstable expansion of CAG repeats encoding polyglutamine stretches in DRPLA gene (ATN1). DRPLAp, the gene product of ATN1, is located in the nucleus and has been shown to function as a transcriptional coregulator. Its target genes, however, remain largely unknown. Aim: To elucidate the physiological functions of DRPLAp based on determination of the target genes regulated by wild‐type DRPLAp. Methods: Using the cultured cells expressing wild‐type full‐length DRPLA cDNA (Q19) encoding DRPLAp under the control of the doxycycline (Dox)‐On system, we searched for genes whose expression levels were regulated by wild‐type DRPLAp (Q19). Results: On the basis of detailed RNA‐seq analyses, we identified seven up‐regulated genes and five down‐regulated genes in Q19 cells. In gene ontology analysis, the up‐regulated differentially expressed genes formed clusters featuring skeletal system morphogenesis, transcriptional regulation, and RNA metabolism, which is consistent with the recent report that the mutations in ATN1 lead to recognizable facial form and variable congenital anomalies. Conclusion: These findings support that DRPLAp may play a role as a transcriptional coregulator involved in the regulation of development. [ABSTRACT FROM AUTHOR]

Machado-Joseph disease (MJD) is a dominant neurodegenerative disease caused by an expanded CAG repeat in the ATXN3 gene encoding the ataxin-3 protein. Several cellular processes, including transcription and apoptosis, are disrupted in MJD. To gain further insights into the extent of dysregulation of mitochondrial apoptosis in MJD and to evaluate if expression alterations of specific apoptosis genes/proteins can be used as transcriptional biomarkers of disease, the expression levels of BCL2, BAX and TP53 and the BCL2/BAX ratio (an indicator of susceptibility to apoptosis) were assessed in blood and post-mortem brain samples from MJD subjects and MJD transgenic mice and controls. While patients show reduced levels of blood BCL2 transcripts, this measurement displays low accuracy to discriminate patients from matched controls. However, increased levels of blood BAX transcripts and decreased BCL2/BAX ratio are associated with earlier onset of disease, indicating a possible association with MJD pathogenesis. Post-mortem MJD brains show increased BCL2/BAX transcript ratio in the dentate cerebellar nucleus (DCN) and increased BCL2/BAX insoluble protein ratio in the DCN and pons, suggesting that in these regions, severely affected by degeneration in MJD, cells show signs of apoptosis resistance. Interestingly, a follow-up study of 18 patients further shows that blood BCL2 and TP53 transcript levels increase over time in MJD patients. Furthermore, while the similar levels of blood BCL2, BAX, and TP53 transcripts observed in preclinical subjects and controls is mimicked by pre-symptomatic MJD mice, the expression profile of these genes in patient brains is partially replicated by symptomatic MJD mice. Globally, our findings indicate that there is tissue-specific vulnerability to apoptosis in MJD subjects and that this tissue-dependent behavior is partially replicated in a MJD mouse model.

Spinal and Bulbar Muscular Atrophy (SBMA) is an X-linked adult-onset progressive neuromuscular disease that affects the spinal and bulbar motor neurons and skeletal muscles. SBMA is caused by expansion of polymorphic CAG trinucleotide repeats in the Androgen Receptor (AR) gene, resulting in expanded glutamine tract in the AR protein. Polyglutamine (polyQ) expansion renders the mutant AR protein toxic, resulting in the formation of mutant protein aggregates and cell death. This classifies SBMA as one of the nine known polyQ diseases. Like other polyQ disorders, the expansion of the polyQ tract in the AR protein is the main genetic cause of the disease; however, multiple other mechanisms besides the polyQ tract expansion also contribute to the SBMA disease pathophysiology. Posttranslational modifications (PTMs), including phosphorylation, acetylation, methylation, ubiquitination, and SUMOylation are a category of mechanisms by which the functionality of AR has been found to be significantly modulated and can alter the neurotoxicity of SBMA. This review summarizes the different PTMs and their effects in regulating the AR function and discusses their pathogenic or protective roles in context of SBMA. This review also includes the therapeutic approaches that target the PTMs of AR in an effort to reduce the mutant AR-mediated toxicity in SBMA. [ABSTRACT FROM AUTHOR]

Genome-wide association studies (GWASs) of Huntington disease (HD) have identified six DNA maintenance gene loci (among others) as modifiers and implicated a two step-mechanism of pathogenesis: somatic instability of the causative HTT CAG repeat with subsequent triggering of neuronal damage. The largest studies have been limited to HD individuals with a rater-estimated age at motor onset. To capitalize on the wealth of phenotypic data in several large HD natural history studies, we have performed algorithmic prediction by using common motor and cognitive measures to predict age at other disease landmarks as additional phenotypes for GWASs. Combined with imputation with the Trans-Omics for Precision Medicine reference panel, predictions using integrated measures provided objective landmark phenotypes with greater power to detect most modifier loci. Importantly, substantial differences in the relative modifier signal across loci, highlighted by comparing common modifiers at MSH3 and FAN1 , revealed that individual modifier effects can act preferentially in the motor or cognitive domains. Individual components of the DNA maintenance modifier mechanisms may therefore act differentially on the neuronal circuits underlying the corresponding clinical measures. In addition, we identified additional modifier effects at the PMS1 and PMS2 loci and implicated a potential second locus on chromosome 7. These findings indicate that broadened discovery and characterization of HD genetic modifiers based on additional quantitative or qualitative phenotypes offers not only the promise of in-human validated therapeutic targets but also a route to dissecting the mechanisms and cell types involved in both the somatic instability and toxicity components of HD pathogenesis. [ABSTRACT FROM AUTHOR]

