Polyglutamine Stretch Research Articles

Polymorphic CAG and GGN repeats in Cryptorchidism patient risk: A meta-analytical study

Genetic mutations in the androgen receptor (AR) gene have been identified as the cause of androgen insensitivity syndrome. These mutations are linked to inconsistent development of the Wolffian duct and may result in conditions such as micropenis, hypospadias, and cryptorchidism. The androgen receptor has two polymorphic sites located in exon 1, which consists of varying amounts of CAG and GGN repeats. These repetitions lead to the formation of polyglutamine and polyglycine stretches of varied lengths. Increased CAG repeats lead to a decrease in androgen receptor transcriptional activity, but the impact of GGN triplets is less well understood. This research examined the CAG and GGN repeat lengths in males who had a past medical record of cryptorchidism. Prospective and retrospective observational studies from PubMed, Science Direct, and Embase were systematically searched up to 15th November 2020. Primary outcomes were analyzed using a fixed or random effect model regarding its heterogeneity and continued with multilevel modeling of each polymorphism and ethnicity. CAG and GGN repeat polymorphism was found to be significantly different compared to control in contributing to cryptorchidism (CAG: 0.55 [CI 95%=0.19-0.91]; p-value=0.003 and GGN 0.90 [CI95%=0.65-1.15]; p value<0.000). In conclusion, CAG and GGN repeat polymorphism have an essential role in the incidence of cryptorchidism.

Differential Effects of Post-translational Modifications on the Membrane Interaction of Huntingtin Protein.

Huntington's disease is a neurodegenerative disorder caused by an expanded polyglutamine stretch near the N-terminus of the huntingtin (HTT) protein, rendering the protein more prone to aggregate. The first 17 residues in HTT (Nt17) interact with lipid membranes and harbor multiple post-translational modifications (PTMs) that can modulate HTT conformation and aggregation. In this study, we used a combination of biophysical studies and molecular simulations to investigate the effect of PTMs on the helicity of Nt17 in the presence of various lipid membranes. We demonstrate that anionic lipids such as PI4P, PI(4,5)P2, and GM1 significantly enhance the helical structure of unmodified Nt17. This effect is attenuated by single acetylation events at K6, K9, or K15, whereas tri-acetylation at these sites abolishes Nt17-membrane interaction. Similarly, single phosphorylation at S13 and S16 decreased but did not abolish the POPG and PIP2-induced helicity, while dual phosphorylation at these sites markedly diminished Nt17 helicity, regardless of lipid composition. The helicity of Nt17 with phosphorylation at T3 is insensitive to the membrane environment. Oxidation at M8 variably affects membrane-induced helicity, highlighting a lipid-dependent modulation of the Nt17 structure. Altogether, our findings reveal differential effects of PTMs and crosstalks between PTMs on membrane interaction and conformation of HTT. Intriguingly, the effects of phosphorylation at T3 or single acetylation at K6, K9, and K15 on Nt17 conformation in the presence of certain membranes do not mirror that observed in the absence of membranes. Our studies provide novel insights into the complex relationship between Nt17 structure, PTMs, and membrane binding.

Spinocerebellar Ataxia Type 3 Pathophysiology-Implications for Translational Research and Clinical Studies.

The spinocerebellar ataxias (SCA) comprise a group of inherited neurodegenerative diseases. Machado-Joseph Disease (MJD) or spinocerebellar ataxia 3 (SCA3) is the most common autosomal dominant form, caused by the expansion of CAG repeats within the ataxin-3 (ATXN3) gene. This mutation results in the expression of an abnormal protein containing long polyglutamine (polyQ) stretches that confers a toxic gain of function and leads to misfolding and aggregation of ATXN3 in neurons. As a result of the neurodegenerative process, SCA3 patients are severely disabled and die prematurely. Several screening approaches, e.g., druggable genome-wide and drug library screenings have been performed, focussing on the reduction in stably overexpressed ATXN3(polyQ) protein and improvement in the resultant toxicity. Transgenic overexpression models of toxic ATXN3, however, missed potential modulators of endogenous ATXN3 regulation. In another approach to identify modifiers of endogenous ATXN3 expression using a CRISPR/Cas9-modified SK-N-SH wild-type cell line with a GFP-T2A-luciferase (LUC) cassette under the control of the endogenous ATXN3 promotor, four statins were identified as potential activators of expression. We here provide an overview of the high throughput screening approaches yet performed to find compounds or genomic modifiers of ATXN3(polyQ) toxicity in different SCA3 model organisms and cell lines to ameliorate and halt SCA3 progression in patients. Furthermore, the putative role of cholesterol in neurodegenerative diseases (NDDs) in general and SCA3 in particular is discussed.

