Dystonia, a debilitating movement disorder characterized by abnormal fixed positions and/or twisting postures, is associated with dysfunction of motor control networks. While gross brain lesions can produce secondary dystonias, advanced neuroimaging techniques have been required to identify network abnormalities in primary dystonias. Prior neuroimaging studies have provided valuable insights into the pathophysiology of dystonia, but few directly assessed the gross volume of motor control regions, and to our knowledge, none identified abnormalities common to multiple types of idiopathic focal dystonia.

Upstream open reading frames (uORFs) have emerged as major post-transcriptional regulatory elements in eukaryotic species. In general, uORFs are initiated by a translation start codon within the 5' untranslated region of a gene (upstream ATG; uATG), and they are negatively correlated with translational efficiency. In addition to their translational regulatory role, some uORFs can code for biologically active short peptides. The importance of uATGs/uORFs is further underscored by human diseases associated with single nucleotide polymorphisms (SNPs), which disrupt existing uORFs or introduce novel uORFs. Although several functional proteins translated from naturally occurring uORFs have been described, the coding potential of uORFs created by SNPs has been ignored because of the a priori assumption that these proteins are short-lived with no likely impact on protein homeostasis. Thus, studies on SNP-created uORFs are limited to their translational effects, leaving unexplored the potential cellular consequences of a SNP/uORF-encoded protein. Here, we investigate functionality of a uATG/uORF introduced by a +142C>T SNP within the GCH1 gene and associated with a familial form of DOPA Responsive Dystonia. We report that the +142C>T SNP represses GCH1 translation, and introduces a short, frame shifted uORF that encodes a 73-amino acid peptide. This peptide is localized within the nucleus and compromises cell viability upon proteasome inhibition. Our work extends the list of uATG/uORF associated diseases and advances research on peptides translated from SNP-introduced uORFs, a neglected component of the proteome.

Dystonia 16 (DYT16) is caused by mutations in PACT, the protein activator of interferon-induced double-stranded RNA-activated protein kinase (PKR). PKR regulates the integrated stress response (ISR) via phosphorylation of the translation initiation factor eIF2α. This post-translational modification attenuates general protein synthesis while concomitantly triggering enhanced translation of a few specific transcripts leading either to recovery and homeostasis or cellular apoptosis depending on the intensity and duration of stress signals. PKR plays a regulatory role in determining the cellular response to viral infections, oxidative stress, endoplasmic reticulum (ER) stress, and growth factor deprivation. In the absence of stress, both PACT and PKR are bound by their inhibitor transactivation RNA-binding protein (TRBP) thereby keeping PKR inactive. Under conditions of cellular stress these inhibitory interactions dissociate facilitating PACT-PACT interactions critical for PKR activation. While both PACT-TRBP and PKR-TRBP interactions are pro-survival, PACT-PACT and PACT-PKR interactions are pro-apoptotic. In this study we evaluate if five DYT16 substitution mutations alter PKR activation and ISR. Our results indicate that the mutant DYT16 proteins show stronger PACT-PACT interactions and enhanced PKR activation. In DYT16 patient derived lymphoblasts the enhanced PACT-PKR interactions and heightened PKR activation leads to a dysregulation of ISR and increased apoptosis. More importantly, this enhanced sensitivity to ER stress can be rescued by luteolin, which disrupts PACT-PKR interactions. Our results not only demonstrate the impact of DYT16 mutations on regulation of ISR and DYT16 etiology but indicate that therapeutic interventions could be possible after a further evaluation of such strategies.