Abstract Intracellular aggregates are a common pathological hallmark of neurodegenerative diseases such as polyglutamine (polyQ) diseases, amyotrophic lateral sclerosis (ALS), Parkinson’s disease (PD), and multiple system atrophy (MSA). Aggregates are mainly formed by aberrant disease-specific proteins and are accompanied by accumulation of other aggregate-interacting proteins. Although aggregate-interacting proteins have been considered to modulate the formation of aggregates and to be involved in molecular mechanisms of disease progression, the components of aggregate-interacting proteins remain unknown. In this study, we showed that small glutamine-rich tetratricopeptide repeat-containing protein alfa (SGTA) is an aggregate-interacting protein in neurodegenerative diseases. Immunohistochemistry showed that SGTA interacted with intracellular aggregates in Huntington disease (HD) cell models and neurons of HD model mice. We also revealed that SGTA colocalized with intracellular aggregates in postmortem brains of patients with polyQ diseases including spinocerebellar ataxia (SCA)1, SCA2, SCA3, and dentatorubral–pallidoluysian atrophy. In addition, SGTA colocalized with glial cytoplasmic inclusions in the brains of MSA patients, whereas no accumulation of SGTA was observed in neurons of PD and ALS patients. In vitro study showed that SGTA bound to polyQ aggregates through its C-terminal domain and SGTA overexpression reduced intracellular aggregates. These results suggest that SGTA may play a role in the formation of aggregates and may act as potential modifier of molecular pathological mechanisms of polyQ diseases and MSA.

Spinal and Bulbar Muscular Atrophy (SBMA) is an X-linked adult-onset progressive neuromuscular disease that affects the spinal and bulbar motor neurons and skeletal muscles. SBMA is caused by expansion of polymorphic CAG trinucleotide repeats in the Androgen Receptor (AR) gene, resulting in expanded glutamine tract in the AR protein. Polyglutamine (polyQ) expansion renders the mutant AR protein toxic, resulting in the formation of mutant protein aggregates and cell death. This classifies SBMA as one of the nine known polyQ diseases. Like other polyQ disorders, the expansion of the polyQ tract in the AR protein is the main genetic cause of the disease; however, multiple other mechanisms besides the polyQ tract expansion also contribute to the SBMA disease pathophysiology. Posttranslational modifications (PTMs), including phosphorylation, acetylation, methylation, ubiquitination, and SUMOylation are a category of mechanisms by which the functionality of AR has been found to be significantly modulated and can alter the neurotoxicity of SBMA. This review summarizes the different PTMs and their effects in regulating the AR function and discusses their pathogenic or protective roles in context of SBMA. This review also includes the therapeutic approaches that target the PTMs of AR in an effort to reduce the mutant AR-mediated toxicity in SBMA.

Machado-Joseph disease (MJD/SCA3) is a neurodegenerative polyglutamine disorder exhibiting a wide spectrum of phenotypes. The abnormal size of the (CAG)n at ATXN3 explains ~55% of the age at onset variance, suggesting the involvement of other factors, namely genetic modifiers, whose identification remains limited. Our aim was to find novel genetic modifiers, analyse their epistatic effects and identify disease-modifying pathways contributing to MJD variable expressivity. We performed whole-exome sequencing in a discovery sample of four age at onset concordant and four discordant first-degree relative pairs of Azorean patients, to identify candidate variants which genotypes differed for each discordant pair but were shared in each concordant pair. Variants identified by this approach were then tested in an independent multi-origin cohort of 282 MJD patients. Whole-exome sequencing identified 233 candidate variants, from which 82 variants in 53 genes were prioritized for downstream analysis. Eighteen disease-modifying pathways were identified; two of the most enriched pathways were relevant for the nervous system, namely the neuregulin signaling and the agrin interactions at neuromuscular junction. Variants at PARD3, NFKB1, CHD5, ACTG1, CFAP57, DLGAP2, ITGB1, DIDO1 and CERS4 modulate age at onset in MJD, with those identified in CFAP57, ACTG1 and DIDO1 showing consistent effects across cohorts of different geographical origins. Network analyses of the nine novel MJD modifiers highlighted several important molecular interactions, including genes/proteins previously related with MJD pathogenesis, namely between ACTG1/APOE and VCP/ITGB1. We describe novel pathways, modifiers, and their interaction partners, providing a broad molecular portrait of age at onset modulation to be further exploited as new disease-modifying targets for MJD and related diseases.

BackgroundMachado-Joseph disease (or spinocerebellar ataxia type 3) is a late-onset polyglutamine neurodegenerative disorder caused by a mutation in the ATXN3 gene, which encodes for the ubiquitously expressed protein ataxin-3. Previous studies on cell and animal models have suggested that mutated ataxin-3 is involved in transcriptional dysregulation. Starting with a whole-transcriptome profiling of peripheral blood samples from patients and controls, we aimed to confirm abnormal expression profiles in Machado-Joseph disease and to identify promising up-regulated genes as potential candidate biomarkers of disease status.MethodsThe Illumina Human V4-HT12 array was used to measure transcriptome-wide gene expression in peripheral blood samples from 12 patients and 12 controls. Technical validation and validation in an independent set of samples were performed by quantitative real-time polymerase chain reaction (PCR).ResultsBased on the results from the microarray, twenty six genes, found to be up-regulated in patients, were selected for technical validation by quantitative real-time PCR (validation rate of 81% for the up-regulation trend). Fourteen of these were further tested in an independent set of 42 patients and 35 controls; 10 genes maintained the up-regulation trend (FCGR3B, CSR2RA, CLC, TNFSF14, SLA, P2RY13, FPR2, SELPLG, YIPF6, and GPR96); FCGR3B, P2RY13, and SELPLG were significantly up-regulated in patients when compared with controls.ConclusionsOur findings support the hypothesis that mutated ataxin-3 is associated with transcription dysregulation, detectable in peripheral blood cells. Furthermore, this is the first report suggesting a pool of up-regulated genes in Machado-Joseph disease that may have the potential to be used for fine phenotyping of this disease. © 2015 International Parkinson and Movement Disorder Society.

Abstract A 61 year old man with facial diplegia, quadruparesis, tongue atrophy/fasciculations, bulbar speech, muscle weakness/wasting, hypotonia, tremor, dysdiadochokinesia, absent tendon reflexes, fasciculations, and gynecomastia, received immunoglobulins for suspected immune‐neuropathy with limited benefit. After reconsideration, Kennedy disease was diagnosed upon 44 CAG repeats in AR. In conclusion, immunoglobulins exhibit limited benefit on immune‐neuropathy in patients with coexisting KD.

