polyQ Expansion Disorders Research Articles

Association with proteasome determines pathogenic threshold of polyglutamine expansion diseases

Expansion of glutamine residue track (polyQ) within soluble protein is responsible for eight autosomal-dominant genetic neurodegenerative disorders. These disorders affect cerebellum, striatum, basal ganglia and other brain regions. Each disease develops when polyQ expansion exceeds a pathogenic threshold (Qth). A pathogenic threshold is unique for each disease but the reasons for variability in Qth within this family of proteins are poorly understood. In the previous publication we proposed that polarity of the regions flanking polyQ track in each protein plays a key role in defining Qth value [1]. To explain the correlation between the polarity of the flanking sequences and Qth we performed quantitative analysis of interactions between polyQ-expanded proteins and proteasome. Based on structural and theoretical modeling, we predict that Qth value is determined by the energy of polar interaction of the flanking regions with the polyQ and proteasome. More polar flanking regions facilitate unfolding of α-helical polyQ conformation adopted inside the proteasome and as a result, increase Qth. Predictions of our model are consistent with Qth values observed in clinic for each of the eight polyQ-expansion disorders. Our results suggest that the agents that can destabilize polyQ α-helical structure may have a beneficial therapeutic effect for treatment of polyQ-expansion disorders.

The 2.2-Angstrom resolution crystal structure of the carboxy-terminal region of ataxin-3.

An expansion of polyglutamine (polyQ) sequence in ataxin‐3 protein causes spinocerebellar ataxia type 3, an inherited neurodegenerative disorder. The crystal structure of the polyQ‐containing carboxy‐terminal fragment of human ataxin‐3 was solved at 2.2‐Å resolution. The Atxn3 carboxy‐terminal fragment including 14 glutamine residues adopts both random coil and α‐helical conformations in the crystal structure. The polyQ sequence in α‐helical structure is stabilized by intrahelical hydrogen bonds mediated by glutamine side chains. The intrahelical hydrogen‐bond interactions between glutamine side chains along the axis of the polyQ α‐helix stabilize the secondary structure. Analysis of this structure furthers our understanding of the polyQ‐structural characteristics that likely underlie the pathogenesis of polyQ‐expansion disorders.

The role of the immune system in triplet repeat expansion diseases.

Trinucleotide repeat expansion disorders (TREDs) are a group of dominantly inherited neurological diseases caused by the expansion of unstable repeats in specific regions of the associated genes. Expansion of CAG repeat tracts in translated regions of the respective genes results in polyglutamine- (polyQ-) rich proteins that form intracellular aggregates that affect numerous cellular activities. Recent evidence suggests the involvement of an RNA toxicity component in polyQ expansion disorders, thus increasing the complexity of the pathogenic processes. Neurodegeneration, accompanied by reactive gliosis and astrocytosis is the common feature of most TREDs, which may suggest involvement of inflammation in pathogenesis. Indeed, a number of immune response markers have been observed in the blood and CNS of patients and mouse models, and the activation of these markers was even observed in the premanifest stage of the disease. Although inflammation is not an initiating factor of TREDs, growing evidence indicates that inflammatory responses involving astrocytes, microglia, and the peripheral immune system may contribute to disease progression. Herein, we review the involvement of the immune system in the pathogenesis of triplet repeat expansion diseases, with particular emphasis on polyglutamine disorders. We also present various therapeutic approaches targeting the dysregulated inflammation pathways in these diseases.

Pathogenic polyglutamine expansion length correlates with polarity of the flanking sequences.

BackgroundPolyglutamine (polyQ) repeat expansion within coding sequence of a soluble protein is responsible for eight autosomal-dominant genetic neurodegenerative disorders. These disorders affect cerebellum, striatum, basal ganglia and other brain regions. The pathogenic polyQ-expansion threshold in these proteins varies from 32Q to 54Q. Understanding the reasons for variability in pathogenic polyQ threshold may provide insights into pathogenic mechanisms responsible for development of these disorders.FindingsHere we established a quantitative correlation between the polarity of the flanking sequences and pathogenic polyQ-expansion threshold in this protein family. We introduced an “edge polarity index” (EPI) to quantify polarity effects of the flanking regions and established a strong correlation between EPI index and critical polyQ expansion length in this protein family. Based on this analysis we subdivided polyQ-expanded proteins into 2 groups – with strong and weak dependence of polyQ threshold on EPI index. The main difference between members of the first and the second group is a polarity profile of these proteins outside of polyQ and flanking regions. PolyQ proteins are known substrates for proteasome and most likely mechanistic explanation for the observed correlation is that proteasome may have an impaired ability to process continuous non-polar regions of proteins.ConclusionsThe proposed hypothesis provides a quantitative explanation for variability in pathogenic threshold among polyQ-expansion disorders, which we established to correlate with polarity of flanking regions. To explain these results we propose that proteasome is not efficient in processing continuous non-polar regions of proteins, resulting in release of undigested and partially digested fragments. If supported experimentally, our hypothesis may have wide implications for further understanding the pathogensis of polyglutamine expansion disorders.