Huntington's Disease: Complex Pathogenesis and Therapeutic Strategies.

Huntington's disease (HD) arises from the abnormal expansion of CAG repeats in the huntingtin gene (HTT), resulting in the production of the mutant huntingtin protein (mHTT) with a polyglutamine stretch in its N-terminus. The pathogenic mechanisms underlying HD are complex and not yet fully elucidated. However, mHTT forms aggregates and accumulates abnormally in neuronal nuclei and processes, leading to disruptions in multiple cellular functions. Although there is currently no effective curative treatment for HD, significant progress has been made in developing various therapeutic strategies to treat HD. In addition to drugs targeting the neuronal toxicity of mHTT, gene therapy approaches that aim to reduce the expression of the mutant HTT gene hold great promise for effective HD therapy. This review provides an overview of current HD treatments, discusses different therapeutic strategies, and aims to facilitate future therapeutic advancements in the field.

A short peptide protects from age-onset proteotoxicity.

Aberrant protein aggregation jeopardizes cellular functionality and underlies the development of a myriad of late-onset maladies including Alzheimer's disease (AD) and Huntington's disease (HD). Accordingly, molecules that mitigate the toxicity of hazardous protein aggregates are of great interest as potential future therapeutics. Here we asked whether a small peptide, composed of five amino acids (5MER peptide) that was derived from the human pro-inflammatory CD44 protein, could protect model nematodes from the toxicity of aggregative proteins that underlie the development of neurodegenerative disorders in humans. We found that the 5MER peptide mitigates the toxicity that stems from both; the AD-causing Aβ peptide and a stretch of poly-glutamine that is accountable for the development of several disorders including HD, while minimally affecting lifespan. This protection was dependent on the activity of aging-regulating transcription factors and associated with enhanced Aβ and polyQ35-YFP aggregation. A transcriptomic analysis unveiled that the peptide modifies signaling pathways, thereby modulating the expression of various genes, including these, which are known as protein homeostasis (proteostasis) regulators such as txt-13 and modifiers of proteasome activity. The knockdown of txt-13 protects worms from proteotoxicity to the same extent as the 5MER peptide, suggesting that the peptide activates the transcellular chaperone signaling to promote proteostasis. Together, our results propose that the 5MER peptide should be considered as a component of future therapeutic cocktails for the treatment of neurodegenerative maladies.

High throughput compound screening in neuronal cells identifies statins as activators of ataxin 3 expression

The spinocerebellar ataxias (SCA) comprise a group of inherited neurodegenerative diseases. SCA3 is the most common form, caused by the expansion of CAG repeats within the ataxin 3 (ATXN3) gene. The mutation results in the expression of an abnormal protein, containing long polyglutamine (polyQ) stretches. The polyQ stretch confers a toxic gain of function and leads to misfolding and aggregation of ATXN3 in neurons. Thus, modulators of ATXN3 expression could potentially ameliorate the pathology in SCA3 patients. Therefore, we generated a CRISPR/Cas9 modified ATXN3-Exon4-Luciferase (ATXN3-LUC) genomic fusion- and control cell lines to perform a reporter cell line-based high-throughput screen comprising 2640 bioactive compounds, including the FDA approved drugs. We found no unequivocal inhibitors of, but identified statins as activators of the LUC signal in the ATXN3-LUC screening cell line. We further confirmed that Simvastatin treatment of wild type SK-N-SH cells increases ATXN3 mRNA and protein levels which likely results from direct binding of the activated sterol regulatory element binding protein 1 (SREBP1) to the ATXN3 promotor. Finally, we observed an increase of normal and expanded ATXN3 protein levels in a patient-derived cell line upon Simvastatin treatment, underscoring the potential medical relevance of our findings.

Pathogen-specific structural features of Candida albicans Ras1 activation complex: uncovering new antifungal drug targets.

