Glial dysregulation in the human brain in fragile X-associated tremor/ataxia syndrome.

Short trinucleotide expansions at the FMR1 locus are associated with the late-onset condition fragile X-associated tremor/ataxia syndrome (FXTAS), which shows very different clinical and pathological features from fragile X syndrome (associated with longer expansions), with no clear molecular explanation for these marked differences. One prevailing theory posits that the shorter, premutation expansion uniquely causes extreme neurotoxic increases in FMR1 mRNA (i.e., four to eightfold increases), but evidence to support this hypothesis is largely derived from analysis of peripheral blood. We applied single-nucleus RNA sequencing to postmortem frontal cortex and cerebellum from 7 individuals with premutation and matched controls (n = 6) to assess cell type-specific molecular neuropathology. We found only modest upregulation (~1.3-fold) of FMR1 in some glial populations associated with premutation expansions. In premutation cases, we also identified decreased astrocyte proportions in the cortex. Differential expression and gene ontology analysis demonstrated altered neuroregulatory roles of glia. Using network analyses, we identified cell type-specific and region-specific patterns of FMR1 protein target gene dysregulation unique to premutation cases, with notable network dysregulation in the cortical oligodendrocyte lineage. We used pseudotime trajectory analysis to determine how oligodendrocyte development was altered and identified differences in early gene expression in oligodendrocyte trajectories in premutation cases specifically, implicating early cortical glial developmental perturbations. These findings challenge dogma regarding extremely elevated FMR1 increases in FXTAS and implicate glial dysregulation as a critical facet of premutation pathophysiology, representing potential unique therapeutic targets directly derived from the human condition.

FXTAS Neuropathology Includes Widespread Reactive Astrogliosis and White Matter Specific Astrocyte Degeneration.

OBJECTIVE: Fragile X-associated tremor/ataxia syndrome (FXTAS) is a late-onset progressive genetic neurodegenerative disorder that occurs in FMR1 premutation carriers. The temporal, spatial, and cell-type specific patterns of neurodegeneration in the FXTAS brain remain incompletely characterized. Intranuclear inclusion bodies are the neuropathological hallmark of FXTAS, which are largest and occur most frequently in astrocytes, glial cells that maintain brain homeostasis. Here, we characterized neuropathological alterations in astrocytes in multiple regions of the FXTAS brain. METHODS: Striatal and cerebellar sections from FXTAS cases (n = 12) and controls (n = 12) were stained for the astrocyte markers glial fibrillary acidic protein (GFAP) and aldehyde dehydrogenase 1L1 (ALDH1L1) using immunohistochemistry. Reactive astrogliosis severity, the prevalence of GFAP+ fragments, and astrocyte density were scored. Double label immunofluorescence was utilized to detect co-localization of GFAP and cleaved caspase-3. RESULTS: FXTAS cases showed widespread reactive gliosis in both grey and white matter. GFAP staining also revealed remarkably severe astrocyte pathology in FXTAS white matter - characterized by a significant and visible reduction in astrocyte density (-38.7% in striatum and - 32.2% in cerebellum) and the widespread presence of GFAP+ fragments reminiscent of apoptotic bodies. White matter specific reductions in astrocyte density were confirmed with ALDH1L1 staining. GFAP+ astrocytes and fragments in white matter were positive for cleaved caspase-3, suggesting that apoptosis-mediated degeneration is responsible for reduced astrocyte counts. INTERPRETATION: We have established that FXTAS neuropathology includes robust degeneration of astrocytes, which is specific to white matter. Because astrocytes are essential for maintaining homeostasis within the central nervous system, a loss of astrocytes likely further exacerbates neuropathological progression of other cell types in the FXTAS brain. ANN NEUROL 2024;95:558-575.

Identification of PSMB5 as a genetic modifier of fragile X-associated tremor/ataxia syndrome.

