Posttranscriptional regulation of FAN1 by miR-124-3p at rs3512 underlies onset-delaying genetic modification in Huntington's disease.

Many Mendelian disorders, such as Huntington's disease (HD) and spinocerebellar ataxias, arise from expansions of CAG trinucleotide repeats. Despite the clear genetic causes, additional genetic factors may influence the rate of those monogenic disorders. Notably, genome-wide association studies discovered somewhat expected modifiers, particularly mismatch repair genes involved in the CAG repeat instability, impacting age at onset of HD. Strikingly, FAN1, previously unrelated to repeat instability, produced the strongest HD modification signals. Diverse FAN1 haplotypes independently modify HD, with rare genetic variants diminishing DNA binding or nuclease activity of the FAN1 protein, hastening HD onset. However, the mechanism behind the frequent and the most significant onset-delaying FAN1 haplotype lacking missense variations has remained elusive. Here, we illustrated that a microRNA acting on 3'-UTR (untranslated region) SNP rs3512, rather than transcriptional regulation, is responsible for the significant FAN1 expression quantitative trait loci signal and allelic imbalance in FAN1 messenger ribonucleic acid (mRNA), accounting for the most significant and frequent onset-delaying modifier haplotype in HD. Specifically, miR-124-3p selectively targets the reference allele at rs3512, diminishing the stability of FAN1 mRNA harboring that allele and consequently reducing its levels. Subsequent validation analyses, including the use of antagomir and 3'-UTR reporter vectors with swapped alleles, confirmed the specificity of miR-124-3p at rs3512. Together, these findings indicate that the alternative allele at rs3512 renders the FAN1 mRNA less susceptible to miR-124-3p-mediated posttranscriptional regulation, resulting in increased FAN1 levels and a subsequent delay in HD onset by mitigating CAG repeat instability.

Huntington's disease: A clinical primer for acute and general physicians.

Huntington's disease (HD) usually manifests in adulthood and is characterised by progressive neurodegeneration in the brain that causes worsening involuntary movements, mental health and cognition over many years. Depression, anxiety and apathy are common. HD is autosomal dominant and affects about 1 in 8,000 people in the UK. There are currently no disease-modifying treatments and so patient care centres on multidisciplinary therapy support and medical treatments to relieve distressing symptoms. Progression of HD is usually slow, and so acute deteriorations often indicate another problem, such as intercurrent infections, constipation, urinary retention, gastro-oesophageal reflux disease or poor dentition. In this review we outline common presentations in HD patients, both acute and chronic, consider therapeutic options and discuss specific considerations in advanced HD.

Modification of Huntington's disease by short tandem repeats.

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.

Mutant huntingtin confers cell-autonomous phenotypes on Huntington's disease iPSC-derived microglia.

Huntington's disease (HD) is a neurodegenerative disorder caused by a dominantly inherited CAG repeat expansion in the huntingtin gene (HTT). Neuroinflammation and microglia have been implicated in HD pathology, however it has been unclear if mutant HTT (mHTT) expression has an adverse cell-autonomous effect on microglial function, or if they are only activated in response to the neurodegenerative brain environment in HD. To establish a human cell model of HD microglia function, we generated isogenic controls for HD patient-derived induced pluripotent stem cells (iPSC) with 109 CAG repeats (Q109). Q109 and isogenic Q22 iPSC, as well as non-isogenic Q60 and Q33 iPSC lines, were differentiated to iPSC-microglia. Our study supports a model of basal microglia dysfunction in HD leading to elevated pro-inflammatory cytokine production together with impaired phagocytosis and endocytosis capacity, in the absence of immune stimulation. These findings are consistent with early microglia activation observed in pre-manifest patients and indicate that mHTT gene expression affects microglia function in a cell-autonomous way.

Repeat Detector: versatile sizing of expanded tandem repeats and identification of interrupted alleles from targeted DNA sequencing.

