The fragile X mental retardation 1 gene (FMR1)‐related disorder fragile X syndrome (FXS) is the most common heritable form of cognitive impairment and the second most common cause of comorbid autism. ...FXS usually results when a premutation trinucleotide CGG repeat in the 5′ untranslated region of the FMR1 gene (CGG 55–200) expands over generations to a full mutation allele (CGG >200). This expansion is associated with silencing of the FMR1 promoter via an epigenetic mechanism that involves DNA methylation of the CGG repeat and the surrounding regulatory regions. Decrease in FMR1 transcription is associated with loss of the FMR1 protein that is needed for typical brain development. The past decade has seen major advances in our understanding of the genetic and epigenetic processes that underlie FXS. Here we review these advances and their implications for diagnosis and treatment for individuals who have FMR1‐related disorders.
What This Paper Adds
Improved analysis of DNA methylation allows better epigenetic evaluation of the fragile X gene.
New testing techniques have unmasked interindividual variation among children with fragile X syndrome.
New testing methods have also detected additional cases of fragile X.
Resumen
Epigenética del síndrome X frágil y sus trastornos relacionados
El síndrome de discapacidad intelectual– gen 1 (FMR1) relacionada al síndrome de X frágil (FXS) es la forma hereditaria más común de deterioro cognitivo y la segunda causa más común de autismo comórbido. El FXS generalmente se produce cuando una repetición CGT de trinucleótido premutación en la región 5 ‘no traducida del gen FMR1 (CGG 55‐200) se expande a lo largo de varias generaciones hasta un alelo de mutación completo (CGG> 200). Esta expansión está asociada con el silenciamiento del promotor de FMR1 a través de un mecanismo epigenético que implica la metilación del ADN de la repetición de CGG y las regiones reguladoras circundantes. La disminución en la transcripción de FMR1 se asocia con la pérdida de la proteína FMR1 que se necesita para el desarrollo cerebral típico. La última década ha visto avances importantes en nuestra comprensión de los procesos genéticos y epigenéticos que subyacen al FXS. Aquí revisamos estos avances y sus implicaciones para el diagnóstico y el tratamiento de las personas que tienen trastornos relacionados con la FMR1.
Resumo
Epigenética da síndrome do X frágil e desordens relacionadas ao X frágil
A síndrome do X‐frágil (SXF) com deficiência mental relacionada a desordem do gene 1 da deficiência mental do X frágil (FMR1) é a forma mais comum de deficiência cognitiva herdável e a segunda causa mais comum de autismo como comorbidade. A SXF usualmente resulta quando uma repetição pré‐mutação do trinucleotídeo CGG na 5′ região não translacionada do gente FMR1 (CGG 55‐200) se expande por gerações para uma mutacão complete do alelo (CGG >200). Esta expansão está associada com o silenciamento do promotor do FMR1 por meio de um mecanismo epigenético que envolve metilação da repetição do CGG e das regiões regulatórias ao redor. A redução da transcrição no FMR1 está associada com perda da proteina FMR1 necessária para o desenvolvimento típico do cérebro. A última década tem visto grandes avanços na compreensão de processos genéticos e epigenéticos que explicam a SFX. Aqui, revisamos estes avanços e suas implicações para o diagnóstico e tratamento de indivíduos com desordens relacionadas ao FMR1.
What This Paper Adds
Improved analysis of DNA methylation allows better epigenetic evaluation of the fragile X gene.
New testing techniques have unmasked interindividual variation among children with fragile X syndrome.
New testing methods have also detected additional cases of fragile X.
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Relationships between Fragile X Mental Retardation 1 (FMR1) mRNA levels in blood and intragenic FMR1 CGG triplet expansions support the pathogenic role of RNA gain of function toxicity in premutation ...(PM: 55-199 CGGs) related disorders. Real-time PCR (RT-PCR) studies reporting these findings normalised FMR1 mRNA level to a single internal control gene called β-glucuronidase (GUS). This study evaluated FMR1 mRNA-CGG correlations in 33 PM and 33 age- and IQ-matched control females using three normalisation strategies in peripheral blood mononuclear cells (PBMCs): (i) GUS as a single internal control; (ii) the mean of GUS, Eukaryotic Translation Initiation Factor 4A2 (EIF4A2) and succinate dehydrogenase complex flavoprotein subunit A (SDHA); and (iii) the mean of EIF4A2 and SDHA (with no contribution from GUS). GUS mRNA levels normalised to the mean of EIF4A2 and SDHA mRNA levels and EIF4A2/SDHA ratio were also evaluated. FMR1mRNA level normalised to the mean of EIF4A2 and SDHA mRNA levels, with no contribution from GUS, showed the most significant correlation with CGG size and the greatest difference between PM and control groups (p = 10-11). Only 15% of FMR1 mRNA PM results exceeded the maximum control value when normalised to GUS, compared with over 42% when normalised to the mean of EIF4A2 and SDHA mRNA levels. Neither GUS mRNA level normalised to the mean RNA levels of EIF4A2 and SDHA, nor to the EIF4A2/SDHA ratio were correlated with CGG size. However, greater variability in GUS mRNA levels were observed for both PM and control females across the full range of CGG repeat as compared to the EIF4A2/SDHA ratio. In conclusion, normalisation with multiple control genes, excluding GUS, can improve assessment of the biological significance of FMR1 mRNA-CGG size relationships.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Fragile X syndrome (FXS) is a leading single-gene cause of intellectual disability (ID) with autism features. This study analysed diagnostic and prognostic utility of the Fragile X-Related Epigenetic ...Element 2 DNA methylation (FREE2m) assessed by Methylation Specific-Quantitative Melt Analysis and the EpiTYPER system, in retrospectively retrieved newborn blood spots (NBS) and newly created dried blood spots (DBS) from 65 children with FXS (~2-17 years). A further 168 NBS from infants from the general population were used to establish control reference ranges, in both sexes. FREE2m analysis showed sensitivity and specificity approaching 100%. In FXS males, NBS FREE2m strongly correlated with intellectual functioning and autism features, however associations were not as strong for FXS females. Fragile X mental retardation 1 gene (
) mRNA levels in blood were correlated with FREE2m in both NBS and DBS, for both sexes. In females, DNAm was significantly increased at birth with a decrease in childhood. The findings support the use of FREE2m analysis in newborns for screening, diagnostic and prognostic testing in FXS.
