CGG Repeat Size Research Articles

Neurodegeneration of White and Gray Matter in the Hippocampus with FXTAS.

Fragile X-associated tremor/ataxia syndrome (FXTAS) is a neurodegenerative disorder that affects older premutation carriers (55-200 CGG repeats) of the fragile X gene. Despite the high prevalence of the FXTAS disorder, neuropathology studies of individuals affected by FXTAS are limited. We performed hematoxylin and eosin (H&E) staining in the hippocampus of 26 FXTAS cases and analyzed the tissue microscopically. The major neuropathological characteristics were white matter disease, intranuclear inclusions in neurons and astrocytes, and neuron loss. Astrocytes contained more and larger inclusions than neurons. There was a negative correlation between age of death and CGG repeat length in cases over the age of 60. The number of astroglial inclusions (CA3 and dentate gyrus) and the number of CA3 neuronal inclusions increased with elevated CGG repeat length. In the two cases with a CGG repeat size less than 65, FXTAS intranuclear inclusions were not present in the hippocampus, while in the two cases with less than 70 (65-70) CGG repeat expansion, neurons and astrocytes with inclusions were occasionally identified in the CA1 sub-region. These findings add hippocampus neuropathology to the previously reported changes in other areas of the brain in FXTAS patients, with implications for understanding FXTAS pathogenesis.

Stratification of the risk of ovarian dysfunction by studying the complexity of intermediate and premutation alleles of the FMR1 gene.

FMR1 premutation female carriers are at risk of developing premature/primary ovarian insufficiency (POI) with an incomplete penetrance. In this study, we determined the CGG repeat size among 1095 women with diminished ovarian reserve (DOR) / POI and characterized the CGG/AGG substructure in 44 women carrying an abnormal FMR1 repeat expansion number, compared to a group of 25 pregnant women carrying an abnormal FMR1 CGG repeat size. Allelic complexity scores of the FMR1 gene were calculated and compared between the two groups. In the DOR/POI cohort, 2.1% of women presented with an intermediate repeat size and 1.9% with a premutation. Our results suggest that the risk of POI is highest in the mid-range of CGG repeats. We observed that the allelic score is significantly higher in POI women compared to the pregnant women group (p-value = 0.02). We suggest that a high allelic score due to more than 2 AGG interspersions in the context of an intermediate number of repetitions could favor POI. Larger studies are still needed to evaluate the relevance of this new tool for the determination of the individual risk of developing POI in women with abnormal number of CGG repeats.

Relationships of Motor Changes with Cognitive and Neuropsychiatric Features in FMR1 Male Carriers Affected with Fragile X-Associated Tremor/Ataxia Syndrome.

The premutation expansion of the Fragile X Messenger Ribonucleoprotein 1 (FMR1) gene on the X chromosome has been linked to a range of clinical and subclinical features. Nearly half of men with FMR1 premutation develop a neurodegenerative disorder; Fragile X-Associated Tremor/Ataxia Syndrome (FXTAS). In this syndrome, cognitive executive decline and psychiatric changes may co-occur with major motor features, and in this study, we explored the interrelationships between these three domains in a sample of adult males affected with FXTAS. A sample of 23 adult males aged between 48 and 80 years (mean = 62.3; SD = 8.8), carrying premutation expansions between 45 and 118 CGG repeats, and affected with FXTAS, were included in this study. We employed a battery of cognitive assessments, two standard motor rating scales, and two self-reported measures of psychiatric symptoms. When controlling for age and/or educational level, where appropriate, there were highly significant correlations between motor rating score for ICARS gait domain, and the scores representing global cognitive decline (ACE-III), processing speed (SDMT), immediate memory (Digit Span), and depression and anxiety scores derived from both SCL90 and DASS instruments. Remarkably, close relationships of UPDRS scores, representing the contribution of Parkinsonism to FXTAS phenotypes, were exclusive to psychiatric scores. Highly significant relationships between CGG repeat size and most scores for three phenotypic domains suggest a close tracking with genetic liability. These findings of relationships between a constellation of phenotypic domains in male PM carriers with FXTAS are reminiscent of other conditions associated with disruption to cerebro-cerebellar circuits.

Subclinical peripheral neuropathy is common in neuronal intranuclear inclusion disease with dominant encephalopathy.

