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Vascular pathology of donor one and two. Brown iron-like deposits are seen in both donors (A,E), which stain blue in a Perls staining (B,F). Both donors showed abnormal increased iron deposits in the ventral part of the substantia nigra (C,G). Throughout the brain, inclusions were seen in the endothelial cells (black arrowhead) and the pericytes (white arrowhead) (D,H). Scale bar A,B,D,E,F,H is 10 μm, scale bar C,G is 0.5 cm.
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CGG repeat expansions within the premutation range (55-200) of the FMR1 gene can lead to Fragile-X associated tremor/ataxia syndrome and Fragile-X associated neuropsychiatric disorders. These CGG repeats are translated into a toxic polyglycine-containing protein, FMRpolyG. Pathology of Fragile-X associated tremor/ataxia syndrome and Fragile-X assoc...
Citations
... These findings indicate that all brains with FXTAS show MRI changes consistent with ischemia, although hemorrhages could occur concurrently or even were the predominant injury in about 24% of the brains. Consistent with our findings, a recently published study [47] reported p62positive intranuclear inclusions in the pericytes and endothelial cells of brain vasculature, as well as vascular infarcts such as lacunae and strokes throughout the brain and iron deposits resulting from disrupted vasculature throughout the cerebral cortex and hippocampus, in two male premutation carriers. These two men displayed mild motor impairments, no MCP sign or confluent WMHs, but prominent clinical symptoms of fragile X-associated neuropsychiatric disorders (FXAND), including apathy, aggression and depression [48]. ...
... These two men displayed mild motor impairments, no MCP sign or confluent WMHs, but prominent clinical symptoms of fragile X-associated neuropsychiatric disorders (FXAND), including apathy, aggression and depression [48]. In addition, the toxic polyglycine-containing protein, FMRpolyG protein [49], was also detected throughout the brain and brain vasculature [47], suggesting that compromised inclusion-bearing vasculature can be an important feature for both FXTAS and FXAND [19]. Brain regions showing significant atrophy in FXTAS cases were the cerebellar white matter, red nucleus and dentate nucleus (Table 3), among which atrophy of the red nucleus and dentate nucleus have not been reported in FXTAS. ...
Brain changes at the end-stage of fragile X-associated tremor/ataxia syndrome (FXTAS) are largely unknown due to mobility impairment. We conducted a postmortem MRI study of FXTAS to quantify cerebrovascular disease, brain atrophy and iron content, and examined their relationships using principal component analysis (PCA). Intracranial hemorrhage (ICH) was observed in 4/17 FXTAS cases, among which one was confirmed by histologic staining. Compared with seven control brains, FXTAS cases showed higher ratings of T2-hyperintensities (indicating cerebral small vessel disease) in the cerebellum, globus pallidus and frontoparietal white matter, and significant atrophy in the cerebellar white matter, red nucleus and dentate nucleus. PCA of FXTAS cases revealed negative associations of T2-hyperintensity ratings with anatomic volumes and iron content in the white matter, hippocampus and amygdala, that were independent from a highly correlated number of regions with ICH and iron content in subcortical nuclei. Post-hoc analysis confirmed PCA findings and further revealed increased iron content in the white matter, hippocampus and amygdala in FXTAS cases compared to controls, after adjusting for T2-hyperintensity ratings. These findings indicate that both ischemic and hemorrhagic brain damage may occur in FXTAS, with the former being marked by demyelination/iron depletion and atrophy, and the latter by ICH and iron accumulation in basal ganglia.
... Consistent with our ndings, a recently published study 31 reported p62-positive intranuclear inclusions in the pericytes and endothelial cells of brain vasculature as well as vascular infarcts such as lacunae and strokes throughout the brain and iron deposits resulting from disrupted vasculature throughout the cerebral cortex and hippocampus in two male premutation carriers. These two men displayed mild motor impairments, no MCP sign or con uent WMHs, but prominent clinical symptoms of fragile X-associated neuropsychiatric disorders (FXAND) including apathy, aggression, and depression 32 . In addition, the toxic polyglycine-containing protein, FMRpolyG protein 33 , was also detected throughout the brain and brain vasculature 31 symptomology. ...