Fragile X syndrome (FXS) is a common cause of intellectual disability and autism spectrum disorder (ASD) usually associated with a CGG expansion, termed full mutation (FM: CGG ≥ 200), increased DNA ...methylation of the FMR1 promoter and silencing of the gene. Mosaicism for presence of cells with either methylated FM or smaller unmethylated pre-mutation (PM: CGG 55-199) alleles in the same individual have been associated with better cognitive functioning. This study compares age- and sex-matched FM-only and PM/FM mosaic individuals on intellectual functioning, ASD features and maladaptive behaviours.
This study comprised a large international cohort of 126 male and female participants with FXS (aged 1.15 to 43.17 years) separated into FM-only and PM/FM mosaic groups (90 males, 77.8% FM-only; 36 females, 77.8% FM-only). Intellectual functioning was assessed with age appropriate developmental or intelligence tests. The Autism Diagnostic Observation Schedule-2nd Edition was used to examine ASD features while the Aberrant Behavior Checklist-Community assessed maladaptive behaviours.
Comparing males and females (FM-only + PM/FM mosaic), males had poorer intellectual functioning on all domains (p < 0.0001). Although females had less ASD features and less parent-reported maladaptive behaviours, these differences were no longer significant after controlling for intellectual functioning. Participants with PM/FM mosaicism, regardless of sex, presented with better intellectual functioning and less maladaptive behaviours compared with their age- and sex-matched FM-only counterparts (p < 0.05). ASD features were similar between FM-only and PM/FM mosaics within each sex, after controlling for overall intellectual functioning.
Males with FXS had significantly lower intellectual functioning than females with FXS. However, there were no significant differences in ASD features and maladaptive behaviours, after controlling for intellectual functioning, independent of the presence or absence of mosaicism. This suggests that interventions that primarily target cognitive abilities may in turn reduce the severity of maladaptive behaviours including ASD features in FXS.
Fragile X syndrome (FXS) is a common monogenic cause of intellectual disability with autism features. While it is caused by loss of the
1 product (FMRP), mosaicism for active and inactive
alleles, ...including alleles termed premutation (PM: 55-199 CGGs), is not uncommon. Importantly, both PM and active full mutation (FM: ≥ 200 CGGs) alleles often express elevated levels of mRNA that are thought to be toxic. This study determined if complete
mRNA silencing from FM alleles and/or levels of
mRNA (if present) in blood are associated with intellectual functioning and autism features in FXS.
The study cohort included 98 participants (70.4% male) with FXS (FM-only and PM/FM mosaic) aged 1-43 years. A control group of 14 females were used to establish control
mRNA reference range. Intellectual functioning and autism features were assessed using the Mullen Scales of Early Learning or an age-appropriate Wechsler Scale and the Autism Diagnostic Observation Schedule-2nd Edition (ADOS-2), respectively.
mRNA was analysed in venous blood collected at the time of assessments, using the real-time PCR relative standard curve method.
Females with FXS had significantly higher levels of
mRNA (
< 0.001) than males.
mRNA levels were positively associated with age (
< 0.001), but not with intellectual functioning and autistic features in females. FM-only males (aged < 19 years) expressing FM
mRNA had significantly higher ADOS calibrated severity scores compared to FM-only males with completely silenced
(
= 0.011). However, there were no significant differences between these subgroups on intellectual functioning. In contrast, decreased levels of
mRNA were associated with decreased intellectual functioning in FXS males (
= 0.029), but not autism features, when combined with the PM/FM mosaic group.
Incomplete silencing of toxic FM RNA may be associated with autistic features, but not intellectual functioning in FXS males. While decreased levels of mRNA may be more predictive of intellectual functioning than autism features. If confirmed in future studies, these findings may have implications for patient stratification, outcome measure development, and design of clinical and pre-clinical trials in FXS.