Spinocerebellar ataxias (SCAs) are a group of genetically heterogeneous inherited neurodegenerative disorders characterized by progressive ataxia and cerebellar degeneration. Here, we used a mouse model to test a possible connection between SCA and Ronin (Thap11), a polyglutamine-containing transcriptional regulator encoded in a region of human chromosome 16q22.1 that has been genetically linked to SCA type 4. We report that transgenic expression of Ronin in mouse cerebellar Purkinje cells leads to detrimental loss of these cells and the development of severe ataxia as early as 10 weeks after birth. Mechanistically, we find that several SCA-causing genes harbor Ronin DNA-binding motifs and are transcriptionally deregulated in transgenic animals. In addition, ectopic expression of Ronin in embryonic stem cells significantly increases the protein level of Ataxin-1, the protein encoded by Atxn1, alterations of which cause SCA type 1. This increase is also seen in the cerebellum of transgenic animals, although the latter was not statistically significant. Hence, our data provide evidence for a link between Ronin and SCAs, and suggest that Ronin may be involved in the development of other neurodegenerative diseases.

We report an autopsy case of a 56‐year‐old male patient with the coexistence of dentatorubral‐pallidoluysian atrophy (DRPLA) and Parkinson's disease (PD). He presented with gait instability and dysarthria for 10 years. The removed brain showed general atrophy (988 g) with depigmentation of the substantia nigra. The neocortex and deep gray matter, including the red nucleus, subthalamic nuclei, and globus pallidus, were atrophic, and grumose degeneration of the cerebellar dentate nucleus was observed. Polyglutamine‐ and p62‐positive neuronal inclusions were present and widespread in the areas mentioned above. Interestingly, this case also had brainstem‐predominant PD pathology with α‐synuclein‐positive Lewy bodies and Lewy neurites. Generalized white matter atrophy with patchy loss of astrocytes in the white matter suggested glial dysfunction by elongated CAG repeats in the atrophin 1 gene (atrophin 1). Polymerase chain reaction (PCR) fragment analysis revealed increased CAG repeats (61) on atrophin 1 encoding atrophin 1. The patient had a family history of DRPLA, including his daughter, who was confirmed positive on genetic testing (CAG repeat: 65). His father, brother, and niece were suspected of having the disease. Clinicopathologically, all of the above findings are consistent with the coexistence of DRPLA and PD. So far, various overlapping neurodegenerative disorders have been reported, but the coexistence of DRPLA and PD has never been demonstrated in the published literature. Even though the exact time of PD development is unknown in this case, PD might develop after DRPLA, and the overwhelming symptoms of DRPLA might mask those of PD. Here, we report a clinicopathologically definite case of the coexistence of DRPLA and PD. White matter degeneration with patchy loss of astrocytes was another remarkable finding of this case. [ABSTRACT FROM AUTHOR]

The RNA-binding protein Ataxin-2 regulates translation and mRNA stability through cytoplasmic polyadenylation of the targets. Here we newly identified DDX6 as a positive regulator of the cytoplasmic polyadenylation. Analysis of Ataxin-2 interactome using LC-MS/MS revealed prominent interaction with the DEAD-box RNA helicase DDX6. DDX6 interacted with components of the Ataxin-2 polyadenylation machinery; Ataxin-2, PABPC1 and PAPD4. As in the case for Ataxin-2 downregulation, DDX6 downregulation led to an increase in Ataxin-2 target mRNAs with short poly(A) tails as well as a reduction in their protein expression. In contrast, Ataxin-2 target mRNAs with short poly(A) tails were decreased by the overexpression of Ataxin-2, which was compromised by the DDX6 downregulation. However, polyadenylation induced by Ataxin-2 tethering was not affected by the DDX6 downregulation. Taken together, these results suggest that DDX6 positively regulates Ataxin-2-induced cytoplasmic polyadenylation to maintain poly(A) tail length of the Ataxin-2 targets provably through accelerating binding of Ataxin-2 to the target mRNAs. • Ataxin-2 interactome was analyzed by LC-MS/MS. • DDX6 is a component of the Ataxin-2/PAPD4 polyadenylation machinery. • DDX6 is required for maintaining poly(A) tail length of the Ataxin-2 target mRNAs. • DDX6 is required for protein expression of the Ataxin-2 target mRNAs. • DDX6 positively regulates Ataxin-2-induced cytoplasmic polyadenylation. [ABSTRACT FROM AUTHOR]

Intracellular aggregates are a common pathological hallmark of neurodegenerative diseases such as polyglutamine (polyQ) diseases, amyotrophic lateral sclerosis (ALS), Parkinson's disease (PD), and multiple system atrophy (MSA). Aggregates are mainly formed by aberrant disease-specific proteins and are accompanied by accumulation of other aggregate-interacting proteins. Although aggregate-interacting proteins have been considered to modulate the formation of aggregates and to be involved in molecular mechanisms of disease progression, the components of aggregate-interacting proteins remain unknown. In this study, we showed that small glutamine-rich tetratricopeptide repeat-containing protein alfa (SGTA) is an aggregate-interacting protein in neurodegenerative diseases. Immunohistochemistry showed that SGTA interacted with intracellular aggregates in Huntington disease (HD) cell models and neurons of HD model mice. We also revealed that SGTA colocalized with intracellular aggregates in postmortem brains of patients with polyQ diseases including spinocerebellar ataxia (SCA)1, SCA2, SCA3, and dentatorubral–pallidoluysian atrophy. In addition, SGTA colocalized with glial cytoplasmic inclusions in the brains of MSA patients, whereas no accumulation of SGTA was observed in neurons of PD and ALS patients. In vitro study showed that SGTA bound to polyQ aggregates through its C-terminal domain and SGTA overexpression reduced intracellular aggregates. These results suggest that SGTA may play a role in the formation of aggregates and may act as potential modifier of molecular pathological mechanisms of polyQ diseases and MSA. [ABSTRACT FROM AUTHOR]