Fragile X­associated tremor/ataxia syndrome (FXTAS) is a debilitating late-onset neurodegenerative disease in premutation carriers of the expanded CGG repeat in FMR1 that presents with a spectrum of neurological manifestations, such as gait ataxia, intention tremor, and parkinsonism [P. J. Hagerman, R. J. Hagerman, Ann. N. Y. Acad. Sci. 1338, 58­70 (2015); S. Jacquemont et al., JAMA 291, 460­469 (2004)]. Here, we performed whole-genome sequencing (WGS) on male premutation carriers (CGG55­200) and prioritized candidate variants to screen for candidate genetic modifiers using a Drosophila model of FXTAS. We found 18 genes that genetically modulate CGG-associated neurotoxicity in Drosophila, such as Prosbeta5 (PSMB5), pAbp (PABPC1L), e(y)1 (TAF9), and CG14231 (OSGEPL1). Among them, knockdown of Prosbeta5 (PSMB5) suppressed CGG-associated neurodegeneration in the fly as well as in N2A cells. Interestingly, an expression quantitative trait locus variant in PSMB5, PSMB5rs11543947-A, was found to be associated with decreased expression of PSMB5 and delayed onset of FXTAS in human FMR1 premutation carriers. Finally, we demonstrate evidence that PSMB5 knockdown results in suppression of CGG neurotoxicity via both the RAN translation and RNA-mediated toxicity mechanisms, thereby presenting a therapeutic strategy for FXTAS.

Mechanistic convergence across initiation sites for RAN translation in fragile X associated tremor ataxia syndrome.

Repeat associated non-AUG (RAN) translation of CGG repeats in the 5'UTR of FMR1 produces toxic proteins that contribute to fragile X-associated tremor/ataxia syndrome (FXTAS) pathogenesis. The most abundant RAN product, FMRpolyG, initiates predominantly at an ACG upstream of the repeat. Accurate FMRpolyG measurements in FXTAS patients are lacking. We used data-dependent acquisition and parallel reaction monitoring (PRM) mass spectrometry coupled with stable isotope labeled standard peptides to identify signature FMRpolyG fragments in patient samples. Following immunoprecipitation, PRM detected FMRpolyG signature peptides in transfected cells, and FXTAS tissues and cells, but not in controls. We identified two amino-terminal peptides: an ACG-initiated Ac-MEAPLPGGVR and a GUG-initiated Ac-TEAPLPGGVR, as well as evidence for RAN translation initiation within the CGG repeat itself in two reading frames. Initiation at all sites increased following cellular stress, decreased following eIF1 overexpression and was eIF4A and M7G cap-dependent. These data demonstrate that FMRpolyG is quantifiable in human samples and FMR1 RAN translation initiates via similar mechanisms for near-cognate codons and within the repeat through processes dependent on available initiation factors and cellular environment.

RNA toxicity in non-coding repeat expansion disorders.

Several neurodegenerative disorders like amyotrophic lateral sclerosis (ALS) and spinocerebellar ataxia (SCA) are caused by non-coding nucleotide repeat expansions. Different pathogenic mechanisms may underlie these non-coding repeat expansion disorders. While gain-of-function mechanisms, such as toxicity associated with expression of repeat RNA or toxicity associated with repeat-associated non-ATG (RAN) products, are most frequently connected with these disorders, loss-of-function mechanisms have also been implicated. We review the different pathways that have been linked to non-coding repeat expansion disorders such as C9ORF72-linked ALS/frontotemporal dementia (FTD), myotonic dystrophy, fragile X tremor/ataxia syndrome (FXTAS), SCA, and Huntington's disease-like 2. We discuss modes of RNA toxicity focusing on the identity and the interacting partners of the toxic RNA species. Using the C9ORF72 ALS/FTD paradigm, we further explore the efforts and different methods used to disentangle RNA vs. RAN toxicity. Overall, we conclude that there is ample evidence for a role of RNA toxicity in non-coding repeat expansion diseases.

Association of Gene Expression and Tremor Network Structure.

