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Dystonia due to pathogenic variants in the THAP1 gene (DYT-THAP1) shows variable expressivity and reduced penetrance of ~ 50%. Since THAP1 encodes a transcription factor, modifiers influencing this variability likely operate at the gene expression level. This study aimed to assess the transferability of differentially expressed genes (DEGs) in neuronal cells related to pathogenic variants in the THAP1 gene, which were previously identified by transcriptome analyses. For this, we performed quantitative (qPCR) and Digital PCR (dPCR) in cultured fibroblasts. RNA was extracted from THAP1 manifesting (MMCs) and non-manifesting mutation carriers (NMCs) as well as from healthy controls. The expression profiles of ten of 14 known neuronal DEGs demonstrated differences in fibroblasts between these three groups. This included transcription factors and targets (ATF4, CLN3, EIF2A, RRM1, YY1), genes involved in G protein-coupled receptor signaling (BDKRB2, LPAR1), and a gene linked to apoptosis and DNA replication/repair (CRADD), which all showed higher expression levels in MMCs and NMCs than in controls. Moreover, the analysis of genes linked to neurological disorders (STXBP1, TOR1A) unveiled differences in expression patterns between MMCs and controls. Notably, the genes CUEDC2, DRD4, ECH1, and SIX2 were not statistically significantly differentially expressed in fibroblast cultures. With > 70% of the tested genes being DEGs also in fibroblasts, fibroblasts seem to be a suitable model for DYT-THAP1 research despite some restrictions. Furthermore, at least some of these DEGs may potentially also serve as biomarkers of DYT-THAP1 and influence its penetrance and expressivity., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Background: The THAP1 gene encodes a transcription factor, and pathogenic variants cause a form of autosomal dominant, isolated dystonia (DYT-THAP1) with reduced penetrance. Factors underlying both reduced penetrance and the disease mechanism of DYT-THAP1 are largely unknown., Methods: We performed transcriptome analysis on 29 cortical neuronal precursors derived from human-induced pluripotent stem cell lines generated from manifesting and nonmanifesting THAP1 mutation carriers and control individuals., Results: Whole transcriptome analysis showed a penetrance-linked signature with expressional changes more pronounced in the group of manifesting (MMCs) than in nonmanifesting mutation carriers (NMCs) when compared to controls. A direct comparison of the transcriptomes in MMCs versus NMCs showed significant upregulation of the DRD4 gene in MMCs. A gene set enrichment analysis demonstrated alterations in various neurotransmitter release cycle pathways, extracellular matrix organization, and deoxyribonucleic acid methylation between MMCs and NMCs. When specifically considering transcription factors, the expression of YY1 and SIX2 differed in MMCs versus NMCs. Further, THAP1 was upregulated in the group of MMCs., Conclusions: To our knowledge, this is the first report systematically analyzing reduced penetrance in DYT-THAP1 in a human model using transcriptomes. Our findings indicate that transcriptional alterations during cortical development influence DYT-THAP1 pathogenesis and penetrance. We reinforce previously linked pathways including dopamine and eukaryotic translation initiation factor 2 alpha signaling in the pathogenesis of dystonia including DYT-THAP1 and suggest extracellular matrix organization and deoxyribonucleic acid methylation as mediators of disease protection. © 2021 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society., (© 2021 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.)

Mutations in THAP1 (THAP domain-containing apoptosis-associated protein 1) cause a form of early-onset, isolated dystonia (DYT-THAP1, aka DYT6). Here, we describe the generation of eight human induced pluripotent stem cell (iPSC) lines of manifesting and non-manifesting carriers of the THAP1 mutations p.Lys158Asnfs*23 or p.Arg13His (each 4 lines). Dermal fibroblasts were reprogrammed using non-integrating Sendai virus. The iPSC lines were comprehensively characterized including expression analyses of pluripotency markers, the potential to differentiate into cells of all three germ layers, and stable karyotypes. These lines provide a valuable resource for studying the impact of THAP1 mutations on the pathology of dystonia., (Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.)