Spinal and bulbar muscular atrophy (SBMA) is a neuromuscular degenerative disease caused by a polyglutamine expansion in the androgen receptor (AR). This mutation causes AR to misfold and aggregate, contributing to toxicity in and degeneration of motor neurons and skeletal muscle. There is currently no effective treatment or cure for this disease. The role of an interdomain interaction between the amino- and carboxyl-termini of AR, termed the N/C interaction, has been previously identified as a component of androgen receptor-induced toxicity in cell and mouse models of SBMA. However, the mechanism by which this interaction contributes to disease pathology is unclear. This work seeks to investigate this mechanism by interrogating the role of AR homodimerization- a unique form of the N/C-interaction- in SBMA. We show that, although the AR N/C-interaction is reduced by polyglutamine-expansion, homodimers of 5α-dihydrotestosterone (DHT)-bound AR are increased. Additionally, blocking homodimerization results in decreased AR aggregation and toxicity in cell models. Blocking homodimerization results in the increased degradation of AR, which likely plays a role in the protective effects of this mutation. Overall, this work identifies a novel mechanism in SBMA pathology that may represent a novel target for the development of therapeutics for this disease., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Huntington's disease (HD) is a progressive neurodegenerative disorder caused by a trinucleotide repeat expansion in the huntingtin (HTT) gene, which is expressed ubiquitously throughout the brain and peripheral tissues. Whilst the focus of much research has been on the cognitive, psychiatric and motor symptoms of HD, the extent of peripheral pathology and its potential impact on central symptoms has been less intensely explored. Disruption of the gastrointestinal microbiome (gut dysbiosis) has been recently reported in a number of neurological and psychiatric disorders, and therefore we hypothesized that it might also occur in HD. We have used 16S rRNA amplicon sequencing to characterize the gut microbiome in the R6/1 transgenic mouse model of HD, relative to littermate wild-type controls. We report that there is a significant difference in microbiota composition in HD mice at 12 weeks of age. Specifically, we observed an increase in Bacteriodetes and a proportional decrease in Firmicutes in the HD gut microbiome. In addition, we observed an increase in microbial diversity in male HD mice, compared to wild-type controls, but no differences in diversity were observed in female HD mice. The gut dysbiosis observed coincided with impairment in body weight gain despite higher food intake as well as motor deficits at 12 weeks of age. Gut dysbiosis was also associated with a change in the gut microenvironment, as we observed higher fecal water content in HD mice at 12 weeks of age. This study provides the first evidence of gut dysbiosis in HD.

Polyglutamine (polyQ) diseases, including Huntington’s disease, are a group of late-onset progressive neurological disorders caused by CAG repeat expansions. Although recently, many studies have investigated the pathological features and development of polyQ diseases, many questions remain unanswered. The advancement of new gene-editing technologies, especially the CRISPR-Cas9 technique, has undeniable value for the generation of relevant polyQ models, which substantially support the research process. Here, we review how these tools have been used to correct disease-causing mutations or create isogenic cell lines with different numbers of CAG repeats. We characterize various cellular models such as HEK 293 cells, patient-derived fibroblasts, human embryonic stem cells (hESCs), induced pluripotent stem cells (iPSCs) and animal models generated with the use of genome-editing technology.

Key points: Muscle‐derived neurotrophic factors may offer therapeutic promise for treating neuromuscular diseases.We report that a muscle‐derived neurotrophic factor, BDNF, rescues synaptic and muscle function in a muscle‐type specific manner in mice modelling Kennedy's disease (KD).We also find that BDNF rescues select molecular mechanisms in slow and fast muscle that may underlie the improved cellular function.We also report for the first time that expression of BDNF, but not other members of the neurotrophin family, is perturbed in muscle from patients with KD.Given that muscle BDNF had divergent therapeutic effects that depended on muscle type, a combination of neurotrophic factors may optimally rescue neuromuscular function via effects on both pre‐ and postsynaptic function, in the face of disease. Deficits in muscle brain‐derived neurotrophic factor (BDNF) correlate with neuromuscular deficits in mouse models of Kennedy's disease (KD), suggesting that restoring muscle BDNF might restore function. To test this possibility, transgenic mice expressing human BDNF in skeletal muscle were crossed with '97Q' KD mice. We found that muscle BDNF slowed disease, doubling the time between symptom onset and endstage. BDNF also improved expression of genes in muscle known to play key roles in neuromuscular function, including counteracting the expression of neonatal isoforms induced by disease. Intriguingly, BDNF's ameliorative effects differed between muscle types: synaptic strength was rescued only in slow‐twitch muscle, while contractile strength was improved only in fast‐twitch muscle. In sum, muscle BDNF slows disease progression, rescuing select cellular and molecular mechanisms that depend on fibre type. Muscle BDNF expression was also affected in KD patients, reinforcing its translational and therapeutic potential for treating this disorder. Key points: Muscle‐derived neurotrophic factors may offer therapeutic promise for treating neuromuscular diseases.We report that a muscle‐derived neurotrophic factor, BDNF, rescues synaptic and muscle function in a muscle‐type specific manner in mice modelling Kennedy's disease (KD).We also find that BDNF rescues select molecular mechanisms in slow and fast muscle that may underlie the improved cellular function.We also report for the first time that expression of BDNF, but not other members of the neurotrophin family, is perturbed in muscle from patients with KD.Given that muscle BDNF had divergent therapeutic effects that depended on muscle type, a combination of neurotrophic factors may optimally rescue neuromuscular function via effects on both pre‐ and postsynaptic function, in the face of disease. [ABSTRACT FROM AUTHOR]

