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RETT syndrome is caused by mutations in X-linked MECP2, encoding methyl-CpG-binding protein 2
Abstract
Rett syndrome (RTT, MIM 312750) is a progressive neurodevelopmental disorder and one of the most common causes of mental retardation in females, with an incidence of 1 in 10,000-15,000 (ref. 2). Patients with classic RTT appear to develop normally until 6-18 months of age, then gradually lose speech and purposeful hand use, and develop microcephaly, seizures, autism, ataxia, intermittent hyperventilation and stereotypic hand movements. After initial regression, the condition stabilizes and patients usually survive into adulthood. As RTT occurs almost exclusively in females, it has been proposed that RTT is caused by an X-linked dominant mutation with lethality in hemizygous males. Previous exclusion mapping studies using RTT families mapped the locus to Xq28 (refs 6,9,10,11). Using a systematic gene screening approach, we have identified mutations in the gene (MECP2 ) encoding X-linked methyl-CpG-binding protein 2 (MeCP2) as the cause of some cases of RTT. MeCP2 selectively binds CpG dinucleotides in the mammalian genome and mediates transcriptional repression through interaction with histone deacetylase and the corepressor SIN3A (refs 12,13). In 5 of 21 sporadic patients, we found 3 de novo missense mutations in the region encoding the highly conserved methyl-binding domain (MBD) as well as a de novo frameshift and a de novo nonsense mutation, both of which disrupt the transcription repression domain (TRD). In two affected half-sisters of a RTT family, we found segregation of an additional missense mutation not detected in their obligate carrier mother. This suggests that the mother is a germline mosaic for this mutation. Our study reports the first disease-causing mutations in RTT and points to abnormal epigenetic regulation as the mechanism underlying the pathogenesis of RTT.
... A disorder that highlights The value of pre-clinical research is Rett syndrome (RTT). a severe neurological disorder caused by mutations in the X-linked gene MECP2 (Amir et al., 1999). Mouse models recapitulate many features of the human condition (Chen et al. 2001;Guy et al. 2001), supporting the conclusion that The function of the MeCP2 protein is the same in mice as in humans. ...
... Additionally, certain classes of restriction enzymes play key roles in DNA mapping, epigenome mapping, and constructing DNA libraries. [2] 2. Zinc finger nucleases: increased recognition potential ...
... Additionally, ZFNs have been used to edit tumorinfiltrating lymphocytes as a treatment strategy for metastatic melanoma. [2] ...
Abstact:-Recent advancements in our understanding of the genetic basis of neurological disorders have spurred significant efforts to develop gene-based therapies. These therapies aim to correct the underlying genetic defects, either by introducing a functional copy of the mutated gene or by directly editing the genomic sequence. This review explores the main genetic strategies currently being investigated for the treatment of monogenic neurological disorders, highlighting the challenges and ethical considerations associated with these approaches. Introduction:
... The MeCP2 protein binds to methylated cytosines, and regulates expression of thousands of genes. 1,2 Loss of function or deletion mutations of MECP2 cause Rett syndrome (RTT, MIM# 312750), 3 a severe neurodevelopmental disorder (NDD) characterized by developmental delay/intellectual disability (DD/ID), dysautonomia, epilepsy, gastrointestinal problems, sleep disturbances, and hand stereotypies. 4 However, there are males with loss of function or hypomorphic alleles who survive into infancy or childhood and demonstrate RTT-like features. ...
... The remaining electrophysiological studies, including VEP assessments, are provided in the Notes S1. (2)(3)(4) in the indicated brain areas. The MDS group had significantly higher delta power than the TD group at the frontal, central and temporal regions during wake stage. ...
Objective
Rett syndrome (RTT) and MECP2 duplication syndrome (MDS) result from under‐ and overexpression of MECP2, respectively. Preclinical studies using genetic‐based treatment showed robust phenotype recovery for both MDS and RTT. However, there is a risk of converting MDS to RTT, or vice versa, if accurate MeCP2 levels are not achieved. The aim of this study was to identify biomarkers distinguishing RTT from MDS.
Materials and Methods
We prospectively enrolled 11 MDS and 6 male RTT like (MRL) individuals for a panel of clinical and neurophysiological assessments over two visits, 8–10 months apart.