Thanatos-associated protein domain containing, apoptosis-associated protein 1 (THAP1), the gene mutated in DYT6 dystonia, encodes a transcription factor. While the N-terminal THAP domain allows for specific DNA-binding, the functional relevance of the other regions is largely unknown. The C-terminus contains a 4-amino-acid-spanning host cell factor 1 (HCFC1)-binding domain (HBM) that mediates the interaction with HCFC1. Interestingly, three mutations affecting the HBM (p.N136S, p.N136K, p.Y137C) have been reported in dystonia patients. We investigated the consequences of these mutations on the interaction of THAP1 with HCFC1 and demonstrated that all three mutations abolished HCFC1-THAP1 complex formation. Notably, HCFC1 co-localization was found in >90% of the almost 3,500 chromatin regions loaded with THAP1 in publicly available genome-wide ChIP data. By siRNA-mediated depletion of HCFC1, we detected an increase of THAP1 expression, indicating a co-repressor activity of HCFC1 for THAP1. Quantitative ChIP on selected promoters revealed that none of the mutations significantly decreased the DNA-binding ability of THAP1 while HCFC1 binding was highly reduced. Our findings indicate a THAP1-mediated recruitment of HCFC1 to THAP1 target sites. Of note, dystonia-causing mutations within the HBM in THAP1 abolished this interaction. Thus, we demonstrate disrupted THAP1-HCFC1 complex formation as another mechanism of dystonia-causing mutations leading to transcriptional dysregulation., (© The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Mutations in the THAP1 gene encoding the transcription factor THAP1 have been shown to cause DYT6 dystonia. THAP1 contains a highly conserved THAP zinc finger at its N-terminal region which allows specific binding to its target sequences as well as a coiled-coil domain (amino acids 139-190) towards its C-terminus postulated as a protein-protein-binding motif. While several DYT6-causing mutations within the THAP domain were shown to decrease THAP1 activity in transcriptional regulation and DNA-binding, the role of mutations within the coiled-coil domain is rather unknown. Therefore, assigning a function to this domain may enable functional testing of mutations in this region. Notably, THAP1 and other THAP proteins form homodimers; however, the responsible domain has not been elucidated in detail. We show that the region of amino acids 139-185 is involved in formation of THAP1 homodimers by using yeast-two-hybrid, GST pull-down, and cross-linking assays. Surprisingly, all nine reported DYT6-causing missense mutations within this region had no effect on dimerization of THAP1 in GST pull-down and formaldehyde cross-linking assays. In conclusion, we demonstrated that a region of 47 amino acids is involved in THAP1 homodimerization but mutations in this region seem not to impair this mechanism.

Deep brain stimulation (DBS) of the globus pallidus internus (GPi) is an established treatment in patients with severe dystonia. However, factors predicting outcome are largely unknown and motor improvement in DYT6 patients after DBS has been reported to be poorer as compared to, e.g., DYT1 patients. Here, we report the course of clinical improvement for up to 11 years of pallidal DBS in three male patients belonging to the same family with early-onset generalized or segmental dystonia due to a heterozygous THAP1 gene mutation (DYT6). All patients showed an initial effective response to pallidal DBS with a mean of 56.9 ± 11.7% improvement in the Burke-Fahn-Marsden Dystonia motor and 45.5 ± 22.4% in the disability score at 1-year follow-up. The long-term outcome of pallidal DBS was favorable in two patients (39, 67% motor improvement, respectively). Our findings demonstrate that motor improvement is variable and may depend on disease severity, disease duration, and clinical presentation. Overall, our observation supports pallidal DBS as an important treatment option in patients with DYT6 dystonia.