Neuronal intranuclear inclusion disease (NIID) is associated with CGG repeat expansion in the NOTCH2NLC gene. Although pure or dominant peripheral neuropathy has been described as a subtype of NIID in a few patients, most NIID patients predominantly show involvements of the central nervous system (CNS). It is necessary to further explore whether these patients have subclinical peripheral neuropathy. Twenty-eight NIID patients, clinically characterized by CNS-dominant involvements, were recruited from two tertiary hospitals. Standard nerve conduction studies were performed in all patients. Skin and sural nerve biopsies were performed in 28 and 15 patients, respectively. Repeat-primed polymerase chain reaction and amplicon length polymerase chain reaction were used to screen the CGG repeat expansion in NOTCH2NLC. All 28 patients can be diagnosed with NIID based on skin pathological and genetic changes. All patients predominantly showed CNS symptoms mainly characterized by episodic encephalopathy and cognitive impairments, but no clinical symptoms of peripheral neuropathy could be observed initially. Electrophysiological abnormalities were found in 96.4% (27/28) of these patients, indicating that subclinical peripheral neuropathy is common in NIID patients with CNS-dominant type. Electrophysiological and neuropathological studies revealed that demyelinating degeneration was the main pathological pattern in these patients, although mild axonal degeneration was also observed in some patients. No significant association between CGG repeat size and the change of nerve conduction velocity was found in these patients. This study demonstrated that most patients with CNS-dominant NIID had subclinical peripheral neuropathy. Electrophysiological examination should be the routinely diagnostic workflow for every NIID patient.

Triplet-Primed PCR Assays for Accurate Screening of FMR1 CGG Repeat Expansion and Genotype Verification.

Fragile X syndrome and other fragile X-associated disorders are caused by the full-mutation (>200 copies) and premutation (55 to 200 copies) expansion, respectively, of the CGG short tandem repeat in the fragile X messenger ribonucleoprotein 1 (FMR1) gene. Clinical diagnostic laboratories use Southern blot analysis and polymerase chain reaction (PCR)-based tests to detect and/or size the FMR1 CGG repeats. The development of sensitive and high-throughput triplet-primed PCR (TP-PCR) assays has diminished the need to subject all samples to Southern blot analysis, which is both labor- and time-intensive. In this article, we describe two direct TP-PCR (dTP-PCR) assays for the detection of FMR1 CGG repeat expansions. We outline a protocol that is based on melting curve analysis of dTP-PCR amplicons for a rapid and cost-effective first-tier screening and identification of individuals with premutation and full-mutation expansions. We also describe a protocol that employs capillary electrophoresis to resolve the dTP-PCR amplicon fragments and to estimate the repeat sizes of normal (5 to 44 copies), intermediate (45 to 54 copies), and premutation alleles, as well as to detect full mutations and determine the structure of the FMR1 alleles. © 2022 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Direct triplet-primed PCR master mix preparation and amplification of the FMR1 CGG repeat locus for melting curve analysis Basic Protocol 2: Melting curve analysis of direct triplet-primed PCR amplicons on the Rotor-Gene Q MD × 5plex high-resolution melt platform Alternate Protocol: Melting curve analysis of direct triplet-primed PCR amplicons on the LightCycler 480 system Basic Protocol 3: Generation of direct triplet-primed PCR melting curve analysis profiles Basic Protocol 4: Direct triplet-primed PCR master mix preparation and amplification of the FMR1 CGG repeat locus for capillary electrophoresis Basic Protocol 5: Generation of control FMR1 plasmids for direct triplet-primed PCR melting curve analysis Basic Protocol 6: Sanger sequencing assay to verify FMR1 CGG repeat size and structure of plasmid DNA controls.

Refining reproductive risk for FMR1 premutation carriers in the general obstetric population.