Results
We identified numerous clinical and physiological features as promising biomarkers. MRL individuals exhibited large amplitude whole body tremor, midline stereotypies (vs. hand flapping at sides in MDS), earlier neuromotor regression, and earlier onset but less commonly refractory epilepsy. In the neurophysiological domain, we observed several marked differences in sleep physiology between MDS/MRL and typically developing (TD) individuals including reduced sleeping time, increased delta power during rapid eye movement (REM) sleep, decreased occipital alpha and increased brain‐wide delta power during wakefulness, and reduced spindle density and duration. MRL individuals also had much lower delta power during NREM 2 and 3 stages than the TD group. We found differences in spindle duration in the temporal lobes and spindle amplitude in the frontal lobes between MDS and MRL.
Discussion
Our study revealed distinct clinical features of MDS and MRL that can be monitored during a clinical trial and may serve as target engagement, disease progression, or safety biomarkers for interventional studies.
... Rettov sindrom (RS) progresivni je pervazivni neurorazvojni poremećaj koji pogađa gotovo isključivo djevojčice i drugi je najčešći uzrok razvojnog zaostajanja/intelektualnih poteškoća kod osoba ženskog spola, s pojavnošću od 1:10 000 do 1:15 000 živorođenih djevojčica (1,2). Klasični oblik RS-a u 95% slučajeva nastaje na X kromosomu de novo mutacijom gena koji kodira metil-CpG-vezajući protein 2 (MeCP2) (3). MeCP2 kontrolira ekspresiju gena i modulira arhitekturu kromatina vezanjem na metilirani DNA te kao transkripcijski čimbenik sudjeluje u regulaciji gena važnih za funkcioniranje sinapsi u mozgu (3). ...
... Klasični oblik RS-a u 95% slučajeva nastaje na X kromosomu de novo mutacijom gena koji kodira metil-CpG-vezajući protein 2 (MeCP2) (3). MeCP2 kontrolira ekspresiju gena i modulira arhitekturu kromatina vezanjem na metilirani DNA te kao transkripcijski čimbenik sudjeluje u regulaciji gena važnih za funkcioniranje sinapsi u mozgu (3). ...
Rettov sindrom je progresivni pervazivni neurorazvojni poremećaj koji pogađa gotovo isključivo djevojčice i drugi je najčešći uzrokintelektualnih poteškoća kod osoba ženskog spola, s pojavnošću od 1:10 000 do 1:15 000 živorođenih djevojčica. Klasični oblik Rettovog sindroma u 95% slučajeva nastaje na kromosomu X de novo mutacijom gena koji kodira metil-CpG-vezajući protein 2. Karakteriziran je razvojnim zaostajanjem/intelektualnim poteškoćama, ataksijom, epilepsijom, karakterističnim stereotipnim kretnjamaruku i gubitkom njihove normalne funkcije te poremećajem disanja. Nema učinkovitog lijeka, ali ranim prepoznavanjem bolesti tepočetkom terapije može se usporiti nastanak daljnjih komplikacija. Poremećaji disanja koji uključuju apneje u budnosti, hiperventilaciju, ubrzano plitko disanje te zadržavanje daha rezultat su brojnih faktora, uključujući oksidativni stres i kroničnu subkliničkuupalu.U ovom radu prikazujemo djevojčicu koja boluje od Rettovog sindroma i u nekoliko je navrata liječena antibioticima zbog radiološkiperzistirajućih promjena gornjeg desnog plućnog režnja sa smetnjama ventilacije i prominentnim desnim hilusom. Nađene promjene gornjeg desnog režnja pluća na kompjuteriziranoj tomografiji toraksa, prema podatcima iz literatrure, mogu se naći u 50% djeces Rettovim sindromom i kroničnog su karaktera te ne zahtijevaju liječenje antibioticima.
... Rett syndrome (RTT, OMIM #312,750) is a rare X-linked neurodevelopmental disorder caused in over 95% of cases by mutations in the Methyl-CpG-binding Protein 2 (MECP2) gene 1,2 . However, some people with RTTcausing MECP2 mutations show neurodevelopmental abnormalities but lack the characteristic clinical features of RTT 3 . ...
Mutations of the MECP2 gene lead to Rett syndrome (RTT), a rare developmental disease causing severe intellectual and physical disability. How the loss or defective function of MeCP2 mediates RTT is still poorly understood. MeCP2 is a global gene expression regulator, acting at transcriptional and post-transcriptional levels. Little attention has been given so far to the contribution of alternative splicing (AS) dysregulation to RTT pathophysiology. To perform a comparative analysis of publicly available RNA sequencing (RNA-seq) studies and generate novel data resources for AS, we explored 100 human datasets and 130 mouse datasets from Mecp2-mutant models, processing data for gene expression and alternative splicing. Our comparative analysis across studies indicates common species-specific differentially expressed genes (DEGs) and differentially alternatively spliced (DAS) genes. Human and mouse dysregulated genes are involved in two main functional categories: cell-extracellular matrix adhesion regulation and synaptic functions, the first category more significantly enriched in human datasets. Our extensive bioinformatics study indicates, for the first time, a significant dysregulation of AS in human RTT datasets, suggesting the crucial contribution of altered RNA processing to the pathophysiology of RTT.