BACKGROUND: Transcriptomic changes in the essential tremor (ET)-associated cerebello-thalamo-cortical "tremor network" and their association to brain structure have not been investigated. OBJECTIVE: The aim was to characterize molecular changes associated with network-level imaging-derived phenotypes (IDP) found in ET. METHODS: We performed an imaging-transcriptomic study in British adults using imaging-genome-wide association study summary statistics (UK Biobank "BIG40" cohort; n = 33,224, aged 40-69 years). We imputed imaging-transcriptomic associations for 184 IDPs and analyzed functional enrichment of gene modules and aggregate network-level phenotypes. Validation was performed in cerebellar-tissue RNA-sequencing data from ET patients and controls (n = 55). RESULTS: Among 237,896 individual predicted gene expression levels for 6063 unique genes/transcripts, we detected 2269 genome-wide significant associations (Bonferroni P < 2.102e-7, 0.95%). These were concentrated in intracellular volume fraction measures of white matter pathways and in genes with putative links to tremor (MAPT, ARL17A, KANSL1, SPPL2C, LRRC37A4P, PLEKHM1, and FMNL1). Whole-tremor-network cortical thickness was associated with a gene module linked to mitochondrial organization and protein quality control (r = 0.91, P = 2e-70), whereas white-gray T1-weighted magnetic resonance imaging (MRI) contrast in the tremor network was associated with a gene module linked to sphingolipid synthesis and ethanolamine metabolism (r = -0.90, P = 2e-68). Imputed association effect sizes and RNA-sequencing log-fold change in the validation dataset were significantly correlated for cerebellar peduncular diffusion MRI phenotypes, and there was a close overlap of significant associations between both datasets for gray matter phenotypes (χ2 = 6.40, P = 0.006). CONCLUSIONS: The identified genes and processes are potential treatment targets for ET, and our results help characterize molecular changes that could in future be used for patient treatment selection or prognosis prediction. © 2024 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Reversible encephalitis-like episodes in fragile X-associated tremor/ataxia syndrome: a case report.

BACKGROUND: Fragile X-associated tremor/ataxia syndrome (FXTAS) is a neurodegenerative disorder caused by CGG repeat expansion of FMR1 gene. Both FXTAS and neuronal intranuclear inclusion disease (NIID) belong to polyglycine diseases and present similar clinical, radiological, and pathological features, making it difficult to distinguish these diseases. Reversible encephalitis-like attacks are often observed in NIID. It is unclear whether they are presented in FXTAS and can be used for differential diagnosis of NIID and FXTAS. CASE PRESENTATION: A 63-year-old Chinese male with late-onset gait disturbance, cognitive decline, and reversible attacks of fever, consciousness impairment, dizziness, vomiting, and urinary incontinence underwent neurological assessment and examinations, including laboratory tests, electroencephalogram test, imaging, skin biopsy, and genetic test. Brain MRI showed T2 hyperintensities in middle cerebellar peduncle and cerebrum, in addition to cerebellar atrophy and DWI hyperintensities along the corticomedullary junction. Lesions in the brainstem were observed. Skin biopsy showed p62-positive intranuclear inclusions. The possibilities of hypoglycemia, lactic acidosis, epileptic seizures, and cerebrovascular attacks were excluded. Genetic analysis revealed CGG repeat expansion in FMR1 gene, and the number of repeats was 111. The patient was finally diagnosed as FXTAS. He received supportive treatment as well as symptomatic treatment during hospitalization. His encephalitic symptoms were completely relieved within one week. CONCLUSIONS: This is a detailed report of a case of FXTAS with reversible encephalitis-like episodes. This report provides new information for the possible and rare features of FXTAS, highlighting that encephalitis-like episodes are common in polyglycine diseases and unable to be used for differential diagnosis.

Mitochondrial dysfunction in Fragile X syndrome and Fragile X-associated tremor/ataxia syndrome: prospect use of antioxidants and mitochondrial nutrients.

Fragile X syndrome (FXS) is a genetic disorder characterized by mutation in the FMR1 gene, leading to the absence or reduced levels of fragile X Messenger Ribonucleoprotein 1 (FMRP). This results in neurodevelopmental deficits, including autistic spectrum conditions. On the other hand, Fragile X-associated tremor/ataxia syndrome (FXTAS) is a distinct disorder caused by the premutation in the FMR1 gene. FXTAS is associated with elevated levels of FMR1 mRNA, leading to neurodegenerative manifestations such as tremors and ataxia.Mounting evidence suggests a link between both syndromes and mitochondrial dysfunction (MDF). In this minireview, we critically examine the intricate relationship between FXS, FXTAS, and MDF, focusing on potential therapeutic avenues to counteract or mitigate their adverse effects. Specifically, we explore the role of mitochondrial cofactors and antioxidants, with a particular emphasis on alpha-lipoic acid (ALA), carnitine (CARN) and Coenzyme Q10 (CoQ10). Findings from this review will contribute to a deeper understanding of these disorders and foster novel therapeutic strategies to enhance patient outcomes.