Targeted DNA sequencing approaches will improve how the size of short tandem repeats is measured for diagnostic tests and preclinical studies. The expansion of these sequences causes dozens of disorders, with longer tracts generally leading to a more severe disease. Interrupted alleles are sometimes present within repeats and can alter disease manifestation. Determining repeat size mosaicism and identifying interruptions in targeted sequencing datasets remains a major challenge. This is in part because standard alignment tools are ill-suited for repetitive and unstable sequences. To address this, we have developed Repeat Detector (RD), a deterministic profile weighting algorithm for counting repeats in targeted sequencing data. We tested RD using blood-derived DNA samples from Huntington's disease and Fuchs endothelial corneal dystrophy patients sequenced using either Illumina MiSeq or Pacific Biosciences single-molecule, real-time sequencing platforms. RD was highly accurate in determining repeat sizes of 609 blood-derived samples from Huntington's disease individuals and did not require prior knowledge of the flanking sequences. Furthermore, RD can be used to identify alleles with interruptions and provide a measure of repeat instability within an individual. RD is therefore highly versatile and may find applications in the diagnosis of expanded repeat disorders and in the development of novel therapies.

Huntington's disease age at motor onset is modified by the tandem hexamer repeat in TCERG1.

Huntington's disease is caused by an expanded CAG tract in HTT. The length of the CAG tract accounts for over half the variance in age at onset of disease, and is influenced by other genetic factors, mostly implicating the DNA maintenance machinery. We examined a single nucleotide variant, rs79727797, on chromosome 5 in the TCERG1 gene, previously reported to be associated with Huntington's disease and a quasi-tandem repeat (QTR) hexamer in exon 4 of TCERG1 with a central pure repeat. We developed a method for calling perfect and imperfect repeats from exome-sequencing data, and tested association between the QTR in TCERG1 and residual age at motor onset (after correcting for the effects of CAG length in the HTT gene) in 610 individuals with Huntington's disease via regression analysis. We found a significant association between age at onset and the sum of the repeat lengths from both alleles of the QTR (p = 2.1 × 10-9), with each added repeat hexamer reducing age at onset by one year (95% confidence interval [0.7, 1.4]). This association explained that previously observed with rs79727797. The association with age at onset in the genome-wide association study is due to a QTR hexamer in TCERG1, translated to a glutamine/alanine tract in the protein. We could not distinguish whether this was due to cis-effects of the hexamer repeat on gene expression or of the encoded glutamine/alanine tract in the protein. These results motivate further study of the mechanisms by which TCERG1 modifies onset of HD.

Exome sequencing of individuals with Huntington's disease implicates FAN1 nuclease activity in slowing CAG expansion and disease onset.

The age at onset of motor symptoms in Huntington's disease (HD) is driven by HTT CAG repeat length but modified by other genes. In this study, we used exome sequencing of 683 patients with HD with extremes of onset or phenotype relative to CAG length to identify rare variants associated with clinical effect. We discovered damaging coding variants in candidate modifier genes identified in previous genome-wide association studies associated with altered HD onset or severity. Variants in FAN1 clustered in its DNA-binding and nuclease domains and were associated predominantly with earlier-onset HD. Nuclease activities of purified variants in vitro correlated with residual age at motor onset of HD. Mutating endogenous FAN1 to a nuclease-inactive form in an induced pluripotent stem cell model of HD led to rates of CAG expansion similar to those observed with complete FAN1 knockout. Together, these data implicate FAN1 nuclease activity in slowing somatic repeat expansion and hence onset of HD.

Genetic modifiers of Huntington disease differentially influence motor and cognitive domains.

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.

Neurological consultation with an autistic patient.

Autism is a neurodevelopmental condition with a very heterogeneous presentation. Autistic people are more likely to have unmet healthcare needs, making it essential that healthcare professionals are 'autism-aware'. In this article, we provide an overview of how autism presents and use case studies to illustrate how a neurological consultation in an outpatient clinic environment could prove challenging for a autistic person. We suggest how to improve communication with autistic patients in clinic and highlight the importance of a patient-centred and flexible approach.

Timing and Impact of Psychiatric, Cognitive, and Motor Abnormalities in Huntington Disease.