... Of note, cardiac abnormalities including thickened aortic valve, atrial septal defect, patent ductus arteriosus, and patent foramen ovale have also been identified in subjects with Witteveen-Kolk syndrome 21,22 . Also found in the Sin3A complex is MECP2, an X-linked methyl CpG binding protein associated with Rett syndrome, the most common cause of cognitive impairment in females 23 . Further, the Sin3A complex is known to regulate DNA methylation (DNAm) via interactions with the Tet family of methylcytosine dioxygenases. ...
Hypoplastic left heart syndrome (HLHS) is a severe congenital heart disease associated with microcephaly and poor neurodevelopmental outcomes. Here we show that the Ohia HLHS mouse model, with mutations in Sap130, a chromatin modifier, and Pcdha9, a cell adhesion protein, also exhibits microcephaly associated with mitotic block and increased apoptosis leading to impaired cortical neurogenesis. Transcriptome profiling, DNA methylation, and Sap130 ChIPseq analyses all demonstrate dysregulation of genes associated with autism and cognitive impairment. This includes perturbation of REST transcriptional regulation of neurogenesis, disruption of CREB signaling regulating synaptic plasticity, and defects in neurovascular coupling mediating cerebral blood flow. Adult mice harboring either the Pcdha9 mutation, which show normal brain anatomy, or forebrain-specific Sap130 deletion via Emx1-Cre, which show microcephaly, both demonstrate learning and memory deficits and autism-like behavior. These findings provide mechanistic insights indicating the adverse neurodevelopment in HLHS may involve cell autonomous/nonautonomous defects and epigenetic dysregulation.
... The discovery in 1999 that Rett patients had a mutation in the MECP2 gene represented a turning point in understanding the pathogenesis of this disease [9]. Subsequent research has indicated that 95% of individuals with RS exhibit spontaneous mutations in the X-linked MECP2 gene. ...
Rett syndrome (RS) is a rare neurodevelopmental disorder primarily caused by mutations in the X-linked methyl-CpG binding protein 2 (MECP2) gene, responsible for encoding MECP2 which plays a pivotal role in regulating gene expression. The neurological and non-neurological manifestations of RS vary widely in severity depending on the specific mutation type. Bone complications, mostly scoliosis but also osteoporosis, hip displacement, and a high rate of fractures, are among the most prevalent non-neurological comorbidities observed in girls with RS. Low bone mineral density (BMD) is primarily due to a slow rate of bone formation due to dysfunctional osteoblast activity. The use of anticonvulsants, immobilization, low physical activity, poor nutrition, and inadequate vitamin D intake all significantly hamper skeletal maturation and the accumulation of bone mass in RS girls, making them more susceptible to fragility fractures. In RS patients, the upper and lower limbs are the most common sites for fractures which are due to both a reduced BMD and a diminished bone size. This review summarizes the knowledge on risk factors for fragility fracture in patients with RS and proposes a potential diagnostic and therapeutic pathway to enhance low BMD and mitigate the risk of fragility fractures. In particular, this review focused on the importance of clinical and instrumental evaluation of bone status as a basis for adequate planning of nutritional, pharmacological, and surgical interventions to be undertaken. Additionally, the management of bone defects in individuals with RS should be customized to meet each person’s specific needs, abilities, and general health.
... The MECP2 gene encodes methyl CpG binding protein 2 (MECP2), which is involved in various functions, including chromatin architecture, RNA splicing and transcriptional regulation, needed for normal brain development [1][2][3]. Loss-of-function mutations in MECP2 in females cause classic Rett syndrome (OMIM#312750), while hemizygous mutations in males are typically lethal [4]. However, in some cases, specific MECP2 mutations in males can lead to a spectrum of phenotypes, ranging from severe neonatal encephalopathy to intellectual disability with non-specific features [5,6]. ...