An important feature associated with Candida albicans pathogenicity is its ability to switch between yeast and hyphal forms, a process in which CaRas1 plays a key role. CaRas1 is activated by the guanine nucleotide exchange factor (GEF) CaCdc25, triggering hyphal growth-related signaling pathways through its conserved GTP-binding (G)-domain. An important function in hyphal growth has also been proposed for the long hypervariable region downstream the G-domain, whose unusual content of polyglutamine stretches and Q/N repeats make CaRas1 unique within Ras proteins. Despite its biological importance, both the structure of CaRas1 and the molecular basis of its activation by CaCdc25 remain unexplored. Here, we show that CaRas1 has an elongated shape and limited conformational flexibility and that its hypervariable region contains helical structural elements, likely forming an intramolecular coiled-coil. Functional assays disclosed that CaRas1-activation by CaCdc25 is highly efficient, with activities up to 2,000-fold higher than reported for human GEFs. The crystal structure of the CaCdc25 catalytic region revealed an active conformation for the α-helical hairpin, critical for CaRas1-activation, unveiling a specific region exclusive to CTG-clade species. Structural studies on CaRas1/CaCdc25 complexes also revealed an interaction surface clearly distinct from that of homologous human complexes. Furthermore, we identified an inhibitory synthetic peptide, prompting the proposal of a key regulatory mechanism for CaCdc25. To our knowledge, this is the first report of specific inhibition of the CaRas1-activation via targeting its GEF. This, together with their unique pathogen-structural features, disclose a set of novel strategies to specifically block this important virulence-related mechanism. IMPORTANCE Candida albicans is the main causative agent of candidiasis, the commonest fungal infection in humans. The eukaryotic nature of C. albicans and the rapid emergence of antifungal resistance raise the challenge of identifying novel drug targets to battle this prevalent and life-threatening disease. CaRas1 and CaCdc25 are key players in the activation of signaling pathways triggering multiple virulence traits, including the yeast-to-hypha interconversion. The structural similarity of the conserved G-domain of CaRas1 to those of human homologs and the lack of structural information on CaCdc25 has impeded progress in targeting these proteins. The unique structural and functional features for CaRas1 and CaCdc25 presented here, together with the identification of a synthetic peptide capable of specifically inhibiting the GEF activity of CaCdc25, open new possibilities to uncover new antifungal drug targets against C. albicans virulence.

Autophagy in Spinocerebellar Ataxia Type 3: From Pathogenesis to Therapeutics.

Machado-Joseph disease (MJD) or spinocerebellar ataxia 3 (SCA3) is a rare, inherited, monogenic, neurodegenerative disease, and the most common SCA worldwide. MJD/SCA3 causative mutation is an abnormal expansion of the triplet CAG at exon 10 within the ATXN3 gene. The gene encodes for ataxin-3, which is a deubiquitinating protein that is also involved in transcriptional regulation. In normal conditions, the ataxin-3 protein polyglutamine stretch has between 13 and 49 glutamines. However, in MJD/SCA3 patients, the size of the stretch increases from 55 to 87, contributing to abnormal protein conformation, insolubility, and aggregation. The formation of aggregates, which is a hallmark of MJD/SCA3, compromises different cell pathways, leading to an impairment of cell clearance mechanisms, such as autophagy. MJD/SCA3 patients display several signals and symptoms in which the most prominent is ataxia. Neuropathologically, the regions most affected are the cerebellum and the pons. Currently, there are no disease-modifying therapies, and patients rely only on supportive and symptomatic treatments. Due to these facts, there is a huge research effort to develop therapeutic strategies for this incurable disease. This review aims to bring together current state-of-the-art strategies regarding the autophagy pathway in MJD/SCA3, focusing on evidence for its impairment in the disease context and, importantly, its targeting for the development of pharmacological and gene-based therapies.

Quantifying mutant huntingtin protein in human cerebrospinal fluid to support the development of huntingtin-lowering therapies

Huntington’s disease (HD) is caused by a cytosine adenine guanine-repeat expansion in the huntingtin gene. This results in the production of toxic mutant huntingtin protein (mHTT), which has an elongated polyglutamine (polyQ) stretch near the protein’s N-terminal end. The pharmacological lowering of mHTT expression in the brain targets the underlying driver of HD and is one of the principal therapeutic strategies being pursued to slow or stop disease progression. This report describes the characterisation and validation of an assay designed to quantify mHTT in the cerebrospinal fluid of individuals with HD, for use in registrational clinical trials. The assay was optimised, and its performance was characterised with recombinant huntingtin protein (HTT) varying in overall and polyQ-repeat length. The assay was successfully validated by two independent laboratories in regulated bioanalytical environments and showed a steep signal increase as the polyQ stretch of recombinant HTTs pivoted from wild-type to mutant protein forms. Linear mixed effects modelling confirmed highly parallel concentration–response curves for HTTs, with only a minor impact of individual slopes of the concentration–response for different HTTs (typically < 5% of the overall slope). This implies an equivalent quantitative signal behaviour for HTTs with differing polyQ-repeat lengths. The reported method may be a reliable biomarker tool with relevance across the spectrum of HD mutations, which can facilitate the clinical development of HTT-lowering therapies in HD.