Potential Prodromal Digital Postural Sway Markers for Fragile X-Associated Tremor/Ataxia Syndrome (FXTAS) Detected via Dual-Tasking and Sensory Manipulation.

FXTAS is a neurodegenerative disorder occurring in some Fragile X Messenger Ribonucleoprotein 1 (FMR1) gene premutation carriers (PMCs) and is characterized by cerebellar ataxia, tremor, and cognitive deficits that negatively impact balance and gait and increase fall risk. Dual-tasking (DT) cognitive-motor paradigms and challenging balance conditions may have the capacity to reveal markers of FXTAS onset. Our objectives were to determine the impact of dual-tasking and sensory and stance manipulation on balance in FXTAS and potentially detect subtle postural sway deficits in FMR1 PMCs who are asymptomatic for signs of FXTAS on clinical exam. Participants with FXTAS, PMCs without FXTAS, and controls underwent balance testing using an inertial sensor system. Stance, vision, surface stability, and cognitive demand were manipulated in 30 s trials. FXTAS participants had significantly greater total sway area, jerk, and RMS sway than controls under almost all balance conditions but were most impaired in those requiring vestibular control. PMCs without FXTAS had significantly greater RMS sway compared with controls in the feet apart, firm, single task conditions both with eyes open and closed (EC) and the feet together, firm, EC, DT condition. Postural sway deficits in the RMS postural sway variability domain in asymptomatic PMCs might represent prodromal signs of FXTAS. This information may be useful in providing sensitive biomarkers of FXTAS onset and as quantitative balance measures in future interventional trials and longitudinal natural history studies.

Apolipoproteine and KLOTHO Gene Variants Do Not Affect the Penetrance of Fragile X-Associated Tremor/Ataxia Syndrome.

In this study, the potential role and interaction of the APOε and KLOTHO genes on the penetrance of fragile X-associated tremor/ataxia syndrome (FXTAS) and on the IQ trajectory were investigated. FXTAS was diagnosed based on molecular, clinical and radiological criteria. Males with the premutation (PM) over 50 years, 165 with and 34 without an FXTAS diagnosis, were included in this study and were compared based on their APO (ε2-ε3-ε4) and KLOTHO variant (KL-VS) genotypes. The effect of APOε4 on FXTAS stage and on diagnosis did not differ significantly by KL-VS genotype with interaction effect p = 0.662 and p = 0.91, respectively. In the FXTAS individuals with an APOε2 allele, a marginal significance was observed towards a larger decline in verbal IQ (VIQ) in individuals with an APOε4 allele compared to those without an APOε4 allele (p = 0.071). In conclusion, our findings suggest that the APOε4 and KL-VS genotypes alone or through their interaction effect do not appear to predispose to either FXTAS diagnosis or stage in male carriers of the PM allele. A further study is needed to establish the trend of IQ decline in the FXTAS individuals who carry APOε4 with APOε2 compared to those without APOε4.

Repeat RNA expansion disorders of the nervous system: post-transcriptional mechanisms and therapeutic strategies.

Dozens of incurable neurological disorders result from expansion of short repeat sequences in both coding and non-coding regions of the transcriptome. Short repeat expansions underlie microsatellite repeat expansion (MRE) disorders including myotonic dystrophy (DM1, CUG50-3,500 in DMPK; DM2, CCTG75-11,000 in ZNF9), fragile X tremor ataxia syndrome (FXTAS, CGG50-200 in FMR1), spinal bulbar muscular atrophy (SBMA, CAG40-55 in AR), Huntington's disease (HD, CAG36-121 in HTT), C9ORF72- amyotrophic lateral sclerosis (ALS)/frontotemporal dementia (FTD and C9-ALS/FTD, GGGGCC in C9ORF72), and many others, like ataxias. Recent research has highlighted several mechanisms that may contribute to pathology in this heterogeneous class of neurological MRE disorders - bidirectional transcription, intranuclear RNA foci, and repeat associated non-AUG (RAN) translation - which are the subject of this review. Additionally, many MRE disorders share similar underlying molecular pathologies that have been recently targeted in experimental and preclinical contexts. We discuss the therapeutic potential of versatile therapeutic strategies that may selectively target disrupted RNA-based processes and may be readily adaptable for the treatment of multiple MRE disorders. Collectively, the strategies under consideration for treatment of multiple MRE disorders include reducing levels of toxic RNA, preventing RNA foci formation, and eliminating the downstream cellular toxicity associated with peptide repeats produced by RAN translation. While treatments are still lacking for the majority of MRE disorders, several promising therapeutic strategies have emerged and will be evaluated within this review.