OBJECTIVE: To assess the prevalence, timing, and functional impact of psychiatric, cognitive, and motor abnormalities in Huntington disease (HD) gene carriers, we analyzed retrospective clinical data from individuals with manifest HD. METHODS: Clinical features of patients with HD were analyzed for 6,316 individuals in an observational study of the European Huntington's Disease Network (REGISTRY) from 161 sites across 17 countries. Data came from clinical history and the patient-completed Clinical Characteristics Questionnaire that assessed 8 symptoms: motor, cognitive, apathy, depression, perseverative/obsessive behavior, irritability, violent/aggressive behavior, and psychosis. Multiple logistic regression was used to analyze relationships between symptoms and functional outcomes. RESULTS: The initial manifestation of HD is increasingly likely to be motor and less likely to be psychiatric as age at presentation increases and is independent of pathogenic CAG repeat length. The Clinical Characteristics Questionnaire captures data on nonmotor symptom prevalence that correlate specifically with validated clinical measures. Psychiatric and cognitive symptoms are common in HD gene carriers, with earlier onsets associated with longer CAG repeats. Of patients with HD, 42.4% reported at least 1 psychiatric or cognitive symptom before motor symptoms, with depression most common. Each nonmotor symptom was associated with significantly reduced total functional capacity scores. CONCLUSIONS: Psychiatric and cognitive symptoms are common and functionally debilitating in HD gene carriers. They require recognition and targeting with clinical outcome measures and treatments. However, because it is impossible to distinguish confidently between nonmotor symptoms arising from HD and primary psychiatric disorders, particularly in younger premanifest patients, nonmotor symptoms should not be used to make a clinical diagnosis of HD. TRIAL REGISTRATION INFORMATION: ClinicalTrials.gov Identifier: NCT01590589.

Dentatorubral-pallidoluysian Atrophy: An Update.

Background: Dentatorubral-pallidoluysian atrophy (DRPLA) is a rare, autosomal dominantly inherited disorder characterized by myoclonus, epilepsy, ataxia, and dementia. Diagnosis is challenging due to the heterogeneous presentation and symptomatic overlap with other spinocerebellar ataxias. Symptoms vary according to age of onset, with a mean age at onset of 31 years. A CAG repeat expansion in the ATN1 gene results in neuronal intranuclear inclusions, variable neuronal loss, and astrocytosis in the globus pallidus, dentate and red nuclei. No disease-modifying or curative treatments are currently available. Methods: We performed an online literature search using PubMed for all articles published in an English Language format on the topics of DRPLA or ATN1 over the last 10 years. Where these articles cited other research as support for findings, or statements, these articles were also reviewed. Contemporary articles from related research fields (e.g., Huntington's Disease) were also included to support statements. Results: Forty-seven articles were identified, 10 were unobtainable and 10 provided no relevant information. The remaining 27 articles were then used for the review template: seven case reports, seven case series, six model system articles (one review article), four population clinical and genetic studies (one review article), two general review articles, and one human gene expression study. Other cited articles or research from related fields gave a further 42 articles, producing a total of 69 articles cited: 15 case series (including eight family studies), 14 model systems (one review article), 14 population clinical and genetic studies (two review articles), 10 case reports, eight clinical trials/guidelines, four genetic methodology articles, three general review articles, and one human gene expression study. Discussion: DRPLA remains an intractable, progressive, neurodegenerative disorder without effective treatment. Early recognition of the disorder may improve patient understanding, and access to services and treatments. Large-scale studies are lacking, but are required to characterize the full allelic architecture of the disorder in all populations and the heterogeneous phenotypic spectrum, including neuroimaging findings, possible biomarkers, and responses to treatment.

The central role of DNA damage and repair in CAG repeat diseases.

Diseases such as Huntington's disease and certain spinocerebellar ataxias are caused by the expansion of genomic cytosine-adenine-guanine (CAG) trinucleotide repeats beyond a specific threshold. These diseases are all characterised by neurological symptoms and central neurodegeneration, but our understanding of how expanded repeats drive neuronal loss is incomplete. Recent human genetic evidence implicates DNA repair pathways, especially mismatch repair, in modifying the onset and progression of CAG repeat diseases. Repair pathways might operate directly on repeat sequences by licensing or inhibiting repeat expansion in neurons. Alternatively, or in addition, because many of the genes containing pathogenic CAG repeats encode proteins that themselves have roles in the DNA damage response, it is possible that repeat expansions impair specific DNA repair pathways. DNA damage could then accrue in neurons, leading to further expansion at repeat loci, thus setting up a vicious cycle of pathology. In this review, we consider DNA damage and repair pathways in postmitotic neurons in the context of disease-causing CAG repeats. Investigating and understanding these pathways, which are clearly relevant in promoting and ameliorating disease in humans, is a research priority, as they are known to modify disease and therefore constitute prevalidated drug targets.