Brain changes at end-stage of fragile X-associated tremor/ataxia syndrome (FXTAS) are largely unknown due to mobility impairment. We conducted a postmortem MRI study of FXTAS to quantify cerebrovascular disease, brain atrophy, and iron content and examined their relationships using principal component analysis (PCA). Intracranial hemorrhage (ICH) was observed in 4/17 FXTAS cases among which one was confirmed by histologic staining. Compared with seven control brains, FXTAS cases showed higher ratings of T2-hyperintensities (indicating cerebral small vessel disease) in the cerebellum, globus pallidus, and frontoparietal white matter and significant atrophy in cerebellar white matter, red nucleus, and dentate nucleus. PCA of FXTAS cases revealed negative associations of T2-hyperintensity ratings with anatomic volumes and iron content in the white matter, hippocampus, and amygdala, that were independent from highly correlated number of regions with ICH and iron content in subcortical nuclei. Post hoc analysis confirmed PCA findings and further revealed increased iron content in the white matter, hippocampus, and amygdala in FXTAS cases than controls after adjusting for T2-hyperintensity ratings. These findings indicate that both ischemic and hemorrhagic brain damage may occur in FXTAS, with the former marked by demyelination/iron depletion and atrophy and the latter, ICH and iron accumulation in basal ganglia.
... 35,36 In the case of FMR1, expanded CGG repeats are predominantly translated into a polyglycinecontaining protein, named FMRpolyG, which forms ubiquitin-positive intranuclear inclusions and which expression is toxic for neurons in cell and animal models. [37][38][39] Intranuclear inclusions of FMRpolyG have been detected in the brains of FXTAS patients, 37,[39][40][41] as well as in non-CNS tissues, 42 including the ovarian stroma of a woman with FXPOI, 43 and in mural granulosa cells from six FMR1 premutation carriers. 44 Furthermore, FMRpolyG protein has been detected in the ovarian stroma of mice expressing expanded CGG repeats. ...
... This suggests that the CGG RNA aggregates and FMRpolyG expression observed are not artefactual but biologically meaningful. 40,41,54 It is possible that the proportion of p62 positive CGG/FMRpolyG positive cells may increase at 72 h post transfection, however this was not explored further due to low cell viability at later time points. ...
Fragile X‐associated premature ovarian insufficiency (FXPOI) is among a family of disorders caused by expansion of a CGG trinucleotide repeat sequence located in the 5′ untranslated region (UTR) of the fragile X messenger ribonucleoprotein 1 (FMR1) gene on the X chromosome. Women with FXPOI have a depleted ovarian follicle population, resulting in amenorrhea, hypoestrogenism, and loss of fertility before the age of 40. FXPOI is caused by expansions of the CGG sequence to lengths between 55 and 200 repeats, known as a FMRI premutation, however the mechanism by which the premutation drives disease pathogenesis remains unclear. Two main hypotheses exist, which describe an mRNA toxic gain‐of‐function mechanism or a protein‐based mechanism, where repeat‐associated non‐AUG (RAN) translation results in the production of an abnormal protein, called FMRpolyG. Here, we have developed an in vitro granulosa cell model of the FMR1 premutation by ectopically expressing CGG‐repeat RNA and FMRpolyG protein. We show that expanded CGG‐repeat RNA accumulated in intranuclear RNA structures, and these aggregates were able to cause significant granulosa cell death independent of FMRpolyG expression. Using an innovative RNA pulldown, mass spectrometry‐based approach we have identified proteins that are specifically sequestered by CGG RNA aggregates in granulosa cells in vitro, and thus may be deregulated as consequence of this interaction. Furthermore, we have demonstrated reduced expression of three proteins identified via our RNA pulldown (FUS, PA2G4 and TRA2β) in ovarian follicles in a FMR1 premutation mouse model. Collectively, these data provide evidence for the contribution of an mRNA gain‐of‐function mechanism to FXPOI disease biology.
... Of interest, the FMRpolyG protein with an expansion over~30 glycine repeats is prone to aggregation and expression of FMRpolyG in various cell and animal models leads to the formation of cytoplasmic and intranuclear aggregates, positives for ubiquitin and p62 (Todd et al., 2013;Hukema et al., 2015;Sellier et al., 2017;Derbis et al., 2018;Hoem et al., 2019;Wenzel et al., 2019). Importantly, various mouse or rabbit antibodies independently developed and directed against FMRpolyG revealed presence of this protein within the typical eosinophilic ubiquitin-positive intranuclear inclusions in cell and tissue sections of individuals with FXTAS (Todd et al., 2013;Buijsen et al., 2014;Sellier et al., 2017;Bonapace et al., 2019;Krans et al., 2019;Dijkstra et al., 2021). Of technical interest, these antibodies are directed against the N-or C-terminal parts of FMRpolyG, but none are directly targeting the polyglycine stretch. ...