THAP1 encodes a transcription factor but its regulation is largely elusive. TOR1A was shown to be repressed by THAP1 in vitro. Notably, mutations in both of these genes lead to dystonia (DYT6 or DYT1). Surprisingly, expressional changes of TOR1A in THAP1 mutation carriers have not been detected indicating additional levels of regulation. Here, we investigated whether THAP1 is able to autoregulate its own expression. Using in-silico prediction, luciferase reporter gene assays, and (quantitative) chromatin immunoprecipitation (ChIP), we defined the THAP1 minimal promoter to a 480bp-fragment and demonstrated specific binding of THAP1 to this region which resulted in repression of the THAP1 promoter. This autoregulation was disturbed by different DYT6-causing mutations. Two mutants (Ser6Phe, Arg13His) were shown to be less stable than wildtype THAP1 adding to the effect of reduced binding to the THAP1 promoter. Overexpressed THAP1 is preferably degraded through the proteasome. Notably, endogenous THAP1 expression was significantly reduced in cells overexpressing wildtype THAP1 as demonstrated by quantitative PCR. In contrast, higher THAP1 levels were detected in induced pluripotent stem cell (iPS)-derived neurons from THAP1 mutation carriers. Thus, we identified a feedback-loop in the regulation of THAP1 expression and demonstrated that mutant THAP1 leads to higher THAP1 expression levels. This compensatory autoregulation may contribute to the mean age at onset in the late teen years or even reduced penetrance in some THAP1 mutation carriers., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Mutations in THAP1 have been associated with dystonia 6 (DYT6). THAP1 encodes a transcription factor that represses the expression of DYT1. To further evaluate the mutational spectrum of THAP1 and its associated phenotype, we sequenced THAP1 in 567 patients with focal (n = 461), segmental (n = 68), or generalized dystonia (n = 38). We identified 10 novel variants, including six missense substitutions within the DNA-binding Thanatos-associated protein domain (Arg13His, Lys16Glu, His23Pro, Lys24Glu, Pro26Leu, Ile80Val), a 1bp-deletion downstream of the nuclear localization signal (Asp191Thrfs*9), and three alterations in the untranslated regions. The effect of the missense variants was assessed using prediction tools and luciferase reporter gene assays. This indicated the Ile80Val substitution as a benign variant. The subcellular localization of Asp191Thrfs*9 suggests a disturbed nuclear import for this mutation. Thus, we consider six of the 10 novel variants as pathogenic mutations accounting for a mutation frequency of 1.1%. Mutation carriers presented mainly with early onset dystonia (<12 years in five of six patients). Symptoms started in an arm or neck and spread to become generalized in three patients or segmental in two patients. Speech was affected in four mutation carriers. In conclusion, THAP1 mutations are rare in unselected dystonia patients and functional analysis is necessary to distinguish between benign variants and pathogenic mutations.

To identify the underlying genetic cause in a consanguineous family with apparently recessively inherited dystonia, we performed genome-wide homozygosity mapping. This revealed 2 candidate regions including the THAP1 gene, where heterozygous mutations cause dystonia 6. A homozygous missense mutation in THAP1 (c.95T>A; p.Leu32His) was found in all 3 affected siblings. Symptoms started in childhood in the legs and became generalized within a few years. Three heterozygous mutation carriers were unaffected. Because THAP1 regulates the expression of the DYT1 gene, we used reporter gene assays to show that DYT1 expression was significantly increased for Leu32His. However, this increase was less pronounced than for other THAP1 mutations that cause dystonia in the heterozygous state. Our data suggest that homozygous THAP1 mutations cause dystonia and may be associated with a less severe dysfunction of the encoded protein compared with heterozygous disease-causing mutations., (Copyright © 2011 Movement Disorder Society.)