Fragile X syndrome (FXS) is caused by hypermethylation of the FMR1 promoter due to the full mutation expansion (full mutation FM: CGG ≥ 200 repeats) and silencing of FMR1. Assessment of mosaicism for ...active‐unmethylated alleles has prognostic utility. This study examined relationships between FMR1 methylation in different tissues with FMR1 messenger ribonucleic acid (mRNA) and intellectual functioning in 87 males with FXS (1.89–43.17 years of age). Methylation sensitive Southern blot (mSB) and Methylation Specific‐Quantitative Melt Aanalysis (MS‐QMA) were used to examine FMR1 methylation. FMR1 mRNA levels in blood showed strong relationships with FMR1 methylation assessed using MS‐QMA in blood (n = 68; R2 = 0.597; p = 1.4 × 10−10) and buccal epithelial cells (BEC) (n = 62; R2 = 0.24; p = 0.003), with these measures also showing relationships with intellectual functioning scores (p < 0.01). However, these relationships were not as strong for mSB, with ~40% of males with only FM alleles that were 100% methylated and non‐mosaic by mSB, showing methylation mosaicism by MS‐QMA. This was confirmed through presence of detectable levels of FMR1 mRNA in blood. In summary, FMR1 methylation levels in blood and BEC examined by MS‐QMA were significantly associated with FMR1 mRNA levels and intellectual functioning in males with FXS. These relationships were not as strong for mSB, which underestimated prevalence of mosaicism.
For years, premutation-carriers of fragile X-syndrome (FXS) were assumed free from any deleterious phenotype. In this review, we discuss the current literature on neurocognitive, emotional and ...neuromotor profiles emerging in females with the fragile-X premutation, and discuss phenotypic profiles in male premutation-carriers to gain insights into possible underlying mechanisms associated with FMR1 gene expression. We contend that this emerging phenotypic profile in females with the fragile-X premutation needs further investigation using experimentally-driven tasks sensitive to neural networks especially vulnerable to FMR1 gene expression. Further investigation of developmental aspects of the female carrier profile is needed to determine the extent to which emotional, cognitive and neurobehavioural challenges indicate at-risk profiles for later degenerative decline, or rather a stable developmental phenotype. These future research avenues will provide critical new information which will enable identification of women at greatest risk for subtle age-dependent neurobehavioural changes well before the onset of more serious clinical consequences alongside the identification of biomarkers which may be useful in establishing the efficacy of future therapeutic interventions.
Developmental delay phenotypes have been associated with FMR1 premutation (PM: 55–200 CGG repeats) and “gray zone” (GZ: 45–54 CGG repeats) alleles. However, these associations have not been confirmed ...by larger studies to be useful in pediatric diagnostic or screening settings.
This study determined the prevalence of PM and GZ alleles in two independent cohorts of 19,076 pediatric referrals to developmental delay diagnostic testing through Victorian Clinical Genetics Service (cohort 1: N = 10,235; cohort 2: N = 8841), compared with two independent general population cohorts (newborn screening N = 1997; carrier screening by the Victorian Clinical Genetics Service prepair program N = 14,249).
PM and GZ prevalence rates were not significantly increased (p > 0.05) in either developmental delay cohort (male PM: 0.12–0.22%; female PM: 0.26–0.33%; male GZ: 0.68–0.69%; female GZ: 1.59–2.13-%) compared with general population cohorts (male PM: 0.20%; female PM: 0.27–0.82%; male GZ: 0.79%; female GZ: 1.43–2.51%). Furthermore, CGG size distributions were comparable across datasets, with each having a modal value of 29 or 30 and ~1/3 females and ~1/5 males having at least one allele with ≤26 CGG repeats.
These data do not support the causative link between PM and GZ expansions and developmental-delay phenotypes in pediatric settings.
There is now growing evidence of cognitive weakness in female premutation carriers (between 55 and 199 CGG repeats) of the fragile X mental retardation gene, including impairments associated with ...executive function. While an age-related decline in assessments of executive function has been found for male premutation carriers, few studies have explored whether female carriers show a similar trajectory with age. A total of 20 female premutation carriers and 21 age- and IQ-matched healthy controls completed a battery of tasks assessing executive function tasks, including the behavioural dyscontrol scale (BDS), symbol digit modalities test (SDMT), paced auditory serial addition test (PASAT), Haylings sentence completion test and the digit span task (forward and backward). Performance was compared between premutation carriers and healthy controls, and the association between task performance and age was also ascertained. Compared to controls, female premutation carriers had significant impairment on the BDS, SDMT, PASAT, and Haylings sentence completion task, all of which rely on quick, or timed, responses. Further analyses revealed no significant association between age and task performance for either premutation carriers or controls. This study demonstrates that a cohort of female premutation carriers have deficits on a range of tasks of executive function that require the rapid temporal resolution of responses. We propose that the understanding of the phenotype of premutation carriers will be advanced through use of such measures.
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