Microsatellites are repeated DNA sequences of 3–6 nucleotides highly variable in length and sequence and that have important roles in genomes regulation and evolution. However, expansion of a subset of these microsatellites over a threshold size is responsible of more than 50 human genetic diseases. Interestingly, some of these disorders are caused by expansions of similar sequences, sizes and localizations and present striking similarities in clinical manifestations and histopathological features, which suggest a common mechanism of disease. Notably, five identical CGG repeat expansions, but located in different genes, are the causes of fragile X-associated tremor/ataxia syndrome (FXTAS), neuronal intranuclear inclusion disease (NIID), oculopharyngodistal myopathy type 1 to 3 (OPDM1-3) and oculopharyngeal myopathy with leukoencephalopathy (OPML), which are neuromuscular and neurodegenerative syndromes with overlapping symptoms and similar histopathological features, notably the presence of characteristic eosinophilic ubiquitin-positive intranuclear inclusions. In this review we summarize recent finding in neuronal intranuclear inclusion disease and FXTAS, where the causing CGG expansions were found to be embedded within small upstream ORFs (uORFs), resulting in their translation into novel proteins containing a stretch of polyglycine (polyG). Importantly, expression of these polyG proteins is toxic in animal models and is sufficient to reproduce the formation of ubiquitin-positive intranuclear inclusions. These data suggest the existence of a novel class of human genetic pathology, the polyG diseases, and question whether a similar mechanism may exist in other diseases, notably in OPDM and OPML.
... In fact, fragile X-associated tremor/ataxia syndrome (FXTAS) patients are characterized by an FMRP mutation in 5′UTR of the gene with CGG repetition comprising between 55 and 200, showing a neurocognitive deficit associated with AD (Aydin et al., 2020). Also, other neurodegenerative disorders are associated to FMR1 premutation, such as Parkinson disease or multiple sclerosis (Sacino et al., 2019;Dijkstra et al., 2021), pinpointing a crucial effect of FMRP mutations in common neuronal function and synaptic dysfunction. ...
Fragile X mental retardation protein (FMRP) is an RNA binding protein (RBP) whose absence is essentially associated to Fragile X Syndrome (FXS). As an RNA Binding Protein (RBP), FMRP is able to bind and recognize different RNA structures and the control of specific mRNAs is important for neuronal synaptic plasticity. Perturbations of this pathway have been associated with the autistic spectrum. One of the FMRP partners is the APP mRNA, the main protagonist of Alzheimer’s disease (AD), thereby regulating its protein level and metabolism. Therefore FMRP is associated to two neurodevelopmental and age-related degenerative conditions, respectively FXS and AD. Although these pathologies are characterized by different features, they have been reported to share a number of common molecular and cellular players. The aim of this review is to describe the double-edged sword of FMRP in autism and AD, possibly allowing the elucidation of key shared underlying mechanisms and neuronal circuits. As an RBP, FMRP is able to regulate APP expression promoting the production of amyloid β fragments. Indeed, FXS patients show an increase of amyloid β load, typical of other neurological disorders, such as AD, Down syndrome, Parkinson’s Disease, etc. Beyond APP dysmetabolism, the two neurodegenerative conditions share molecular targets, brain circuits and related cognitive deficits. In this review, we will point out the potential common neuropathological pattern which needs to be addressed and we will hopefully contribute to clarifying the complex phenotype of these two neurorological disorders, in order to pave the way for a novel, common disease-modifying therapy.
... In addition, FMRpolyG is the only species that have been found in intranuclear aggregates in neural and non-neural tissues (e.g., kidney, heart, thyroid, adrenal gland) [45], and in the cytoplasm of mural granulosa cells [46]. More recently, a vascular phenotype has been observed in the brain of PM carriers linked to the detection of FMRpolyG/p62+ inclusions [47]. Evidence for FMRpolyG toxicity has been collected from Drosophila and mouse models expressing different variants of engineered FMRpolyG aimed at enhancing or precluding such toxicity [42,48]. ...
... However, measurements of ovarian dysfunction (FSH levels) and pituitary-adrenal dysfunction (prolactin, cortisol and ACTH levels) were not correlated with scores in the FXTAS motor rating scale in PM women [91], suggesting that endocrine and motor alterations in PM carriers do not have a simple relationship. Finally, no association has been observed between CGG repeats in the normal range and reproductive parameters (including FSH and AMH levels) [126], but there are some indications regarding high levels of FSH in cases bearing intermediate CGG sizes (35)(36)(37)(38)(39)(40)(41)(42)(43)(44)(45)(46)(47)(48)(49)(50)(51)(52)(53)(54) and fecundity problems and amenorrhea [115], an observation that deserves further confirmation. ...