Spinocerebellar ataxia type 2 (SCA2), a rare polyglutamine neurodegenerative disorder caused by a CAG repeat expansion in the ataxin-2 gene, exhibits common cellular phenotypes with other neurodegenerative disorders, including oxidative stress and mitochondrial dysfunction. Here, we show that SCA2 patient cells exhibit higher levels of caspase-8- and caspase-9-mediated apoptotic activation than control cells, cellular phenotypes that we find to be exacerbated by reactive oxygen species (ROS) and inhibition of autophagy. We also suggest that oligomerization of mutant ataxin-2 protein is likely to be the cause of the observed cellular phenotypes by causing inhibition of autophagy and by inducing ROS generation. Finally, we show that removal of ataxin-2 oligomers, either by increasing autophagic clearance or by oligomer dissolution, appears to alleviate the cellular phenotypes. Our results suggest that oligomerized ataxin-2 and oxidative stress affect autophagic clearance in SCA2 cells, contributing to the pathophysiology, and that activation of autophagy or clearance of oligomers may prove to be effective therapeutic strategies. [ABSTRACT FROM AUTHOR]

The most common dominantly inherited ataxia, spinocerebellar ataxia type 3 (SCA3), is an incurable neurodegenerative disorder caused by a CAG repeat expansion in the ATXN3 gene that encodes an abnormally long polyglutamine tract in the disease protein, ATXN3. Mice lacking ATXN3 are phenotypically normal; hence, disease gene suppression offers a compelling approach to slow the neurodegenerative cascade in SCA3. Here we tested antisense oligonucleotides (ASOs) that target human ATXN3 in two complementary mouse models of SCA3: yeast artificial chromosome (YAC) MJD-Q84.2 (Q84) mice expressing the full-length human ATXN3 gene and cytomegalovirus (CMV) MJD-Q135 (Q135) mice expressing a human ATXN3 cDNA. Intracerebroventricular injection of ASOs resulted in widespread delivery to the most vulnerable brain regions in SCA3. In treated Q84 mice, three of five tested ASOs reduced disease protein levels by >50% in the diencephalon, cerebellum, and cervical spinal cord. Two ASOs also significantly reduced mutant ATXN3 in the mouse forebrain and resulted in no signs of astrogliosis or microgliosis. In Q135 mice expressing a single ATXN3 isoform via a cDNA transgene, ASOs did not result in similar robust ATXN3 silencing. Our results indicate that ASOs targeting full-length human ATXN3 would likely be well tolerated and could lead to a preventative therapy for SCA3.

Huntington's Disease (HD) is a fatal neurodegenerative disease characterized by severe loss of medium spiny neuron (MSN) function and striatal-dependent behaviors. We report that female HdhQ200/200 mice display an earlier onset and more robust deterioration in spontaneous locomotion and motor coordination measured at 8 months of age compared to male HdhQ200/200 mice. Remarkably, HdhQ200/200 mice of both sexes exhibit comparable impaired spontaneous locomotion and motor coordination at 10 months of age and reach moribund stage by 12 months of age, demonstrating reduced life span in this model system. Histopathological analysis revealed enhanced mutant huntingtin protein aggregation in male HdhQ200/200 striatal tissue at 8 months of age compared to female HdhQ200/200. Functional analysis of calcium dynamics in MSNs of female HdhQ200/200 mice using GCaMP6m imaging revealed elevated responses to excitatory cortical-striatal stimulation suggesting increased MSN excitability. Although there was no down-regulation of the expression of common HD biomarkers (DARPP-32, enkephalin and CB1R), we measured a sex-dependent reduction of the astrocytic glutamate transporter, GLT-1, in female HdhQ200/200 mice that was not detected in male HdhQ200/200 mice when compared to respective wild-type littermates. Our study outlines a sex-dependent rapid deterioration of striatal-dependent behaviors occurring in the HdhQ200/200 mouse line that does not involve alterations in the expression of common HD biomarkers and yet includes impaired MSN function.

Perturbation of protein homeostasis and aggregation of misfolded proteins is a major cause of many human diseases. A hallmark of the neurodegenerative disease spinocerebellar ataxia type 7 (SCA7) is the intranuclear accumulation of mutant, misfolded ataxin-7 (polyQ-ATXN7). Here, we show that endogenous ATXN7 is modified by SUMO proteins, thus also suggesting a physiological role for this modification under conditions of proteotoxic stress caused by the accumulation of polyQ-ATXN7. Co-immunoprecipitation experiments, immunofluorescence microscopy and proximity ligation assays confirmed the colocalization and interaction of polyQ-ATXN7 with SUMO2 in cells. Moreover, upon inhibition of the proteasome, both endogenous SUMO2/3 and the RNF4 ubiquitin ligase surround large polyQ-ATXN7 intranuclear inclusions. Overexpression of RNF4 and/or SUMO2 significantly decreased levels of polyQ-ATXN7 and, upon proteasomal inhibition, led to a marked increase in the polyubiquitination of polyQ-ATXN7. This provides a mechanism for the clearance of polyQ-ATXN7 from affected cells that involves the recruitment of RNF4 by SUMO2/3-modified polyQ-ATXN7, thus leading to its ubiquitination and proteasomal degradation. In a SCA7 knock-in mouse model, we similarly observed colocalization of SUMO2/3 with polyQ-ATXN7 inclusions in the cerebellum and retina. Furthermore, we detected accumulation of SUMO2/3 high-molecular-mass species in the cerebellum of SCA7 knock-in mice, compared with their wild-type littermates, and changes in SUMO-related transcripts. Immunohistochemical analysis showed the accumulation of SUMO proteins and RNF4 in the cerebellum of SCA7 patients. Taken together, our results show that the SUMO pathway contributes to the clearance of aggregated ATXN7 and suggest that its deregulation might be associated with SCA7 disease progression.