Female FMR1 premutation (FMR1 PM) carriers for fragile X syndrome (FXS) are at risk to have a child with FXS based on their CGG repeat size and AGG interruption number. Studies examining this risk in unselected populations of female PM carriers are lacking. This retrospective cohort study analyzed carrier status, CGG repeat length, AGG interruption result, and reproductive risk refinement in a population of female patients who underwent routine carrier screening for FXS. A total of 1536 PM carriers (0.43%) were identified, 95% of whom had between 55 and 90 CGG repeats. A number of 1334 carriers underwent AGG interruption testing. The majority had at least one AGG interruption and received a lower reproductive risk for FXS following AGG interruption testing (89% and 85%, respectively) as compared to their risk calculated based on CGG repeat size alone. The average change in risk across the population following AGG interruption testing was -3.4%, with a range from -50.8% to 48.9%. This article describes the range of CGG repeats and AGG interruptions in an unselected population of female PM carriers and suggests that most carriers would benefit from AGG interruption testing to refine their reproductive risk of having a child with FXS.

Cellular Bioenergetics and AMPK and TORC1 Signalling in Blood Lymphoblasts Are Biomarkers of Clinical Status in FMR1 Premutation Carriers.

Fragile X Associated Tremor/Ataxia Syndrome (FXTAS) is a neurodegenerative disorder affecting carriers of premutation alleles (PM) of the X-linked FMR1 gene, which contain CGG repeat expansions of 55–200 range in a non-coding region. This late-onset disorder is characterised by the presence of tremor/ataxia and cognitive decline, associated with the white matter lesions throughout the brain, especially involving the middle cerebellar peduncles. Nearly half of older male and ~ 20% of female PM carriers develop FXTAS. While there is evidence for mitochondrial dysfunction in neural and some peripheral tissues from FXTAS patients (though less obvious in the non-FXTAS PM carriers), the results from peripheral blood mononuclear cells (PBMC) are still controversial. Motor, cognitive, and neuropsychiatric impairments were correlated with measures of mitochondrial and non-mitochondrial respiratory activity, AMPK, and TORC1 cellular stress-sensing protein kinases, and CGG repeat size, in a sample of adult FXTAS male and female carriers. Moreover, the levels of these cellular measures, all derived from Epstein- Barr virus (EBV)- transformed and easily accessible blood lymphoblasts, were compared between the FXTAS (N = 23) and non-FXTAS (n = 30) subgroups, and with baseline data from 33 healthy non-carriers. A significant hyperactivity of cellular bioenergetics components as compared with the baseline data, more marked in the non-FXTAS PMs, was negatively correlated with repeat numbers at the lower end of the CGG-PM distribution. Significant associations of these components with motor impairment measures, including tremor-ataxia and parkinsonism, and neuropsychiatric changes, were prevalent in the FXTAS subgroup. Moreover, a striking elevation of AMPK activity, and a decrease in TORC1 levels, especially in the non-FXTAS carriers, were related to the size of CGG expansion. The bioenergetics changes in blood lymphoblasts are biomarkers of the clinical status of FMR1 carriers. The relationship between these changes and neurological involvement in the affected carriers suggests that brain bioenergetic alterations are reflected in this peripheral tissue. A possible neuroprotective role of stress sensing kinase, AMPK, in PM carriers, should be addressed in future longitudinal studies. A decreased level of TORC1—the mechanistic target of the rapamycin complex, suggests a possible future approach to therapy in FXTAS.

Refining the risk for fragile X–associated primary ovarian insufficiency (FXPOI) by FMR1 CGG repeat size

PurposeApproximately 20–30% of women with an FMR1 premutation experience fragile X–associated primary ovarian insufficiency (FXPOI); however, current risk estimates based on repeat size only identify women with the midrange of repeats to be at the highest risk. MethodsTo better understand the risk by repeat size, we collected self-reported reproductive histories on 1,668 women and divided them into high-resolution repeat size bins of ~5 CGG repeats to determine a more accurate risk for FXPOI in relation to CGG repeat length. ResultsAs previously reported, women with 70–100 CGG repeats were at the highest risk for FXPOI using various statistical models to compare average age at menopause and risk of FXPOI, with women with 85–89 repeats being at the highest risk. Importantly, women with <65 repeats or >120 repeats did not have a significantly increased risk for FXPOI compared to women with <45 repeats. ConclusionUsing a large cross-section study on 1,668 women, we have provided more personalized risk assessment for FXPOI using high-resolution repeat size bins. Understanding the variability in risk has important implications for family planning and overall health among women with a premutation.

FMR1 gene CGG repeat distribution among the three individual cohorts with intellectual disability, autism, and primary ovarian insufficiency from Tamil Nadu, Southern India.