Treatment with the Glycosphingolipid Modulator THI Rescues Myelin Integrity in the Striatum of R6/2 HD Mice

Huntington's disease is one of the most common dominantly inherited neurodegenerative disorders caused by an expansion of a polyglutamine (polyQ) stretch in the N-terminal region of huntingtin (Htt). Among all the molecular mechanisms, affected by the mutation, emerging evidence proposes glycosphingolipid dysfunction as one of the major determinants. High levels of sphingolipids have been found to localize in the myelin sheaths of oligodendrocytes, where they play an important role in myelination stability and functions. In this study, we investigated any potential existing link between sphingolipid modulation and myelin structure by performing both ultrastructural and biochemical analyses. Our findings demonstrated that the treatment with the glycosphingolipid modulator THI preserved myelin thickness and the overall structure and reduced both area and diameter of pathologically giant axons in the striatum of HD mice. These ultrastructural findings were associated with restoration of different myelin marker protein, such as myelin-associated glycoprotein (MAG), myelin basic protein (MBP) and 2', 3' Cyclic Nucleotide 3'-Phosphodiesterase (CNP). Interestingly, the compound modulated the expression of glycosphingolipid biosynthetic enzymes and increased levels of GM1, whose elevation has been extensively reported to be associated with reduced toxicity of mutant Htt in different HD pre-clinical models. Our study further supports the evidence that the metabolism of glycosphingolipids may represent an effective therapeutic target for the disease.

Kynurenine 3-Monooxygenase Interacts with Huntingtin at the Outer Mitochondrial Membrane.

The flavoprotein kynurenine 3-monooxygenase (KMO) is localised to the outer mitochondrial membrane and catalyses the synthesis of 3-hydroxykynurenine from L-kynurenine, a key step in the kynurenine pathway (KP) of tryptophan degradation. Perturbation of KP metabolism due to inflammation has long been associated with the pathogenesis of several neurodegenerative disorders, including Huntington’s disease (HD)—which is caused by the expansion of a polyglutamine stretch in the huntingtin (HTT) protein. While HTT is primarily localised to the cytoplasm, it also associates with mitochondria, where it may physically interact with KMO. In order to test this hypothesis, we employed bimolecular fluorescence complementation (BiFC) and found that KMO physically interacts with soluble HTT exon 1 protein fragment in living cells. Notably, expansion of the disease-causing polyglutamine tract in HTT leads to the formation of proteinaceous intracellular inclusions that disrupt this interaction with KMO, markedly decreasing BiFC efficiency. Using confocal microscopy and ultrastructural analysis, we determined KMO and HTT localisation within the cell and found that the KMO-HTT interaction is localized to the outer mitochondrial membrane. These data suggest that KMO may interact with a pool of HTT at the mitochondrial membrane, highlighting a possible physiological role for mitochondrial HTT. The KMO-HTT interaction is abrogated upon polyglutamine expansion, which may indicate a heretofore unrecognized relevance in the pathogenesis of this disorder.

Huntingtin and Its Partner Huntingtin-Associated Protein 40: Structural and Functional Considerations in Health and Disease.

Since the discovery of the mutation causing Huntington’s disease (HD) in 1993, it has been debated whether an expanded polyglutamine (polyQ) stretch affects the properties of the huntingtin (HTT) protein and thus contributes to the pathological mechanisms responsible for HD. Here we review the current knowledge about the structure of HTT, alone (apo-HTT) or in a complex with Huntingtin-Associated Protein 40 (HAP40), the influence of polyQ-length variation on apo-HTT and the HTT-HAP40 complex, and the biology of HAP40. Phylogenetic analyses suggest that HAP40 performs essential functions. Highlighting the relevance of its interaction with HTT, HAP40 is one of the most abundant partners copurifying with HTT and is rapidly degraded, when HTT levels are reduced. As the levels of both proteins decrease during disease progression, HAP40 could also be a biomarker for HD. Whether declining HAP40 levels contribute to disease etiology is an open question. Structural studies have shown that the conformation of apo-HTT is less constrained but resembles that adopted in the HTT-HAP40 complex, which is exceptionally stable because of extensive interactions between HAP40 and the three domains of HTT. The complex— and to some extent apo-HTT— resists fragmentation after limited proteolysis. Unresolved regions of apo-HTT, constituting about 25% of the protein, are the main sites of post-translational modifications and likely have major regulatory functions. PolyQ elongation does not substantially alter the structure of HTT, alone or when associated with HAP40. Particularly, polyQ above the disease length threshold does not induce drastic conformational changes in full-length HTT. Therefore, models of HD pathogenesis stating that polyQ expansion drastically alters HTT properties should be reconsidered.