Dissecting the roles of EIF4G homologs reveals DAP5 as a modifier of CGG repeat-associated toxicity in a Drosophila model of FXTAS.

Neurodegeneration in Fragile X-associated tremor/ataxia syndrome (FXTAS) is caused by a CGG trinucleotide repeat expansion in the 5' UTR of FMR1. Expanded CGG repeat RNAs form stable secondary structures, which in turn support repeat-associated non-AUG (RAN) translation to produce toxic peptides. The parameters that impact RAN translation initiation efficiency are not well understood. Here we used a Drosophila melanogaster model of FXTAS to evaluate the role of the eIF4G family of eukaryotic translation initiation factors (EIF4G1, EIF4GII and EIF4G2/DAP5) in modulating RAN translation and CGG repeat-associated toxicity. DAP5 knockdown robustly suppressed CGG repeat-associated toxicity and inhibited RAN translation. Furthermore, knockdown of initiation factors that preferentially associate with DAP5 (such as EIF2ß, EIF3F and EIF3G) also selectively suppressed CGG repeat-induced eye degeneration. In mammalian cellular reporter assays, DAP5 knockdown exhibited modest and cell-type specific effects on RAN translation. Taken together, these data support a role for DAP5 in CGG repeat associated toxicity possibly through modulation of RAN translation.

Small molecule 1a reduces FMRpolyG-mediated toxicity in in vitro and in vivo models for FMR1 premutation.

Fragile X-associated tremor and ataxia syndrome (FXTAS) is a late-onset, progressive neurodegenerative disorder characterized by tremors, ataxia and neuropsychological problems. This disease is quite common in the general population with approximately 20 million carriers worldwide. The risk of developing FXTAS increases dramatically with age, with about 45% of male carriers over the age of 50 being affected. FXTAS is caused by a CGG-repeat expansion (CGGexp) in the fragile X mental retardation 1 (FMR1) gene. CGGexp RNA is translated into the FMRpolyG protein by a mechanism called RAN translation. Although both gene and pathogenic trigger are known, no therapeutic interventions are available at this moment. Here, we present, for the first time, primary hippocampal neurons derived from the ubiquitous inducible mouse model which is used as a screening tool for targeted interventions. A promising candidate is the repeat binding, RAN translation blocking, small molecule 1a. Small molecule 1a shields the disease-causing CGGexp from being translated into the toxic FMRpolyG protein. Primary hippocampal neurons formed FMRpolyG-positive inclusions, and upon treatment with 1a, the numbers of FMRpolyG-positive inclusions are reduced. We also describe for the first time the formation of FMRpolyG-positive inclusions in the liver of this mouse model. Treatment with 1a reduced the insoluble FMRpolyG protein fraction in the liver but not the number of inclusions. Moreover, 1a treatment had a reducing effect on the number of Rad23b-positive inclusions and insoluble Rad23b protein levels. These data suggest that targeted small molecule therapy is effective in an FXTAS mouse model and has the potential to treat CGGexp-mediated diseases, including FXTAS.

Ectopic expression of CGG-repeats alters ovarian response to gonadotropins and leads to infertility in a murine FMR1 premutation model.