Ataxin-3 protein and RNA toxicity in spinocerebellar ataxia type 3: current insights and emerging therapeutic strategies.

Ataxin-3 is a ubiquitously expressed deubiqutinating enzyme with important functions in the proteasomal protein degradation pathway and regulation of transcription. The C-terminus of the ataxin-3 protein contains a polyglutamine (PolyQ) region that, when mutationally expanded to over 52 glutamines, causes the neurodegenerative disease spinocerebellar ataxia 3 (SCA3). In spite of extensive research, the molecular mechanisms underlying the cellular toxicity resulting from mutant ataxin-3 remain elusive and no preventive treatment is currently available. It has become clear over the last decade that the hallmark intracellular ataxin-3 aggregates are likely not the main toxic entity in SCA3. Instead, the soluble PolyQ containing fragments arising from proteolytic cleavage of ataxin-3 by caspases and calpains are now regarded to be of greater influence in pathogenesis. In addition, recent evidence suggests potential involvement of a RNA toxicity component in SCA3 and other PolyQ expansion disorders, increasing the pathogenic complexity. Herein, we review the functioning of ataxin-3 and the involvement of known protein and RNA toxicity mechanisms of mutant ataxin-3 that have been discovered, as well as future opportunities for therapeutic intervention.

Beta conformation of polyglutamine track revealed by a crystal structure of Huntingtin N-terminal region with insertion of three histidine residues

Huntington disease is an autosomal-dominant neurodegenerative disorder caused by a polyglutamine (polyQ) expansion (> 35Q) in the first exon (EX1) of huntingtin protein (Htt). mHtt protein is thought to adopt one or more toxic conformation(s) that are involved in pathogenic interactions in cells . However, the structure of mHtt is not known. Here, we present a near atomic resolution structure of mHtt36Q-EX1. To facilitate crystallization, three histidine residues (3H) were introduced within the Htt36Q stretch resulting in the sequence of Q7HQHQHQ27. The Htt36Q3H region adopts α-helix, loop, β-hairpin conformations. Furthermore, we observed interactions between the backbone of the Htt36Q3H β-strand with the aromatic residues mimicking putative-toxic interactions with other proteins. Our findings support previous predictions that the expanded mHtt-polyQ region adopts a β-sheet structure. Detailed structural information about mHtt improves our understanding of the pathogenic mechanisms in HD and other polyQ expansion disorders and may form the basis for rational design of small molecules that target toxic conformations of disease-causing proteins.

Dantrolene is neuroprotective in Huntington's disease transgenic mouse model

BackgroundHuntington's disease (HD) is a progressive neurodegenerative disorder caused by a polyglutamine expansion in the Huntingtin protein which results in the selective degeneration of striatal medium spiny neurons (MSNs). Our group has previously demonstrated that calcium (Ca2+) signaling is abnormal in MSNs from the yeast artificial chromosome transgenic mouse model of HD (YAC128). Moreover, we demonstrated that deranged intracellular Ca2+ signaling sensitizes YAC128 MSNs to glutamate-induced excitotoxicity when compared to wild type (WT) MSNs. In previous studies we also observed abnormal neuronal Ca2+ signaling in neurons from spinocerebellar ataxia 2 (SCA2) and spinocerebellar ataxia 3 (SCA3) mouse models and demonstrated that treatment with dantrolene, a ryanodine receptor antagonist and clinically relevant Ca2+ signaling stabilizer, was neuroprotective in experiments with these mouse models. The aim of the current study was to evaluate potential beneficial effects of dantrolene in experiments with YAC128 HD mouse model.ResultsThe application of caffeine and glutamate resulted in increased Ca2+ release from intracellular stores in YAC128 MSN cultures when compared to WT MSN cultures. Pre-treatment with dantrolene protected YAC128 MSNs from glutamate excitotoxicty, with an effective concentration of 100 nM and above. Feeding dantrolene (5 mg/kg) twice a week to YAC128 mice between 2 months and 11.5 months of age resulted in significantly improved performance in the beam-walking and gait-walking assays. Neuropathological analysis revealed that long-term dantrolene feeding to YAC128 mice significantly reduced the loss of NeuN-positive striatal neurons and reduced formation of Httexp nuclear aggregates.ConclusionsOur results support the hypothesis that deranged Ca2+ signaling plays an important role in HD pathology. Our data also implicate the RyanRs as a potential therapeutic target for the treatment of HD and demonstrate that RyanR inhibitors and Ca2+ signaling stabilizers such as dantrolene should be considered as potential therapeutics for the treatment of HD and other polyQ-expansion disorders.