Generation of induced pluripotent stem cell line, ICGi033-A, by reprogramming peripheral blood mononuclear cells from a patient with Huntington's disease

Huntington's disease (HD) is a hereditary autosomal dominant neurodegenerative disease caused by the polyglutamine stretch expansion in the huntingtin (HTT) protein. In HD, dysregulation of multiple cellular processes occurs, resulting in the death of medium spiny neurons of striatum. A line of induced pluripotent stem cells (iPSCs) ICGi033-A was obtained from peripheral blood mononuclear cells of a patient carrying 77 CAG repeats in the HTT gene. The iPSCs express pluripotency markers, have a normal karyotype, and differentiate into three germ layers: endoderm, ectoderm, mesoderm.

Alternative processing of human HTT mRNA with implications for Huntington's disease therapeutics.

Huntington disease is caused by a CAG repeat expansion in exon 1 of the huntingtin gene (HTT) that is translated into a polyglutamine stretch in the huntingtin protein (HTT). We previously showed that HTT mRNA carrying an expanded CAG repeat was incompletely spliced to generate HTT1a, an exon 1 only transcript, which was translated to produce the highly aggregation-prone and pathogenic exon 1 HTT protein. This occurred in all knock-in mouse models of Huntington's disease and could be detected in patient cell lines and post-mortem brains. To extend these findings to a model system expressing human HTT, we took advantage of YAC128 mice that are transgenic for a yeast artificial chromosome carrying human HTT with an expanded CAG repeat. We discovered that the HTT1a transcript could be detected throughout the brains of YAC128 mice. We implemented RNAscope to visualize HTT transcripts at the single molecule level and found that full-length HTT and HTT1a were retained together in large nuclear RNA clusters, as well as being present as single transcripts in the cytoplasm. Homogeneous time-resolved fluorescence analysis demonstrated that the HTT1a transcript had been translated to produce the exon 1 HTT protein. The levels of exon 1 HTT in YAC128 mice, correlated with HTT aggregation, supportive of the hypothesis that exon 1 HTT initiates the aggregation process. Huntingtin-lowering strategies are a major focus of therapeutic development for Huntington's disease. These approaches often target full-length HTT alone and would not be expected to reduce pathogenic exon 1 HTT levels. We have established YAC128 mouse embryonic fibroblast lines and shown that, together with our QuantiGene multiplex assay, these provide an effective screening tool for agents that target HTT transcripts. The effects of current targeting strategies on nuclear RNA clusters are unknown, structures that may have a pathogenic role or alternatively could be protective by retaining HTT1a in the nucleus and preventing it from being translated. In light of recently halted antisense oligonucleotide trials, it is vital that agents targeting HTT1a are developed, and that the effects of HTT-lowering strategies on the subcellular levels of all HTT transcripts and their various HTT protein isoforms are understood.

Time-resolved FRET screening identifies small molecular modifiers of mutant Huntingtin conformational inflexibility in patient-derived cells.

Huntington's disease (HD) is the most common monogenic neurodegenerative disease and is fatal. CAG repeat expansions in mutant Huntingtin (mHTT) exon 1encode for polyglutamine (polyQ) stretches and influence age of onset and disease severity, depending on their length. mHTT is more structured compared to wild-type (wt) HTT, resulting in a decreased N-terminal conformational flexibility. mHTT inflexibility may contribute to both gain of function toxicity, due to increased mHTT aggregation propensity, but also to loss of function phenotypes, due to decreased interactions with binding partners. High-throughput-screening techniques to identify mHTT flexibility states and potential flexibility modifying small molecules are currently lacking. Here, we propose a novel approach for identifying small molecules that restore mHTT's conformational flexibility in human patient fibroblasts. We have applied a well-established antibody-based time-resolved Förster resonance energy transfer (TR-FRET) immunoassay, which measures endogenous HTT flexibility using two validated HTT-specific antibodies, to a high-throughput screening platform. By performing a small-scale compound screen, we identified several small molecules that can partially rescue mHTT inflexibility, presumably by altering HTT post-translational modifications. Thus, we demonstrated that the HTT TR-FRET immunoassay can be miniaturized and applied to a compound screening workflow in patient cells. This automated assay can now be used in large screening campaigns to identify previously unknown HD drugs and drug targets.