Genetic modifiers of Mendelian disease: Huntington's disease and the trinucleotide repeat disorders.

In the decades since the genes and mutations associated with the commoner Mendelian disorders were first discovered, technological advances in genetic analysis have made finding genomic variation a much less onerous task. Recently, the global efforts to collect subjects with Mendelian disorders, to better define the disorders and to empower appropriate clinical trials, along with improved genetic technologies, have allowed the identification of genetic variation that does not cause disease, but substantially modifies disease presentation. The advantage of this is it identifies biological pathways and molecules, that, if modified in people, might alter disease presentation. In Huntington's disease (HD), caused by an expanded CAG repeat tract in HTT, genetic variation has been uncovered that is associated with change in the onset or progression of disease. Some of this variation lies in genes that are part of the DNA damage response, previously suggested to be important in modulating expansion of the repeat tract in germline and somatic cells. The genetic evidence implicates a DNA damage response-related pathway in modulating the pathogenicity of the repeat tracts in HD, and possibly, in other trinucleotide repeat disorders. These findings offer new targets for drug development in these currently intractable disorders.

Nitrous oxide misuse and vitamin B12 deficiency.

A 36-year-old man presented to hospital with a 5-week history of ascending limb paraesthesiae and balance difficulties. He had no medical or travel history of note, but admitted habitual nitrous oxide (N2O) inhalation. Neurological examination revealed a sensory ataxia with pseudoathetosis in the upper limbs and reduced vibration sensation to the hips bilaterally. Significant investigation results included a low serum vitamin B12 concentration, mild macrocytosis and raised serum homocysteine concentration. T2 MRI of the spinal cord demonstrated increased signal extending from C1 to T11 in keeping with a longitudinal myelitis. The patient was diagnosed with a myeloneuropathy secondary to vitamin B12 deficiency, resulting from heavy N2O inhalation. He was treated with intramuscular vitamin B12 injections and received regular physiotherapy. At discharge, he was able to mobilise short distances with the aid of a zimmer frame, and was independently mobile 8 weeks later.

Double-stranded DNA translocation: structure and mechanism of hexameric FtsK.

FtsK is a DNA translocase that coordinates chromosome segregation and cell division in bacteria. In addition to its role as activator of XerCD site-specific recombination, FtsK can translocate double-stranded DNA (dsDNA) rapidly and directionally and reverse direction. We present crystal structures of the FtsK motor domain monomer, showing that it has a RecA-like core, the FtsK hexamer, and also showing that it is a ring with a large central annulus and a dodecamer consisting of two hexamers, head to head. Electron microscopy (EM) demonstrates the DNA-dependent existence of hexamers in solution and shows that duplex DNA passes through the middle of each ring. Comparison of FtsK monomer structures from two different crystal forms highlights a conformational change that we propose is the structural basis for a rotary inchworm mechanism of DNA translocation.

Asymmetric activation of Xer site-specific recombination by FtsK.

Chromosome dimers, which frequently form in Escherichia coli, are resolved by the combined action of two tyrosine recombinases, XerC and XerD, acting at a specific site on the chromosome, dif, together with the cell division protein FtsK. The C-terminal domain of FtsK (FtsK(C)) is a DNA translocase implicated in helping synapsis of the dif sites and in locally promoting XerD strand exchanges after synapse formation. Here we show that FtsK(C) ATPase activity is directly involved in the local activation of Xer recombination at dif, by using an intermolecular recombination assay that prevents significant DNA translocation, and we confirm that FtsK acts before Holliday junction formation. We show that activation only occurs with a DNA segment adjacent to the XerD-binding site of dif. Only one such DNA extension is required. Taken together, our data suggest that FtsK needs to contact the XerD recombinase to switch its activity on using ATP hydrolysis.