Fragile X syndrome is the most common genetic cause of intellectual disability (ID) and is also well known to have a role in primary ovarian insufficiency (POI) and fragile X-associated tremor ataxia syndrome (FXTAS) that expresses across generations. The objective was to compare the CGG repeat variants in FMR1 gene among three correlating cohorts of ID, autism and idiopathic POI. Thirty-six patients with ID, 12 with autism spectrum disorder (ASD) and 13 females with idiopathic POI were screened for FMR1 CGG repeat size by fluorescent methylation-specific PCR and GeneScan analysis, irrespective of Hagerman checklist clinical scores. Among 29 males and seven females, 11 FMR1 allelic variants ranging from 21 to >200 CGG repeats were observed. Three (CF2-3, 39-5, 44-2) out of 29 males had full mutation alleles accounting for a 10.34% incidence of FXS among idiopathic ID males. One of them was a mosaic for CGG repeats with both premutation and full mutation alleles. The frequency of fragile X syndrome is high among patients with idiopathic ID; they also had a high score for the clinical check list. A cascade testing that begins with checklist evaluation prior to DNA analysis will be cost-effective for establishing early diagnosis in South India. With the huge disease burden, there is a need for the establishment of more molecular diagnostics and self-help groups for fragile X syndrome.

Placebo Response in Fragile X-associated Tremor/Ataxia Syndrome.

Fragile X-associated tremor/ataxia syndrome (FXTAS) is a neurodegenerative disorder characterized by intention tremor, cerebellar ataxia, and executive dysfunction in carriers of a CGG repeat expansion premutation (55-200 repeats) in the fragile X mental retardation 1 (FMR1) gene. Given reports of poor insight in FXTAS, we postulated that patients with FXTAS would be less likely to exhibit placebo response. To analyze placebo response from the first randomized controlled trial in FXTAS that evaluated cognitive and motor outcomes after 1 year of treatment with memantine. Data from the placebo arm of the first randomized controlled trial in FXTAS were analyzed. There were 2 coprimary outcomes. Based on studies in Parkinson's disease, placebo responders were defined as individuals with an improvement of at least 50% in the coprimary outcomes. Improvements of 20% and 30% served as secondary cutoff values based on the suggested magnitude of placebo response in other movement disorders. A total of 36 participants in the placebo group completed baseline and follow-up evaluations. The average age was 66 ± 7 years, and 60% were men. Average CGG repeat size was 86 ± 18. A total of 19 participants had stage 3 disease. Only 1 patient showed 50% improvement in both coprimary outcomes. At 30% and 20% improvement, there were 2 and 3 patients showing placebo response in the coprimary outcomes, respectively. Patients with FXTAS exhibited low rates of placebo response in a randomized controlled trial. Further studies on the relationship between baseline insight and placebo responsivity are applicable to FXTAS and other disorders exhibiting cognitive impairment.

Defining the role of FMR1 gene in unexplained recurrent spontaneous abortion.

Recurrent spontaneous abortion is a multifactorial disorder and till date, various factors have been attributed in its pathogenesis. Still, approximately 50% of RSA cases remain unexplained. Premutation (PM) expanded allele of fragile-X mental retardation 1 (FMR1) gene is known to contribute to ovarian dysfunction in 20% of the cases. Recently, the link between expanded FMR1 allele and recurrent miscarriages has been reported. In the present prospective case-control study, we have investigated the status of CGG repeat size at 5'UTR of the FMR1 gene in women with unexplained RSA in comparison to age-matched healthy control women (n = 100 each). The genomic DNA from these samples was subjected to molecular analysis for characterization of CGG repeat size and composition at FMR1 gene RESULTS: As compared to the control women, the RSA women cohort had a higher frequency of carriers with alleles in gray zone (GZ) and expanded PM range, i.e., 2% (2/100) versus 5% (5/100), respectively. Also, the RSA cohort had a significantly higher number of normal alleles with ≥ 35 CGG repeats (24 out of 200 alleles) as compared to control cohort (8 out of 200 alleles). The number of larger FMR1 alleles with pure CGG repeat tract was found to be significantly higher (P = 0.0063) in the RSA cohort (15 out of 200 alleles) as compared to that in control cohort (3 out of 200 alleles). Henceforth, the CGG expanded uninterrupted FMR1 allele might be associated with recurrent abortions and may help to explain many of these unexplained cases.