Expression profile analysis in cells overexpressing DRPLA cDNA to explore the roles of DRPLAp as a transcriptional coregulator

AbstractBackgroundDentatorubral‐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.AimTo elucidate the physiological functions of DRPLAp based on determination of the target genes regulated by wild‐type DRPLAp.MethodsUsing 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).ResultsOn 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.ConclusionThese findings support that DRPLAp may play a role as a transcriptional coregulator involved in the regulation of development.

Neuropeptide signaling and SKN-1 orchestrate differential responses of the proteostasis network to dissimilar proteotoxic insults.

The protein homeostasis (proteostasis) network (PN) encompasses mechanisms that maintain proteome integrity by controlling various biological functions. Loss of proteostasis leads to toxic protein aggregation (proteotoxicity), which underlies the manifestation of neurodegeneration. How the PN responds to dissimilar proteotoxic challenges and how these responses are regulated at the organismal level are largely unknown. Here, we report that, while torsin chaperones protect from the toxicity of neurodegeneration-causing polyglutamine stretches, they exacerbate the toxicity of the Alzheimer's disease-causing Aβ peptide in neurons and muscles. These opposing effects are accompanied by differential modulations of gene expression, including that of three neuropeptides that are involved in tailoring the organismal response to dissimilar proteotoxic insults. This mechanism is regulated by insulin/IGF signaling and the transcription factor SKN-1/NRF. Our work delineates a mechanism by which the PN orchestrates differential responses to dissimilar proteotoxic challenges and points at potential targets for therapeutic interventions.

Transposable element activation promotes neurodegeneration in a Drosophila model of Huntington's disease

SummaryHuntington's disease (HD) is an autosomal dominant disorder with progressive motor dysfunction and cognitive decline. The disease is caused by a CAG repeat expansion in the IT15 gene, which elongates a polyglutamine stretch of the HD protein, Huntingtin. No therapeutic treatments are available, and new pharmacological targets are needed. Retrotransposons are transposable elements (TEs) that represent 40% and 30% of the human and Drosophila genomes and replicate through an RNA intermediate. Mounting evidence suggests that mammalian TEs are active during neurogenesis and may be involved in diseases of the nervous system. Here we show that TE expression and mobilization are increased in a Drosophila melanogaster HD model. By inhibiting TE mobilization with Reverse Transcriptase inhibitors, polyQ-dependent eye neurodegeneration and genome instability in larval brains are rescued and fly lifespan is increased. These results suggest that TE activation may be involved in polyQ-induced neurotoxicity and a potential pharmacological target.

Huntington's Disease Gene Hunters: An Expanding Tale.

Huntington's Disease Gene Hunters: An Expanding Tale.

Allele-specific targeting of mutant ataxin-3 by antisense oligonucleotides in SCA3-iPSC-derived neurons

Spinocerebellar ataxia type 3 (SCA3) is caused by an expanded polyglutamine stretch in ataxin-3. While wild-type ataxin-3 has important functions, e.g., as a deubiquitinase, downregulation of mutant ataxin-3 is likely to slow down the course of this fatal disease. We established a screening platform with human neurons of patients and controls derived from induced pluripotent stem cells to test antisense oligonucleotides (ASOs) for their effects on ataxin-3 expression. We identified an ASO that suppressed mutant and wild-type ataxin-3 levels by >90% after a singular treatment. Next, we screened pairs of ASOs designed to selectively target the mutant or the wild-type allele by taking advantage of a SNP (c.987G > C) in ATXN3 that is present in most SCA3 patients. We found ASOmut4 to reduce levels of mutant ataxin-3 by 80% after 10 days while leaving expression of wild-type ataxin-3 largely unaffected. In a long-term study we proved this effect to last for about 4 weeks after a single treatment without signs of neurotoxicity. This study provides proof of principle that allele-specific lowering of poly(Q)-expanded ataxin-3 by selective ASOs is feasible and long lasting, with sparing of wild-type ataxin-3 expression in a human cell culture model that is genetically identical to SCA3 patients.