Women heterozygous for an expansion of CGG repeats in the 5'UTR of FMR1 risk developing fragile X-associated primary ovarian insufficiency (FXPOI) and/or tremor and ataxia syndrome (FXTAS). We show that expanded CGGs, independent of FMR1, are sufficient to drive ovarian insufficiency and that expression of CGG-containing mRNAs alone or in conjunction with a polyglycine-containing peptide translated from these RNAs contribute to dysfunction. Heterozygous females from two mouse lines expressing either CGG RNA-only (RNA-only) or CGG RNA and the polyglycine product FMRpolyG (FMRpolyG+RNA) were used to assess ovarian function in aging animals. The expression of FMRpolyG+RNA led to early cessation of breeding, ovulation and transcriptomic changes affecting cholesterol and steroid hormone biosynthesis. Females expressing CGG RNA-only did not exhibit decreased progeny during natural breeding, but their ovarian transcriptomes were enriched for alterations in cholesterol and lipid biosynthesis. The enrichment of CGG RNA-only ovaries for differentially expressed genes related to cholesterol processing provided a link to the ovarian cysts observed in both CGG-expressing lines. Early changes in transcriptome profiles led us to measure ovarian function in prepubertal females that revealed deficiencies in ovulatory responses to gonadotropins. These include impairments in cumulus expansion and resumption of oocyte meiosis, as well as reduced ovulated oocyte number. Cumulatively, we demonstrated the sufficiency of ectopically expressed CGG repeats to lead to ovarian insufficiency and that co-expression of CGG-RNA and FMRpolyG lead to premature cessation of breeding. However, the expression of CGG RNA-alone was sufficient to lead to ovarian dysfunction by impairing responses to hormonal stimulation.

Fragile X premutation rCGG repeats impair synaptic growth and synaptic transmission at Drosophila larval neuromuscular junction.

Fragile X-associated tremor/ataxia syndrome (FXTAS) is a late-onset neurodegenerative disease that develops in some premutation (PM) carriers of the FMR1 gene with alleles bearing 55-200 CGG repeats. The discovery of a broad spectrum of clinical and cell-developmental abnormalities among PM carriers with or without FXTAS and in model systems suggests that neurodegeneration seen in FXTAS could be the inevitable end-result of pathophysiological processes set during early development. Hence, it is imperative to trace early PM-induced pathological abnormalities. Previous studies have shown that transgenic Drosophila carrying PM-length CGG repeats are sufficient to cause neurodegeneration. Here, we used the same transgenic model to understand the effect of CGG repeats on the structure and function of the developing nervous system. We show that presynaptic expression of CGG repeats restricts synaptic growth, reduces the number of synaptic boutons, leads to aberrant presynaptic varicosities, and impairs synaptic transmission at the larval neuromuscular junctions. The postsynaptic analysis shows that both glutamate receptors and subsynaptic reticulum proteins were normal. However, a high percentage of boutons show a reduced density of Bruchpilot protein, a key component of presynaptic active zones required for vesicle release. The electrophysiological analysis shows a significant reduction in quantal content, a measure of total synaptic vesicles released per excitation potential. Together, these findings suggest that synapse perturbation caused by riboCGG (rCGG) repeats mediates presynaptically during larval neuromuscular junction development. We also suggest that the stress-activated c-Jun N-terminal kinase protein Basket and CIDE-N protein Drep-2 positively mediate Bruchpilot active zone defects caused by rCGG repeats.

Recent advances in CGG repeat diseases and a proposal of fragile X-associated tremor/ataxia syndrome, neuronal intranuclear inclusion disease, and oculophryngodistal myopathy (FNOP) spectrum disorder.

While whole genome sequencing and long-read sequencing have become widely available, more and more focuses are on noncoding expanded repeats. Indeed, more than half of noncoding repeat expansions related to diseases have been identified in the five years. An exciting aspect of the progress in this field is an identification of a phenomenon called repeat motif-phenotype correlation. Repeat motif-phenotype correlation in noncoding repeat expansion diseases is first found in benign adult familial myoclonus epilepsy. The concept is extended in the research of CGG repeat expansion diseases. In this review, we focus on newly identified CGG repeat expansion diseases, update the concept of repeat motif-phenotype correlation in CGG repeat expansion diseases, and propose a clinical concept of FNOP (fragile X-associated tremor/ataxia syndrome, neuronal intranuclear inclusion disease, and oculopharyngodistal myopathy)-spectrum disorder, which shares clinical features and thus probably share some common disease pathophysiology, to further facilitate discussion and progress in this field.

Atypical vocal quality in women with the FMR1 premutation: an indicator of impaired sensorimotor control.