Early infantile epileptic encephalopathy-44 (EIEE44, MIM: 617132) is a previously described condition resulting from biallelic variants in UBA5, a gene involved in a ubiquitin-like post-translational modification system called UFMylation. Here we report five children from four families with biallelic pathogenic variants in UBA5 All five children presented with global developmental delay, epilepsy, axial hypotonia, appendicular hypertonia, and a movement disorder, including dystonia in four. Affected individuals in all four families have compound heterozygous pathogenic variants in UBA5 All have the recurrent mild c.1111G > A (p.Ala371Thr) variant in trans with a second UBA5 variant. One patient has the previously described c.562C > T (p. Arg188*) variant, two other unrelated patients have a novel missense variant, c.907T > C (p.Cys303Arg), and the two siblings have a novel missense variant, c.761T > C (p.Leu254Pro). Functional analyses demonstrate that both the p.Cys303Arg variant and the p.Leu254Pro variants result in a significant decrease in protein function. We also review the phenotypes and genotypes of all 15 previously reported families with biallelic UBA5 variants, of which two families have presented with distinct phenotypes, and we describe evidence for some limited genotype-phenotype correlation. The overlap of motor and developmental phenotypes noted in our cohort and literature review adds to the increasing understanding of genetic syndromes with movement disorders-epilepsy.

Functional dystonia (FD) is a disabling and diagnostically challenging functional movement disorder (FMD). We sought to identify historical predictors of FD vs. other primary dystonias (ODs) and develop a practical prediction algorithm to guide neurologists.

X-linked dystonia-parkinsonism (XDP) is a progressive adult-onset neurodegenerative disorder caused by insertion of a SINE-VNTR-Alu (SVA) retrotransposon in the TAF1 gene. The SVA retrotransposon contains a CCCTCT hexameric repeat tract of variable length, whose length is inversely correlated with age at onset. This places XDP in a broader class of repeat expansion diseases, characterized by the instability of their causative repeat mutations. Here, we observe similar inverse correlations between CCCTCT repeat length with age at onset and age at death and no obvious correlation with disease duration. To gain insight into repeat instability in XDP we performed comprehensive quantitative analyses of somatic instability of the XDP CCCTCT repeat in blood and in seventeen brain regions from affected males. Our findings reveal repeat length-dependent and expansion-based instability of the XDP CCCTCT repeat, with greater levels of expansion in brain than in blood. The brain exhibits regional-specific patterns of instability that are broadly similar across individuals, with cerebellum exhibiting low instability and cortical regions exhibiting relatively high instability. The spectrum of somatic instability in the brain includes a high proportion of moderate repeat length changes of up to 5 repeats, as well as expansions of ~ 20- > 100 repeats and contractions of ~ 20-40 repeats at lower frequencies. Comparison with HTT CAG repeat instability in postmortem Huntington's disease brains reveals similar brain region-specific profiles, indicating common trans-acting factors that contribute to the instability of both repeats. Analyses in XDP brains of expansion of a different SVA-associated CCCTCT located in the LIPG gene, and not known to be disease-associated, reveals repeat length-dependent expansion at overall lower levels relative to the XDP CCCTCT repeat, suggesting that expansion propensity may be modified by local chromatin structure. Together, the data support a role for repeat length-dependent somatic expansion in the process(es) driving the onset of XDP and prompt further investigation into repeat dynamics and the relationship to disease.

DYT-PRKRA (dystonia 16 or DYT-PRKRA) is caused by mutations in the PRKRA gene that encodes PACT, the protein activator of interferon (IFN)-induced double-stranded (ds) RNA-activated protein kinase (PKR). PACT participates in several cellular pathways, of which its role as a PKR activator protein during integrated stress response (ISR) is the best characterized. Previously, we have established that the DYT-PRKRA mutations cause enhanced activation of PKR during ISR to sensitize DYT-PRKRA cells to apoptosis. In this study, we evaluate if the most prevalent substitution mutation reported in DYT-PRKRA patients alters PACT's functional role in induction of type I IFNs via the retinoic acid-inducible gene I (RIG-I) signaling. Our results indicate that the P222L mutation augments PACT's ability to induce IFN β in response to dsRNA and the basal expression of IFN β and IFN-stimulated genes (ISGs) is higher in DYT-PRKRA patient cells compared to cells from the unaffected controls. Additionally, IFN β and ISGs are also induced at higher levels in DYT-PRKRA cells in response to dsRNA. These results offer a new avenue for investigations directed towards understanding the underlying molecular pathomechanisms in DYT-PRKRA.