Background: Spinal and bulbar muscular atrophy (SBMA) is a hereditary neuromuscular disorder caused by the expansion of trinucleotide cytosine-adenine-guanine (CAG) repeats, which encodes a polyglutamine (polyQ) tract in the androgen receptor (AR) gene. Recent evidence suggests that, in addition to motor neuron degeneration, defective skeletal muscles are also the primary contributors to the pathogenesis in SBMA. While benefits of physical exercise have been suggested in SBMA, underlying mechanism remains elusive., Methods: We investigated the effect of running exercise in a transgenic mouse model of SBMA carrying human AR with 97 expanded CAGs (AR97Q). We assigned AR97Q mice to exercise and sedentary control groups, and mice in the exercise group received 1-h forced running wheel (5 m/min) 5 days a week for 4 weeks during the early stage of the disease. Motor function (grip strength and rotarod performance) and survival of each group were analysed, and histopathological and biological features in skeletal muscles and motor neurons were evaluated., Results: AR97Q mice in the exercise group showed improvement in motor function (~40% and ~50% increase in grip strength and rotarod performance, respectively, P < 0.05) and survival (median survival 23.6 vs. 16.7 weeks, P < 0.05) with amelioration of neuronal and muscular histopathology (~1.4-fold and ~2.8-fold increase in motor neuron and muscle fibre size, respectively, P < 0.001) compared to those in the sedentary group. Nuclear accumulation of polyQ-expanded AR in skeletal muscles and motor neurons was suppressed in the mice with exercise compared to the sedentary mice (~50% and ~30% reduction in 1C2-positive cells in skeletal muscles and motor neurons, respectively, P < 0.05). We found that the exercise activated 5'-adenosine monophosphate-activated protein kinase (AMPK) signalling and inhibited mammalian target of rapamycin pathway that regulates protein synthesis in skeletal muscles of SBMA mice. Pharmacological activation of AMPK inhibited protein synthesis and reduced polyQ-expanded AR proteins in C2C12 muscle cells., Conclusions: Our findings suggest the therapeutic potential of exercise-induced effect via AMPK activation in SBMA., (© 2023 The Authors. Journal of Cachexia, Sarcopenia and Muscle published by Wiley Periodicals LLC.)

Expansions of glutamine-coding CAG trinucleotide repeats cause a number of neurodegenerative diseases, including Huntington's disease and several of spinocerebellar ataxias. In general, age-at-onset of the polyglutamine diseases is inversely correlated with the size of the respective inherited expanded CAG repeat. Expanded CAG repeats are also somatically unstable in certain tissues, and age-at-onset of Huntington's disease corrected for individual HTT CAG repeat length (i.e. residual age-at-onset), is modified by repeat instability-related DNA maintenance/repair genes as demonstrated by recent genome-wide association studies. Modification of one polyglutamine disease (e.g. Huntington's disease) by the repeat length of another (e.g. ATXN3, CAG expansions in which cause spinocerebellar ataxia 3) has also been hypothesized. Consequently, we determined whether age-at-onset in Huntington's disease is modified by the CAG repeats of other polyglutamine disease genes. We found that the CAG measured repeat sizes of other polyglutamine disease genes that were polymorphic in Huntington's disease participants but did not influence Huntington's disease age-at-onset. Additional analysis focusing specifically on ATXN3 in a larger sample set ( n = 1388) confirmed the lack of association between Huntington's disease residual age-at-onset and ATXN3 CAG repeat length. Additionally, neither our Huntington's disease onset modifier genome-wide association studies single nucleotide polymorphism data nor imputed short tandem repeat data supported the involvement of other polyglutamine disease genes in modifying Huntington's disease. By contrast, our genome-wide association studies based on imputed short tandem repeats revealed significant modification signals for other genomic regions. Together, our short tandem repeat genome-wide association studies show that modification of Huntington's disease is associated with short tandem repeats that do not involve other polyglutamine disease-causing genes, refining the landscape of Huntington's disease modification and highlighting the importance of rigorous data analysis, especially in genetic studies testing candidate modifiers., Competing Interests: J.F.G. was a Scientific Advisory Board member and had a financial interest in Triplet Therapeutics, Inc. His NIH-funded project is using genetic and genomic approaches to uncover other genes that significantly influence when diagnosable symptoms emerge and how rapidly they worsen in Huntington disease. The company is developing new therapeutic approaches to address triplet repeat disorders such Huntington's disease, myotonic dystrophy and SCAs. His interests were reviewed and are managed by Massachusetts General Hospital and Mass General Brigham in accordance with their conflict of interest policies. J.F.G. has also been a consultant for Wave Life Sciences USA, Inc., Biogen, Inc. and Pfizer, Inc. Within the last five years D.G.M. has been a scientific consultant and/or received an honoraria/stock options from AMO Pharma, Dyne, F. Hoffman-La Roche, LoQus23, Novartis, Ono Pharmaceuticals, Rgenta Therapeutics, Sanofi, Sarepta Therapeutics Inc, Script Biosciences, Triplet Therapeutics, and Vertex Pharmaceuticals and held research contracts with AMO Pharma and Vertex Pharmaceuticals. J.D.L. is a paid Advisory Board member for F. Hoffmann-La Roche Ltd and uniQure biopharma B.V., and he is a paid consultant for Vaccinex Inc, Wave Life Sciences USA Inc, Genentech Inc, Triplet Inc, and PTC Therapeutics Inc. T.H.M. is an associate member of the scientific advisory board of LoQus23 Therapeutics. L.J. was a member of the scientific advisory boards of LoQus23 Therapeutics and Triplet Therapeutics. V.C.W. was a Scientific Advisory Board member of Triplet Therapeutics, Inc., a company developing new therapeutic approaches to address triplet repeat disorders such Huntington's disease and myotonic dystrophy. Her financial interests in Triplet Therapeutics were reviewed and are managed by Massachusetts General Hospital and Mass General Brigham in accordance with their conflict of interest policies. She is a scientific advisory board member of LoQus23 Therapeutics and has provided paid consulting services to Alnylam, Acadia Pharmaceuticals Inc., Alnylam Inc., Biogen Inc. and Passage Bio. J.M.L consults for Life Edit Therapeutics and serves in the scientific advisory board of GenEdit, Inc., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Guarantors of Brain.)