Isolation and Analysis of the CGG-Repeat Size in Male and Female Gametes from a Fragile X Mouse Model.

Analysis of individual gametes has a number of applications in the study of the mechanism of repeat expansion in mouse models of the fragile X-related disorders, as well as in mouse models of other repeat expansion diseases. This chapter describes the techniques required to isolate oocytes and male gametes of different stages of maturity, along with the techniques required to accurately determine the repeat number in these gametes.

Curvilinear Association Between Language Disfluency and FMR1 CGG Repeat Size Across the Normal, Intermediate, and Premutation Range.

Historically, investigations of FMR1 have focused almost exclusively on the clinical effects of CGG expansion within the categories of the premutation (55–200 CGG repeats) and fragile X syndrome (>200 CGG repeats). However, emerging evidence suggests that CGG-dependent phenotypes may occur across allele sizes traditionally considered within the “normal” range. This study adopted an individual-differences approach to determine the association between language production ability and CGG repeat length across the full range of normal, intermediate, and premutation alleles. Participants included 61 adult women with CGG repeats within the premutation (n = 37), intermediate (i.e., 41–54 repeats; n = 2), or normal (i.e., 6–40 repeats; n = 22) ranges. All participants were the biological mothers of a child with a developmental disorder, to control for the potential effects of parenting stress. Language samples were collected and the frequency of language disfluencies (i.e., interruptions in the flow of speech) served as an index of language production skills. Verbal inhibition skills, measured with the Hayling Sentence Completion Test, were also measured and examined as a correlate of language disfluency, consistent with theoretical work linking language disfluency with inhibitory deficits (i.e., the Inhibition Deficit Hypothesis). Blood samples were collected to determine FMR1 CGG repeat size. A general linear model tested CGG repeat size of the larger allele (allele-2) as the primary predictor of language disfluency, covarying for education level, IQ, age, and CGG repeats on the other allele. A robust curvilinear association between CGG length and language disfluency was detected, where low-normal (∼ <25 repeats) and mid-premutation alleles (∼90–110 repeats) were linked with higher rates of disfluency. Disfluency was not associated with inhibition deficits, which challenges prior theoretical work and suggests that a primary language deficit could account for elevated language disfluency in FMR1-associated conditions. Findings suggest CGG-dependent variation in language production ability, which was evident across individuals with and without CGG expansions on FMR1.

ASFMR1 splice variant: A predictor of fragile X-associated tremor/ataxia syndrome.

ObjectiveTo explore the association of a splice variant of the antisense fragile X mental retardation 1 (ASFMR1) gene, loss of fragile X mental retardation 1 (FMR1) AGG interspersions and FMR1 CGG repeat size with manifestation, and severity of clinical symptoms of fragile X-associated tremor/ataxia syndrome (FXTAS).MethodsPremutation carriers (PMCs) with FXTAS, without FXTAS, and normal controls (NCs) had a neurologic evaluation and collection of skin and blood samples. Expression of ASFMR1 transcript/splice variant 2 (ASFMR1-TV2), nonspliced ASFMR1, total ASFMR1, and FMR1 messenger RNA were quantified and compared using analysis of variance. Least absolute shrinkage and selection operator (LASSO) logistic regression and receiver operating characteristic analyses were performed.ResultsPremutation men and women both with and without FXTAS had higher ASFMR1-TV2 levels compared with NC men and women (n = 135,135, p < 0.0001), and ASFMR1-TV2 had good discriminating power for FXTAS compared with NCs but not for FXTAS from PMC. After adjusting for age, loss of AGG, larger CGG repeat size (in men), and elevated ASFMR1-TV2 level (in women) were strongly associated with FXTAS compared with NC and PMC (combined).ConclusionsThis study found elevated levels of ASFMR1-TV2 and loss of AGG interruptions in both men and women with FXTAS. Future studies will be needed to determine whether these variables can provide useful diagnostic or predictive information.

Molecular Characterization of FMR1 Gene by TP-PCR in Women of Reproductive Age and Women with Premature Ovarian Insufficiency.