Background: MECP2 duplication syndrome (MDS) (MIM#300260) is a rare X-linked neurodevelopmental disorder. This study aims to (1) develop a specific clinical severity scale, (2) explore its correlation with clinical and molecular variables, and (3) automate diagnosis using the Face2gene platform. Methods: A retrospective study was conducted on genetically confirmed MDS patients who were evaluated at a pediatric hospital between 2012 and 2024. Epidemiological, clinical, and molecular data were collected. A standardized clinical questionnaire was collaboratively developed with input from physicians and parents. Patient photographs were used to train Face2Gene. Results: Thirty-five patients (0–24 years, 30 males) were included. Key features in males comprised intellectual disability (100%), hypotonia (93%), autism spectrum disorder (77%) and developmental regression (52%). Recurrent respiratory infections (79%), dysphagia (73%), constipation (73%) and gastroesophageal reflux (57%) were common. Seizures occurred in 53%, with 33% being treatment-refractory. The Face2Gene algorithm was successfully trained to identify MDS. A specific clinical severity scale (MECPDup) was developed and validated, correlating with the MBA (a scale developed for Rett syndrome). The MECPDup score was significantly higher in males (p < 0.001) and those with early death (p = 0.003). It showed significant positive correlations with age (p < 0.001) and duplication size (p = 0.044). Conclusions: This study expands the understanding of MDS through comprehensive clinical and molecular insights. The integration of AI-based facial recognition technology and the development of the MECPDup severity scale hold promise for enhancing diagnostic accuracy, monitoring disease progression, and evaluating treatment responses in individuals affected by MDS.
... Rett syndrome (RTT) is a neurodevelopmental disorder caused primarily by pathogenic loss-of-function variants on the X-linked MECP2gene [1]. It is a rare disorder that affects approximately 1 in 9000 liveborn females [2]. ...
Introduction
The clinical, research and advocacy communities for Rett syndrome are striving to achieve clinical trial readiness, including having fit-for-purpose clinical outcome assessments. This study aimed to (1) describe psychometric properties of clinical outcome assessment for Rett syndrome and (2) identify what is needed to ensure that fit-for-purpose clinical outcome assessments are available for clinical trials.
Methods
Clinical outcome assessments for the top 10 priority domains identified in the Voice of the Patient Report for Rett syndrome were compiled and available psychometric data were extracted. The clinical outcome assessments measured clinical severity, functional abilities, comorbidities and quality of life, and electrophysiological biomarkers. An international and multidisciplinary panel of 29 experts with clinical, research, psychometric, biostatistical, industry and lived experience was identified through International Rett Syndrome Foundation networks, to discuss validation of the clinical outcome assessments, gaps and next steps, during a workshop and in a follow-up questionnaire. The identified gaps and limitations were coded using inductive content analysis.
Results
Variable validation profiles across 26 clinical outcome assessments of clinical severity, functional abilities, and comorbidities were discussed. Reliability, validity, and responsiveness profiles were mostly incomplete; there were limited content validation data, particularly parent-informed relevance, comprehensiveness and comprehensibility of items; and no data on meaningful change or cross-cultural validity. The panel identified needs for standardised administration protocols and systematic validation programmes.
Conclusion
A pipeline of collaborative clinical outcome assessment development and validation research in Rett syndrome can now be designed, aiming to have fit-for-purpose measures that can evaluate meaningful change, to serve future clinical trials and clinical practice.
... The usual cause of RTT is a pathogenic variant in MECP2, a gene on the X chromosome [1,3]. The elucidation of the usual molecular basis of RTT [28] has stimulated efforts to develop effective treatments [11,18,21,25]. ...
Background
We have held a ‘trouble-shooting’ clinic for Rett syndrome patients from 2003 until the COVID pandemic in 2020. The clinic was multidisciplinary, including clinical genetics, paediatric neurology, adult learning disability psychiatry and physiotherapy. Access to specialist communication support and eye-gaze equipment was also often available. We have reviewed the files of patients seen in the clinic and conducted a survey of parents’ and carers’ satisfaction with the clinic and their experiences during COVID.
Results
Of the 117 patients seen in the clinic, records were reviewed of 103 (97 female, six male) who attended a total of 123 appointments. The records were unavailable for 14 patients. The most common reasons for referral were assessment of ‘episodes’ of uncertain nature (possibly epileptic, possibly autonomic), the wish for a general review by an experienced team, and questions about the diagnosis. We discuss the nature of the advice we were able to provide and offer some brief case vignettes. We wrote to the parents or carers of all patients seen and 63 respondents were willing to be interviewed about the clinic and their experiences during COVID. Respondents were generally complimentary about the clinic team, emphasising the value of a specialist clinic for those affected by a rare condition. Respondents gave insight into the range of problems experienced during COVID, especially the isolation resulting from the withdrawal of services, demonstrating the value of community support. Some respondents mentioned the shift to remote consultations, which they hoped would continue after COVID for its convenience. However, others talked about how difficult it is in a remote consultation to explain the problems of the affected family member to professionals who do not know the patient or know about Rett syndrome.