Abstract Huntington's disease (HD) is caused by an expanded CAG repeat in the huntingtin (HTT) gene, translating into an elongated polyglutamine stretch. In addition to the neurotoxic mutant HTT protein, the mutant CAG repeat RNA can exert toxic functions by trapping RNA-binding proteins. While few examples of proteins that aberrantly bind to mutant HTT RNA and execute abnormal function in conjunction with the CAG repeat RNA have been described, an unbiased approach to identify the interactome of mutant HTT RNA is missing. Here, we describe the analysis of proteins that preferentially bind mutant HTT RNA using a mass spectrometry approach. We show that (I) the majority of proteins captured by mutant HTT RNA belong to the spliceosome pathway, (II) expression of mutant CAG repeat RNA induces mis-splicing in a HD cell model, (III) overexpression of one of the splice factors trapped by mutant HTT ameliorates the HD phenotype in a fly model and (VI) deregulated splicing occurs in human HD brain. Our data suggest that deregulated splicing is a prominent mechanism of RNA-induced toxicity in HD. Graphical Abstract Unlabelled Image Highlights • In HD, both mutant HTT protein and mutant HTT RNA can exert toxic functions. • RNA-mediated toxicity results from aberrant binding of proteins to the mutant transcript. • We describe the systematic analysis of proteins sequestered by mutant HTT RNA. • The majority of proteins captured by mutant HTT RNA belong to the spliceosome pathway. • Deregulated splicing is a prominent mechanism of RNA-induced toxicity in HD. [ABSTRACT FROM AUTHOR]

Abstract More than 90% of the human genome are transcribed as non-coding RNAs. While it is still under debate if all these non-coding transcripts are functional, there is emerging evidence that RNA has several important functions in addition to coding for proteins. For example, microRNAs (miRNAs) are important regulatory RNAs that control gene expression in various biological processes and human diseases. In spinocerebellar ataxia type 3 (SCA3), a devastating neurodegenerative disease, miRNAs are involved in the disease process at different levels, including the deregulation of components of the general miRNA biogenesis machinery, as well as in the cell type-specific control of the expression of the SCA3 disease protein and other SCA3 disease-relevant proteins. However, it remains difficult to predict whether these changes are a cause or a consequence of the neurodegenerative process in SCA3. Further studies using standardized procedures for the analysis of miRNA expression and larger sample numbers are required to enhance our understanding of the miRNA-mediated processes involved in SCA3 disease and may enable the development of miRNA-based therapeutics. In this review, we summarize the findings of independent studies highlighting both the disease-related and cytoprotective roles of miRNAs that have been implicated so far in the disease process of SCA3. Graphical Abstract Unlabelled Image Highlights • The majority of the human transcriptome is made up by non-coding RNAs, although it is unknown if all these transcripts are functional. • Among functional non-coding RNAs, miRNAs play key roles not only during neuronal maturation and brain development but also in neurodegeneration. • In SCA3, miRNAs are involved in the disease process at different levels, including deregulation of the general miRNA biosynthesis machinery, expression of the aggregation-prone disease protein, ATXN3, and other disease process-related proteins like molecular chaperones. • Cell type-specific miRNA changes identified in different cell models of SCA3 may reflect the selective vulnerability of specific cell types affected in SCA3. • Induction of miRNAs that specifically suppress the expression of ATXN3 presents one promising putative therapeutic approach. [ABSTRACT FROM AUTHOR]

Spinal and bulbar muscular atrophy is a neurodegenerative disease caused by extended CAG trinucleotide repeats in the androgen receptor gene, which encodes a ligand-dependent transcription factor. The mutant androgen receptor protein, characterized by polyglutamine expansion, is prone to misfolding and forms aggregates in both the nucleus and cytoplasm in the brain in spinal and bulbar muscular atrophy patients. These aggregates alter protein–protein interactions and compromise transcriptional activity. In this study, we reported that in both cultured N2a cells and mouse brain, mutant androgen receptor with polyglutamine expansion causes reduced expression of mesencephalic astrocyte-derived neurotrophic factor. Overexpression of mesencephalic astrocyte-derived neurotrophic factor ameliorated the neurotoxicity of mutant androgen receptor through the inhibition of mutant androgen receptor aggregation. Conversely, knocking down endogenous mesencephalic astrocyte-derived neurotrophic factor in the mouse brain exacerbated neuronal damage and mutant androgen receptor aggregation. Our findings suggest that inhibition of mesencephalic astrocyte-derived neurotrophic factor expression by mutant androgen receptor is a potential mechanism underlying neurodegeneration in spinal and bulbar muscular atrophy.

Polyglutamine (polyQ) diseases represent a group of progressive neurodegenerative disorders that are caused by abnormal expansion of CAG triplet nucleotides in disease genes. Recent evidence indicates that not only mutant polyQ proteins, but also their corresponding mutant RNAs, contribute to the pathogenesis of polyQ diseases. Here, we describe the identification of a 13-amino-acid peptide, P3, which binds directly and preferentially to long-CAG RNA within the pathogenic range. When administered to cell and Drosophila disease models, as well as to patient-derived fibroblasts, P3 inhibited expanded-CAG-RNA-induced nucleolar stress and suppressed neurotoxicity. We further examined the combined therapeutic effect of P3 and polyQ-binding peptide 1 (QBP1), a well-characterized polyQ protein toxicity inhibitor, on neurodegeneration. When P3 and QBP1 were co-administered to disease models, both RNA and protein toxicities were effectively mitigated, resulting in a notable improvement of neurotoxicity suppression compared with the P3 and QBP1 single-treatment controls. Our findings indicate that targeting toxic RNAs and/or simultaneous targeting of toxic RNAs and their corresponding proteins could open up a new therapeutic strategy for treating polyQ degeneration.