Fragile X syndrome is caused by CGG repeat expansion mutation in the FMR1 gene. Normal alleles have 5-44 CGG repeats with AGG interruptions. The expanded gray zone (GZ) (45-54 repeats) and premutation (PM) (55-200 repeats) alleles are often uninterrupted and are unstably inherited in subsequent generations. The prevalence of PM and GZ carriers is high in the female population, at 1/66 and 1/113, respectively, and PM is associated with fertility problems in 20% of cases. Our objective was to molecularly characterize CGG repeats and AGG interruption sequences in the FMR1 gene in women of reproductive age and in women with premature ovarian insufficiency (POI). We conducted molecular analysis of the FMR1 gene from 300 women of reproductive age and 140 women with POI using triplet primed-polymerase chain reaction. This enabled us to identify carriers and to document CGG repeat size and the AGG interruption pattern. In women of reproductive age, 1.7% were GZ carriers and 0.3% were PM carriers; in women with POI, 3.6% were GZ carriers and 2.14% were PM carriers. The frequency of GZ and PM carriers did not significantly differ between the cohorts (Fisher's exact test: p<2.23 for GZ vs. control and p<0.101 for PM vs. control). Carriers received genetic counselling; family screening identified an additional seven carriers. We documented preliminary data on the prevalence of GZ and PM carriers among the studied cohorts. The identification of PM carriers among women with POI serves a dual purpose of recognizing a cause for ovarian dysfunction and enabling genetic counselling, which will help carriers when making reproductive decisions.

Sustained implantation rate is not impacted by increasing fragile X premutation allele size

Fragile X (FX) premutation carrier patients are at risk for premature ovarian insufficiency (POI), which hinder the effects of IVF. Recent work demonstrated that FX associated embryology is comparable to controls. The questions that remain are 1) how many patients that present for FX PGD have at least one embryo suitable for transfer, and 2) is the sustained implantation rate (SIR) lower for embryos created from a FX premutation carrier? Retrospective Observational Study. Patients from March 2011 to April 2017 from a single IVF center who underwent PGD for FX (with concurrent qPCR-based aneuploidy screening) were included. PGD was performed by a single laboratory that utilized qPCR-based SNP genotyping for linkage analysis. This method determines which embryos inherited the FX-positive maternal chromosome, but cannot determine the size of the CGG repeat in the embryo. Patient characteristics, PGD results, and clinical outcomes including SIR (defined as presence of fetal cardiac activity at 8 weeks of pregnancy/ the number of embryos transferred) were analyzed. Embryos were considered available for transfer if they were euploid and negative for the FX affected haplotype. 28 patients had probes developed for FX PGD and underwent a total of 49 attempted IVF cycles were included. The mean patient age was 31.7±4.8 years. The average CGG repeat size was 100.3 repeats, including 1 full mutation. Due to poor response, 1 patient did not proceed; resulting in 27 patients undergoing a retrieval. 24 patients produced viable blastocysts to undergo PGD. 21 patients had 68 embryos available for transfer, with only 3 patients not producing normal blastocysts. While 4 patients are awaiting their transfer cycles, 17 patients underwent a single embryo transfer. A total of 21 embryos were transferred. 15 patients experienced a successful implantation, yielding 18 ongoing pregnancies. The median number of cycles that a patient underwent was 1.7 cycles. 12 patients only needed 1 cycle to have a successful transfer. The euploid rate per cycle was 68.9%, as expected in this young patient cohort. The SIR for a patient’s first transfer was 88.2%, and overall the SIR was 85.7%. These numbers also suggest that the SIR does not negatively correlate with an increasing maternal premutation allele size, although more patients are needed to fully investigate.Tabled 1Maternal CGG Repeat Size45-5556-7980-99≥100Number of patients who started at least one IVF cycle21286% Patients with at least one usable embryo100% (2/2)82% (9/11)100% (5/5)83% (5/6)Average sustained implantation rate (%)100% (1/1)63% (5/8)100% (4/4)100% (4/4) Open table in a new tab Nearly all patients presenting for PGD for FX were successful in achieving at least one sustained pregnancy. To our knowledge these data are the largest cohort to date of patients undergoing PGD for FX that includes clinical outcomes.