Mutations in the GCH1 gene are associated with childhood onset, dopa-responsive dystonia (DRD). Correct diagnosis of DRD is crucial, given the potential for complete recovery once treated with L-dopa. The majority of DRD associated mutations lie within the coding region of the GCH1 gene, but three additional single nucleotide sequence substitutions have been reported within the 5' untranslated (5'UTR) region of the mRNA. The biologic significance of these 5'UTR GCH1 sequence substitutions has not been analyzed.

X-linked dystonia-parkinsonism (XDP) is a hereditary neurodegenerative disorder involving a progressive loss of striatal medium spiny neurons. The mechanisms underlying neurodegeneration are not known, in part because there have been few cellular models available for studying the disease. The XDP haplotype consists of multiple sequence variations in a region of the X chromosome containingTAF1, a large gene with at least 38 exons, and a multiple transcript system (MTS) composed of five unconventional exons. A previous study identified an XDP-specific insertion of a SINE-VNTR-Alu (SVA)-type retrotransposon in intron 32 ofTAF1, as well as a neural-specific TAF1 isoform, N-TAF1, which showed decreased expression in post-mortem XDP brain compared with control tissue. Here, we generated XDP patient and control fibroblasts and induced pluripotent stem cells (iPSCs) in order to further probe cellular defects associated with this disease. As initial validation of the model, we compared expression ofTAF1and MTS transcripts in XDP versus control fibroblasts and iPSC-derived neural stem cells (NSCs). Compared with control cells, XDP fibroblasts exhibited decreased expression ofTAF1transcript fragments derived from exons 32-36, a region spanning the SVA insertion site. N-TAF1, which incorporates an alternative exon (exon 34'), was not expressed in fibroblasts, but was detectable in iPSC-differentiated NSCs at levels that were ∼threefold lower in XDP cells than in controls. These results support the previous findings that N-TAF1 expression is impaired in XDP, but additionally indicate that this aberrant transcription might occur in neural cells at relatively early stages of development that precede neurodegeneration.

Genetic variation within the transcription factor TCF4 locus can cause the intellectual disability and developmental disorder Pitt-Hopkins syndrome (PTHS), whereas single-nucleotide polymorphisms within noncoding regions are associated with schizophrenia. These genetic findings position TCF4 as a link between transcription and cognition; however, the neurobiology of TCF4 remains poorly understood. Here, we quantitated multiple distinct TCF4 transcript levels in human induced pluripotent stem cell-derived neural progenitors and differentiated neurons, and PTHS patient fibroblasts. We identify two classes of pharmacological treatments that regulate TCF4 expression: WNT pathway activation and inhibition of class I histone deacetylases. In PTHS fibroblasts, both of these perturbations upregulate a subset of TCF4 transcripts. Finally, using chromatin immunoprecipitation sequencing in conjunction with genome-wide transcriptome analysis, we identified TCF4 target genes that may mediate the effect of TCF4 loss on neuroplasticity. Our studies identify new pharmacological assays, tools, and targets for the development of therapeutics for cognitive disorders.