Spinocerebellar ataxia type 7 (SCA7), a neurodegenerative disease characterized by cerebellar ataxia and retinal degeneration, is caused by an abnormal CAG repeat expansion in the ATXN7 gene coding region. The onset and severity of SCA7 are highly variable between patients, thus identification of sensitive biomarkers that accurately diagnose the disease and monitoring its progression are needed. With the aim of identified SCA7-specific metabolites with clinical relevance, we report for the first time, to the best of our knowledge, a metabolomics profiling of circulating acylcarnitines and amino acids in SCA7 patients. We identified 21 metabolites with altered levels in SCA7 patients and determined two different sets of metabolites with diagnostic power. The first signature of metabolites (Valine, Leucine, and Tyrosine) has the ability to discriminate between SCA7 patients and healthy controls, while the second one (Methionine, 3-hydroxytetradecanoyl-carnitine, and 3-hydroxyoctadecanoyl-carnitine) possess the capability to differentiate between early-onset and adult-onset patients, as shown by the multivariate model and ROC analyses. Furthermore, enrichment analyses of metabolic pathways suggest alterations in mitochondrial function, energy metabolism, and fatty acid beta-oxidation in SCA7 patients. In summary, circulating SCA7-specific metabolites identified in this study could serve as effective predictors of SCA7 progression in the clinics, as they are sampled in accessible biofluid and assessed by a relatively simple biochemical assay.

Huntington’s disease (HD) is a neurodegenerative disorder caused by a tandem repeat mutation encoding an expanded polyglutamine tract in the huntingtin protein, which leads to cognitive, psychiatric and motor dysfunction. Exposure to environmental enrichment (EE), which enhances levels of cognitive stimulation and physical activity, has therapeutic effects on cognitive, affective and motor function of transgenic HD mice. The present study investigated gene expression changes and behavioral pharmacology in male and female R6/1 transgenic HD mice at an early time-point in HD progression associated with onset of cognitive and affective abnormalities, following EE and exercise (wheel running) interventions. We have demonstrated changes in expression levels of the serotonin (5-HT) receptor Htr1a, Htr1b, Htr2a and Htr2c genes (encoding the 5-HT1A, 5-HT1B, 5-HT2A and 5-HT2C receptors, respectively) in HD brains at 8 weeks of age, using quantitative real-time PCR. In contrast, expression of the serotonin transporter (SerT, also known as 5-HTT or Slc6a4) was not altered in these brains. Furthermore, we identified region-specific, sex-specific and environmentally regulated (comparing EE, exercise and standard housing conditions) impacts on gene expression of particular 5-HT receptors, as well as SerT. For example, SerT gene expression was upregulated by exercise (wheel running from 6 to 8 weeks of age) in the hippocampus. Interestingly, when EE was introduced from 6 to 8 weeks of age, Htr2a gene expression was upregulated in the cortex, striatum and hippocampus of male mice. EE also rescued the functional activity of 5-HT2 receptors as observed in the head-twitch test, reflecting sexually dimorphic effects of environmental stimulation. These findings demonstrate that disruption of the serotonergic system occurs early in HD pathogenesis and, together with previous findings, show that the timing and duration of environmental interventions are critical in terms of their ability to modify gene expression. This study is the first to show that EE is able to selectively enhance both gene expression of a neurotransmitter receptor and the functional consequences on behavioral pharmacology, and links this molecular modulation to the therapeutic effects of environmental stimulation in this neurodegenerative disease.

A 61 year old man with facial diplegia, quadruparesis, tongue atrophy/fasciculations, bulbar speech, muscle weakness/wasting, hypotonia, tremor, dysdiadochokinesia, absent tendon reflexes, fasciculations, and gynecomastia, received immunoglobulins for suspected immune‐neuropathy with limited benefit. After reconsideration, Kennedy disease was diagnosed upon 44 CAG repeats in AR. In conclusion, immunoglobulins exhibit limited benefit on immune‐neuropathy in patients with coexisting KD. [ABSTRACT FROM AUTHOR]

Polyglutamine (polyQ) diseases are rare autosomal-dominant neurodegenerative diseases associated with the expansion of glutamine-encoding triplet repeats in certain genes. To investigate the functional influence of repeat expansion on disease mechanisms, we applied a biallelic genome-engineering platform that we recently established, called Universal Knock-in System or UKiS, to develop a human cell trio, a set of three isogenic cell lines that are homozygous for two different numbers of repeats (first and second lines) or heterozygous for the two repeat numbers (third line). As an example of a polyQ disease, we chose spinocerebellar ataxia type 2 (SCA2). In a pseudodiploid human cell line, both alleles of the glutamine-encoding triplet repeat in the SCA2-causing gene, ataxin 2 or ATXN2, were first knocked in with a donor sequence encoding both thymidine kinase and either puromycin or blasticidin resistance proteins under dual drug selection. The knocked-in donor alleles were then substituted with a payload having either 22 or 76 triplet repeats in ATXN2 by ganciclovir negative selection. The two-step substitution and subsequent SNP typing and genomic sequencing confirmed that the SCA2-modeling isogenic cell trio was obtained: three clones of 22-repeat homozygotes, two clones of 22/76-repeat heterozygotes and two clones of 76-repeat homozygotes. Finally, RT-PCR and immunoblotting using the obtained clones showed that, consistent with previous observations, glutamine tract expansion reduced transcriptional and translational expression of ATXN2. The cell clones with homozygous long-repeat alleles, which are rarely obtained from patients with SCA2, showed more drastic reduction of ATXN2 expression than the heterozygous clones. This study thus demonstrates the potential of UKiS, which is a beneficial platform for the efficient development of cell models not only for polyQ diseases but also for any other genetic diseases, which may accelerate our deeper understanding of disease mechanisms and cell-based screening for therapeutic drugs.

Inclusion body formation is associated with cytotoxicity in a number of neurodegenerative diseases. However, the molecular basis of the toxicity caused by the accumulation of aggregation-prone proteins remains controversial. In this study, we found that disease-associated inclusions induced by elongated polyglutamine chains disrupt the complex formation of BAG6 with UBL4A, a mammalian homologue of yeast Get5. UBL4A also dissociated from BAG6 in response to proteotoxic stresses such as proteasomal inhibition and mitochondrial depolarization. These findings imply that the cytotoxicity of pathological protein aggregates might be attributed in part to disruption of the BAG6-UBL4A complex that is required for the biogenesis of tail-anchored proteins., (© 2023 The Author(s).)