Expanded Polyglutamine-Binding Peptoid as a Novel Therapeutic Agent for Treatment of Huntington's Disease

Polyglutamine(polyQ)-expanded proteins are potential therapeutic targets for the treatment of polyQ expansion disorders such as Huntington's disease (HD) and spinocerebellar ataxia type 3 (SCA3). Here, we used an amino-terminal fragment of a mutant Huntingtin protein (Htt-N-82Q) as bait in an unbiased screen of a 60,000 peptoid library. Peptoid HQP09 was selected from the isolated hits and confirmed as a specific ligand of Htt-N-82Q and Atxn3-77Q mutant proteins in biochemical experiments. We identified three critical residues in the HQP09 sequence that are important for its activity and generated a minimalderivative, HQP09_9, which maintains the specific polyQ-binding activity. We demonstrated that HQP09 and HQP09_9 inhibited aggregation of Htt-N-53Q invitro and exerted Ca(2+)-stabilizing and neuroprotective effects in experiments with primary striatal neuronal cultures derived from HD mice. We further demonstrated that intracerebroventricular delivery of HQP09 to an HD mouse model resulted in reduced accumulation of mutant Huntingtin aggregates and improved motor behavioral outcomes. These results suggest that HQP09 and similar peptoids hold promise as novel therapeutics for developing treatments for HD, SCA3, and other polyglutamine expansion disorders.

Chemical Inducers of Autophagy That Enhance the Clearance of Mutant Proteins in Neurodegenerative Diseases

Many of the neurodegenerative diseases that afflict people are caused by intracytoplasmic aggregate-prone proteins. These include Parkinson disease, tauopathies, and polyglutamine expansion diseases such as Huntington disease. In Mendelian forms of these diseases, the mutations generally confer toxic novel functions on the relevant proteins. Thus, one potential strategy for dealing with these mutant proteins is to enhance their degradation. This can be achieved by up-regulating macroautophagy, which we will henceforth call autophagy. In this minireview, we will consider the reasons why autophagy up-regulation may be a powerful strategy for these diseases. In addition, we will consider some of the drugs and associated signaling pathways that can be used to induce autophagy with these therapeutic aims in mind.

Therapeutic prospects for spinocerebellar ataxia type 2 and 3.

Spinocerebellar ataxia type 2 (SCA2) and type 3 (SCA3) are autosomal-dominant neurodegenerative disorders. SCA2 primarily affects cerebellar Purkinje neurons. SCA3 primarily affects dentate and pontine nuclei and substantia nigra. Both disorders belong to a class of polyglutamine (polyQ) expansion disorders. SCA2 is caused by a polyQ expansion in the amino-terminal region of a cytosolic protein ataxin-2 (Atxn2). SCA3 is caused by a polyQ expansion in the carboxy-terminal portion of a cytosolic protein ataxin-3 (Atxn3). Both disorders are found worldwide, but SCA2 is common among people of Cuban decent and SCA3 is common among people of Portuguese decent. No effective treatment exist for SCA2, SCA3 or any other polyQ-expansion disorder. Based on anecdotal evidence, a number of small scale clinical trials have been attempted previously for SCA2 and SCA3. These trials were underpowered and did not yield any promising results so far. A number of pathogenic mechanisms have been proposed to explain neuronal dysfunction and degeneration in SCA2 and SCA3. Knockdown of mutant Atxn2 and Atxn3 protein by RNAi or similar approach is most promising avenue of therapeutic development in the long term, but translation of this approach to clinic faces very serious technical challenges. Recent preclinical studies in SCA2 and SCA3 genetic mouse model suggested that abnormal neuronal calcium (Ca2+) signaling may play an important role in SCA2 and SCA3 pathology. These studies also suggested that dantrolene and other Ca2+ signaling inhibitors and stabilizers may have a therapeutic value for treatment of SCA2 and SCA3. Controlled clinical evaluation of dantrolene, memantine, riluzole, dihydropyridines, CoQ10, creatine or other Ca2+ blockers and stabilizers in SCA2 and SCA3 patients is necessary to test clinical importance of these ideas. The EUROSCA consortium provides a potential framework for such clinical evaluation.