Women with the FMR1 premutation are susceptible to motor involvement related to atypical cerebellar function, including risk for developing fragile X tremor ataxia syndrome. Vocal quality analyses are sensitive to subtle differences in motor skills but have not yet been applied to the FMR1 premutation. This study examined whether women with the FMR1 premutation demonstrate differences in vocal quality, and whether such differences relate to FMR1 genetic, executive, motor, or health features of the FMR1 premutation. Participants included 35 women with the FMR1 premutation and 45 age-matched women without the FMR1 premutation who served as a comparison group. Three sustained /a/ vowels were analyzed for pitch (mean F0), variability of pitch (standard deviation of F0), and overall vocal quality (jitter, shimmer, and harmonics-to-noise ratio). Executive, motor, and health indices were obtained from direct and self-report measures and genetic samples were analyzed for FMR1 CGG repeat length and activation ratio. Women with the FMR1 premutation had a lower pitch, larger pitch variability, and poorer vocal quality than the comparison group. Working memory was related to harmonics-to-noise ratio and shimmer in women with the FMR1 premutation. Vocal quality abnormalities differentiated women with the FMR1 premutation from the comparison group and were evident even in the absence of other clinically evident motor deficits. This study supports vocal quality analyses as a tool that may prove useful in the detection of early signs of motor involvement in this population.

Between Order and Chaos: Understanding the Mechanism and Pathology of RAN Translation.

Repeat-associated non-AUG (RAN) translation is a pathogenic mechanism in which repetitive sequences are translated into aggregation-prone proteins from multiple reading frames, even without a canonical AUG start codon. Since its discovery in spinocerebellar ataxia type 8 (SCA8) and myotonic dystrophy type 1 (DM1), RAN translation is now known to occur in the context of 12 disease-linked repeat expansions. This review discusses recent advances in understanding the regulatory mechanisms controlling RAN translation and its contribution to the pathophysiology of repeat expansion diseases. We discuss the key findings in the context of Fragile X Tremor Ataxia Syndrome (FXTAS), a neurodegenerative disorder caused by a CGG repeat expansion in the 5' untranslated region of FMR1.

Men with FMR1 premutation alleles of less than 71 CGG repeats have low risk of being affected with fragile X-associated tremor/ataxia syndrome (FXTAS).

Fragile X-associated tremor/ataxia syndrome (FXTAS) is a late-onset condition characterised by cerebellar ataxia and intention tremor, usually found in individuals with FMR1 premutation alleles (PM-CGG expansion of 55-199 repeats). Population studies estimate that between 1 in 250 and 1 in 1600 men have a PM, with up to 45% of these men suggested to develop FXTAS by age 80. We used a Bayesian approach to compare the probability of finding a specific PM genotype in an ataxia population to a population control group and found an estimated penetrance of <1% (0.031%; CI 0.007% to 0.141%) for men with ≤70 CGGs. These findings suggest that men with a PM of ≤70 CGGs, who comprise the vast majority of those with a PM, have a much lower risk of being affected with FXTAS than previously suggested. This is an issue of growing importance for accurate genetic counselling, as those with a PM of ≤70 CGGs are increasingly detected through community carrier screening or neurodevelopmental assessment programmes.

Cyclic mismatch binding ligands interact with disease-associated CGG trinucleotide repeats in RNA and suppress their translation.

Fragile X-associated tremor/ataxia syndrome (FXTAS) is a late-onset neurodegenerative disorder caused by a limited expansion of CGG repeats in the FMR1 gene. Degeneration of neurons in FXTAS cell models can be triggered by accumulation of polyglycine protein (FMRpolyG), a by-product of translation initiated upstream to the repeats. Specific aims of our work included testing if naphthyridine-based molecules could (i) block FMRpolyG synthesis by binding to CGG repeats in RNA, (ii) reverse pathological alterations in affected cells and (iii) preserve the content of FMRP, translated from the same FMR1 mRNA. We demonstrate that cyclic mismatch binding ligand CMBL4c binds to RNA structure formed by CGG repeats and attenuates translation of FMRpolyG and formation of nuclear inclusions in cells transfected with vectors expressing RNA with expanded CGG repeats. Moreover, our results indicate that CMBL4c delivery can reduce FMRpolyG-mediated cytotoxicity and apoptosis. Importantly, its therapeutic potential is also observed once the inclusions are already formed. We also show that CMBL4c-driven FMRpolyG loss is accompanied by partial FMRP reduction. As complete loss of FMRP induces FXS in children, future experiments should aim at evaluation of CMBL4c therapeutic intervention in differentiated tissues, in which FMRpolyG translation inhibition might outweigh adverse effects related to FMRP depletion.