Abnormal trinucleotide expansions cause rare disorders that compromise quality of life and, in some cases, lifespan. In particular, the expansions of the CGG-repeats stretch at the 5’-UTR of the Fragile X Mental Retardation 1 (FMR1) gene have pleiotropic effects that lead to a variety of Fragile X-associated syndromes: the neurodevelopmental Fragile X syndrome (FXS) in children, the late-onset neurodegenerative disorder Fragile X-associated tremor-ataxia syndrome (FXTAS) that mainly affects adult men, the Fragile X-associated primary ovarian insufficiency (FXPOI) in adult women, and a variety of psychiatric and affective disorders that are under the term of Fragile X-associated neuropsychiatric disorders (FXAND). In this review, we will describe the pathological mechanisms of the adult “gain-of-function” syndromes that are mainly caused by the toxic actions of CGG RNA and FMRpolyG peptide. There have been intensive attempts to identify reliable peripheral biomarkers to assess disease progression and onset of specific pathological traits. Mitochondrial dysfunction, altered miRNA expression, endocrine system failure, and impairment of the GABAergic transmission are some of the affectations that are susceptible to be tracked using peripheral blood for monitoring of the motor, cognitive, psychiatric and reproductive impairment of the CGG-expansion carriers. We provided some illustrative examples from our own cohort. Understanding the association between molecular pathogenesis and biomarkers dynamics will improve effective prognosis and clinical management of CGG-expansion carriers.
Context:
The FMR1 gene consists of 17 exons and codes for the FMRP protein. FMR1 is involved in four genetic disorders depending on the CGG repeats length in its 5'UTR: the full mutation is responsible for the Fragile X syndrome while the premutation is associated with the Fragile X-associated Tremor/Ataxia Syndrome, Fragile X-associated Primary Ovarian Insufficiency (FXPOI) and Fragile X-associated neuropsychiatric disorders. FMR1 presents multiple isoforms resulting from skipping of exons 12 and 14 and the use of alternative splice sites in exons 15 and 17.
Aims:
To investigate the expression of Fmr1 splicing variants during folliculogenesis in the rat.
Methods:
We used preantral, early antral and preovulatory follicles to isolate RNA and characterise, by fluorescent PCR followed by sequencing, all the isoforms present in the different follicular stages.
Key results:
We identified two isoforms resulting from splicing of exon 12, six isoforms resulting from splicing of exon 14 and 15 and one isoform for exon 17.
Conclusions:
The expression levels of the isoforms vary within each follicular stage but not between different stages of folliculogenesis. Importantly, we identify for the first time in rat, an isoform that contains exon 12 and two isoforms, one that includes and one that excludes exon 14 and use the third acceptor site in exon 15.
Implications:
Characterisation of the different FMR1 variants expressed during folliculogenesis will help to understand the potential distinct cellular roles of each of them and the possible implication in the development of FXPOI.
Fragile X-associated premature ovarian insufficiency (FXPOI) is caused by expansion of a CGG repeat sequence located in the 5’ untranslated region of the FMR1 gene. Women with FXPOI have a depleted ovarian reserve, resulting in amenorrhea, hypoestrogenism, and loss of fertility before the age of 40. FXPOI is caused by CGG sequence expansions to lengths between 55 and 200 repeats, known as a FMRI premutation, however the mechanism by which the premutation drives disease pathogenesis remains unclear. Two main hypotheses exist, which describe an mRNA toxic gain-of-function mechanism or that repeat-associated non-AUG (RAN) translation results in the production of an abnormal protein, called FMRpolyG. We have developed an in vitro granulosa cell model of the FMR1 premutation by ectopically expressing CGG-repeat RNA and FMRpolyG protein. We show that expanded CGG-repeat RNA accumulated in intranuclear RNA structures, and these aggregates were able to cause significant granulosa cell death independent of FMRpolyG expression. Furthermore, using an innovative RNA pulldown, mass spectrometry-based approach we have identified proteins that bind CGG-repeat RNA in granulosa cells in vitro , and thus may be deregulated as consequence of this interaction. Collectively, these data provide evidence for the contribution of an mRNA gain-of-function mechanism to FXPOI disease biology.