The purpose of the study was to delineate clinical and electrophysiological characteristics as well as laryngoscopical and transcranial ultrasound (TCS) findings in THAP1 mutation carriers (MutC). According to recent genetic studies, DYT6 (THAP1) gene mutations are an important cause of primary early-onset dystonia. In contrast to DYT1 mutations, THAP1 mutations are associated with primary early-onset segmental or generalised dystonia frequently involving the craniocervical region and the larynx. Blood samples from twelve individuals of three German families with DYT6 positive index cases were obtained to test for THAP1 mutations. Eight THAP1 MutC were identified. Of these, six (three symptomatic and three asymptomatic) THAP1 MutC could be clinically evaluated. Laryngoscopy was performed to evaluate laryngeal dysfunction in patients. Brainstem echogenicity was investigated in all MutC using TCS. Two of the patients had undergone bilateral pallidal DBS. In all three symptomatic MutC, early-onset laryngeal dystonia was a prominent feature. Laryngeal assessment demonstrated adductor-type dystonia in all of them. On clinical examination, the three asymptomatic MutC also showed subtle signs of focal or segmental dystonia. TCS revealed increased substantia nigra (SN) hyperechogenicity in all MutC. Intraoperative microelectrode recordings under general anesthesia in two of the patients showed no difference between THAP1 and previously operated DYT1 MutC. The presence of spasmodic dysphonia in patients with young-onset segmental or generalised dystonia is a hallmark of DYT6 dystonia. SN hyperechogenicity on TCS may represent an endophenotype in these patients. Pallidal DBS in two patients was unsatisfactory.

Mutations in THAP1 have been associated with dystonia 6. THAP1 encodes a transcription factor with mostly unknown targets. We tested the hypothesis that THAP1 regulates the expression of DYT1 (TOR1A), another dystonia-causing gene. After characterization of the TOR1A promoter, we demonstrate that THAP1 binds to the core promoter of TOR1A. Further, we report that wild type THAP1 represses the expression of TOR1A, whereas dystonia 6-associated mutant THAP1 results in decreased repression of TOR1A. Our data demonstrate that THAP1 regulates the transcription of TOR1A, suggesting transcriptional dysregulation as a cause of dystonia.

Background: DYT6 is a primary, early-onset torsion dystonia; however, unlike in DYT1 dystonia, the symptoms of DYT6 dystonia frequently involve the craniocervical region. Recently, two mutations in THAP1, the gene that encodes THAP (thanatos-associated protein) domain-containing apoptosis-associated protein 1 (THAP1), have been identified as a cause of DYT6 dystonia., Methods: We screened THAP1 by sequence analysis and quantitative real-time polymerase chain reaction (PCR) in 160 white patients of European ancestry who had dystonia with an early age at onset (n=64), generalised dystonia (n=35), a positive family history of dystonia (n=56), or facial or laryngeal dystonia. Another 160 patients with dystonia were screened for reported and novel variants in THAP1. 280 neurologically healthy controls were screened for the newly identified and previously reported changes in THAP1 and these and an additional 75 controls were screened for a rare non-coding mutation., Findings: We identified two mutations in THAP1 (388&#95;389delTC and 474delA), respectively, in two (1%) German patients from the 160 patients with dystonia. Both mutation carriers had laryngeal dystonia that started in childhood and both went on to develop generalised dystonia. Thus, two of three patients with early-onset generalised dystonia with orobulbar involvement had mutations in THAP1. One of the identified patients with DYT6 dystonia had two family members with subtle motor signs who also carried the same mutation. A rare substitution in the 5'untranslated region (-236&#95;235GA-->TT) was found in 20 of 320 patients and in seven of 355 controls (p=0.0054)., Interpretation: Although mutations in THAP1 might have only a minor role in patients with different, but mainly focal, forms of dystonia, they do seem to be associated with early-onset generalised dystonia with spasmodic dysphonia. This combination of symptoms might be a characteristic feature of DYT6 dystonia and could be useful in the differential diagnosis of DYT1, DYT4, DYT12, and DYT17 dystonia. In addition to the identified mutations, a rare non-coding substitution in THAP1 might increase the risk of dystonia., Funding: Deutsche Forschungsgemeinschaft; Volkswagen Foundation; Dystonia Medical Research Foundation; University of Lübeck.