Fragile X premutation carriers with mid-range CGG repeat size and reduction in AGG interruptions demonstrate more profoundly diminished ovarian reserve

Fragile X premutation carriers with mid-range CGG repeat size and reduction in AGG interruptions demonstrate more profoundly diminished ovarian reserve

Autistic Phenotype of Permutation and Intermediate Alleles of FMR1 Gene

Background: Mutations in gene are the most common single genetic cause of autism-spectrum disorders, therefore we investigated the possibility that the intermediate alleles may also contribute to the origin of Autistic disorder. Methods: We screened 96 males, recruited from North West of Iran, who were diagnosed with autistic disease for CGG repeat size. The frequencies of either intermediate or premutation alleles were matched with 168 male controls. All patients and normal controls were of Azeri Turkish ethnicity. Results: Three mutated, five intermediate and four premutation carriers were identified among Autistic patients, compared with one premutation carrier in normal controls, representing a significant excess in small size FMR1 allele carriers (= 0.006). There was also a significant excess of the intermediate carriers compared with normal controls (= 0.006). Conclusions: Both of these alleles may show roles in the etiology of autism, possibly as a result of the rise of mRNA. Considering that there are individuals in the general population are carriers of intermediate and premutation alleles, further screening of larger samples of Autism patients is recommended, in order to estimate the role of small size of CGG repeat alleles in the aetiology of autism.

Clinical and molecular correlates in fragile X premutation females.

The prevalence of the fragile X premutation (55–200 CGG repeats) among the general population is relatively high, but there remains a lack of clear understanding of the links between molecular biomarkers and clinical outcomes.In this study we investigated the correlations between molecular measures (CGG repeat size, FMR1 mRNA, FMRP expression levels, and methylation status at the promoter region and in FREE2 site) and clinical phenotypes (anxiety, obsessive compulsive symptoms, depression and executive function deficits) in 36 adult premutation female carriers and compared to 24 normal control subjects.Premutation carriers reported higher levels of obsessive compulsive symptoms, depression, and anxiety, but demonstrated no significant deficits in global cognitive functions or executive function compared to the control group. Increased age in carriers was significantly associated with increased anxiety levels.As expected, FMR1 mRNA expression was significantly correlated with CGG repeat number. However, no significant correlations were observed between molecular (including epigenetic) measures and clinical phenotypes in this sample. Our study, albeit limited by the sample size, establishes the complexity of the mechanisms that link the FMR1 locus to the clinical phenotypes commonly observed in female carriers suggesting that other factors, including environment or additional genetic changes, may have an impact on the clinical phenotypes. However, it continues to emphasize the need for assessment and treatment of psychiatric problems in female premutation carriers.

Prenatal diagnosis and genetic counseling of fragile X syndrome in four pedigrees by using high-resolution multiplex polymerase chain reaction

Objectives To provide prenatal diagnosis and genetic counseling for four athigh-risk pregnant women with a suspected family or personal history of fragile X syndrome (FXS) by genetic screening of fragile X mental retardation (FMR1) gene. Methods This study was conducted on four pregnant women (No.1 to 4) who received outpatient treatment in Peking University First Hospital from August 2014 to June 2016. Genomic DNA was extracted from peripheral blood samples of the pregnant women and six of their family members, four of which were suspected or confirmed FXS and the other two were FMR1 gene carriers. Amplide X kits were used to detect CGG repeat size in FMR1 gene. Two amniocytes and one chorionic villi samples were collected from three pregnant women to extract DNAs for FMR1 gene and karyotyping analyses. Results There were patients diagnosed with FXS in all the families by detecting CGG repeat numbers in FMR1 gene. The pregnant woman No.1 was a permutation carrier; No.2 carried normal FMR1 alleles while her brother had a mutation with over 20 CGG repeats in FMRI gene at chromosome X. No.3 and 4 were full mutation carriers with over 200 CGG repeats in FMR1 gene. After genetic counseling, No.3 decided to terminate the pregnancy due to abnormal fetal karyotype (47, XY,+21) and full mutation of FMR1 alleles. No.1 and 4 continued to pregnancy as their fetuses were normal in FMR1 alleles and karyotype. No.2 continued to pregnancy as her fetus was free of FXS risk. Conclusions Prenatal diagnosis and genetic counseling should be conducted on women at high-risk for FXS to avoid birth defects. People with a family history of FXS should be tested for FMR1 gene carrier status. Key words: Fragile X syndrome; Fragile X mental retardation protein; Polymerase chain reaction; Prenatal diagnosis; Genetic testing; Genetic counseling