RECQ1 is the most abundant RecQ homolog in humans but its functions have remained mostly elusive. Biochemically, RECQ1 displays distinct substrate specificities from WRN and BLM, indicating that these RecQ helicases likely perform non-overlapping functions. Our earlier work demonstrated that RECQ1-deficient cells display spontaneous genomic instability. We have obtained key evidence suggesting a unique role of RECQ1 in repair of oxidative DNA damage. We show that similar to WRN, RECQ1 associates with PARP-1 in nuclear extracts and exhibits direct protein interaction in vitro. Deficiency in WRN or BLM helicases have been shown to result in reduced homologous recombination and hyperactivation of PARP under basal condition. However, RECQ1-deficiency did not lead to PARP activation in undamaged cells and nor did it result in reduction in homologous recombination repair. In stark contrast to what is seen in WRN-deficiency, RECQ1-deficient cells hyperactivate PARP in a specific response to H₂O₂treatment. RECQ1-deficient cells are more sensitive to oxidative DNA damage and exposure to oxidative stress results in a rapid and reversible recruitment of RECQ1 to chromatin. Chromatin localization of RECQ1 precedes WRN helicase, which has been shown to function in oxidative DNA damage repair. However, oxidative DNA damage-induced chromatin recruitment of these RecQ helicases is independent of PARP activity. As other RecQ helicases are known to interact with PARP-1, this study provides a paradigm to delineate specialized and redundant functions of RecQ homologs in repair of oxidative DNA damage.

Objective: To reveal clinical characteristics of suboptimal responses to deep brain stimulation (DBS) in a multi-country DYT1 dystonia cohort. Methods: In this multi-country multi-center retrospective study, we analyzed the clinical data of DYT1 patients who experienced suboptimal responses to DBS defined as <30% improvement in dystonia scales at the last follow-up compared with baseline. We used a literature-driven historical cohort of 112 DYT1 patients for comparison. Results: Approximately 8% of our study cohort (11 out of 132) experienced suboptimal responses to DBS. Compared with the historical cohort, the multi-country cohort with suboptimal responses had a significantly younger age at onset (mean, 7.0 vs. 8.4 years; p = 0.025) and younger age at DBS (mean, 12.0 vs. 18.6 years; p = 0.019). Additionally, cranial involvement was more common in the multi-country cohort (before DBS, 64% vs. 45%, p = 0.074; before or after DBS, 91% vs. 47%, p = 0.001). Mean motor improvement at the last follow-up from baseline were 0% and 66% for the multi-country and historical cohorts, respectively. All 11 patients of the multi-country cohort had generalization of dystonia within 2.5 years after disease onset. All patients experienced dystonia improvement of >30% postoperatively; however, secondary worsening of dystonia commenced between 6 months and 3 years following DBS. The improvement at the last follow-up was less than 30% despite optimally-placed leads, a trial of multiple programming settings, and additional DBS surgeries in all patients. The on-/off-stimulation comparison at the long-term follow-up demonstrated beneficial effects of DBS despite missing the threshold of 30% improvement over baseline. Conclusion: Approximately 8% of patients represent a more aggressive phenotype of DYT1 dystonia characterized by younger age at onset, faster disease progression, and cranial involvement, which seems to be associated with long-term suboptimal responses to DBS (e.g., secondary worsening). This information could be useful for both clinicians and patients in clinical decision making and patient counseling before and following DBS implantations. Patients with this phenotype may have different neuroplasticity, neurogenetics, or possibly distinct neurophysiology.

The Shieldin complex shields double-strand DNA breaks (DSBs) from nucleolytic resection. Curiously, the penultimate Shieldin component, SHLD1, is one of the least abundant mammalian proteins. Here, we report that the transcription factors THAP1, YY1, and HCF1 bind directly to the SHLD1 promoter, where they cooperatively maintain the low basal expression of SHLD1, thereby ensuring a proper balance between end protection and resection during DSB repair. The loss of THAP1-dependent SHLD1 expression confers cross-resistance to poly (ADP-ribose) polymerase (PARP) inhibitor and cisplatin in BRCA1-deficient cells and shorter progression-free survival in ovarian cancer patients. Moreover, the embryonic lethality and PARPi sensitivity of BRCA1-deficient mice is rescued by ablation of SHLD1. Our study uncovers a transcriptional network that directly controls DSB repair choice and suggests a potential link between DNA damage and pathogenic THAP1 mutations, found in patients with the neurodevelopmental movement disorder adult-onset torsion dystonia type 6.

X-linked dystonia-parkinsonism is a neurodegenerative disorder caused by a founder retrotransposon insertion, in which a polymorphic hexanucleotide repeat accounts for ~50% of age at onset variability. Employing a genome-wide association study to identify additional factors modifying age at onset, we establish that three independent loci are significantly associated with age at onset (p < 5 × 10-8). The lead single nucleotide polymorphisms collectively account for 25.6% of the remaining variance not explained by the hexanucleotide repeat and 13.0% of the overall variance in age at onset in X-linked dystonia-parkinsonism with the protective alleles delaying disease onset by seven years. These regions harbor or lie adjacent to MSH3 and PMS2, the genes that were recently implicated in modifying age at onset in Huntington's disease, likely through a common pathway influencing repeat instability. Our work indicates the existence of three modifiers of age at onset in X-linked dystonia-parkinsonism that likely affect the DNA mismatch repair pathway.

Dystonia is a movement disorder characterized by repetitive twisting muscle contractions and postures. Its molecular pathophysiology is poorly understood, in part owing to limited knowledge of the genetic basis of the disorder. Only three genes for primary torsion dystonia (PTD), TOR1A (DYT1), THAP1 (DYT6) and CIZ1 (ref. 5), have been identified. Using exome sequencing in two families with PTD, we identified a new causative gene, GNAL, with a nonsense mutation encoding p.Ser293* resulting in a premature stop codon in one family and a missense mutation encoding p.Val137Met in the other. Screening of GNAL in 39 families with PTD identified 6 additional new mutations in this gene. Impaired function of several of the mutants was shown by bioluminescence resonance energy transfer (BRET) assays.

There has been increasing interest in the interaction of the basal ganglia with the cerebellum and the brainstem in motor control and movement disorders. In addition, it has been suggested that these subcortical connections with the basal ganglia may help to coordinate a network of regions involved in mediating posture and stabilization. While studies in animal models support a role for this circuitry in the pathophysiology of the movement disorder dystonia, thus far, there is only indirect evidence for this in humans with dystonia.

Structural variation (SV) influences genome organization and contributes to human disease. However, the complete mutational spectrum of SV has not been routinely captured in disease association studies.

Dystonia is characterized by sustained or intermittent muscle contractions resulting in abnormal, often repetitive, movements, postures, or both. Neuropathologic research has been essential in understanding the etiology and disease progression of other movement disorders, including Parkinson's disease and cerebellar ataxias. In the field of dystonia, however, research is stymied by the paucity of post-mortem tissue available and the phenotypic heterogeneity found in those with dystonia.

Somatic expansion of the Huntington's disease (HD) CAG repeat drives the rate of a pathogenic process ultimately resulting in neuronal cell death. Although mechanisms of toxicity are poorly delineated, transcriptional dysregulation is a likely contributor. To identify modifiers that act at the level of CAG expansion and/or downstream pathogenic processes, we tested the impact of genetic knockout, in HttQ111 mice, of Hdac2 or Hdac3 in medium-spiny striatal neurons that exhibit extensive CAG expansion and exquisite disease vulnerability. Both knockouts moderately attenuated CAG expansion, with Hdac2 knockout decreasing nuclear huntingtin pathology. Hdac2 knockout resulted in a substantial transcriptional response that included modification of transcriptional dysregulation elicited by the HttQ111 allele, likely via mechanisms unrelated to instability suppression. Our results identify novel modifiers of different aspects of HD pathogenesis in medium-spiny neurons and highlight a complex relationship between the expanded Htt allele and Hdac2 with implications for targeting transcriptional dysregulation in HD.

X-linked dystonia-parkinsonism is a rare neurological disease endemic to the Philippines. Dystonic symptoms appear in males at the mean age of 40 years and progress to parkinsonism with degenerative pathology in the striatum. A retrotransposon inserted in intron 32 of the TAF1 gene leads to alternative splicing in the region and a reduction of the full-length mRNA transcript.