Conclusions
Our findings demonstrate the value of a disease-specific clinic provided by staff experienced with the particular rare condition. Meeting the needs of patients with ultra-rare conditions presents additional challenges. We have also found that the shift to holding a virtual clinic during COVID brought the benefit of convenience but was unsatisfactory in other ways, as it makes clinical assessment more difficult and fails to overcome the sense of isolation during a pandemic.
... The protein fine tunes the expression of thousands of genes in the brain by binding to methylated cytosines predominantly in the CG and CAC context [1][2][3]. Loss-of-function variants, including exonic deletions and indels, of MECP2 cause Rett syndrome (MIM: 312750) [4], a severe postnatal neurodevelopmental disorder that primarily affects females. Conversely, copy number gains encompassing MECP2 lead to MRXSL (MIM: 300260), an X-linked genomic disorder primarily affecting males, with an estimated prevalence of one in 100,000 live male births [5]. ...
Background
MECP2 Duplication Syndrome, also known as X-linked intellectual developmental disorder Lubs type (MRXSL; MIM: 300260), is a neurodevelopmental disorder caused by copy number gains spanning MECP2. Despite varying genomic rearrangement structures, including duplications and triplications, and a wide range of duplication sizes, no clear correlation exists between DNA rearrangement and clinical features. We had previously demonstrated that up to 38% of MRXSL families are characterized by complex genomic rearrangements (CGRs) of intermediate complexity (2 ≤ copy number variant breakpoints < 5), yet the impact of these genomic structures on regulation of gene expression and phenotypic manifestations have not been investigated.
Methods
To study the role of the genomic rearrangement structures on an individual’s clinical phenotypic variability, we employed a comprehensive genomics, transcriptomics, and deep phenotyping analysis approach on 137 individuals affected by MRXSL. Genomic structural information was correlated with transcriptomic and quantitative phenotypic analysis using Human Phenotype Ontology (HPO) semantic similarity scores.
Results
Duplication sizes in the cohort ranging from 64.6 kb to 16.5 Mb were classified into four categories comprising of tandem duplications (48%), terminal duplications (22%), inverted triplications (20%), and other CGRs (10%). Most of the terminal duplication structures consist of translocations (65%) followed by recombinant chromosomes (23%). Notably, 65% of de novo events occurred in the Terminal duplication group in contrast with 17% observed in Tandem duplications. RNA-seq data from lymphoblastoid cell lines indicated that the MECP2 transcript quantity in MECP2 triplications is statistically different from all duplications, but not between other classes of genomic structures. We also observed a significant (p < 0.05) correlation (Pearson R = 0.6, Spearman p = 0.63) between the log-transformed MECP2 RNA levels and MECP2 protein levels, demonstrating that genomic aberrations spanning MECP2 lead to altered MECP2 RNA and MECP2 protein levels. Genotype–phenotype analyses indicated a gradual worsening of phenotypic features, including overall survival, developmental levels, microcephaly, epilepsy, and genitourinary/eye abnormalities in the following order: Tandem duplications, Other complex duplications, Terminal duplications/Translocations, and Triplications encompassing MECP2.
Conclusion
In aggregate, this combined analysis uncovers an interplay between MECP2 dosage, genomic rearrangement structure and phenotypic traits. Whereas the level of MECP2 is a key determinant of the phenotype, the DNA rearrangement structure can contribute to clinical severity and disease expression variability. Employing this type of analytical approach will advance our understanding of the impact of genomic rearrangements on genomic disorders and may help guide more targeted therapeutic approaches.
... study; Population pharmacokinetic modeling; Rett syndrome; Trofinetide; Weight-banded dosing regimen smaller number of male individuals [4]. Almost all cases are caused by de novo loss-of-function mutations in the MECP2 gene, which encodes methyl-CpG-binding protein 2 (MeCP2), an epigenetic regulator of gene expression [5]. MeCP2 deficiency leads to abnormal neuronal maturation and plasticity [6][7][8]. ...
Weight-banded trofinetide dosing improved physician- and caregiver-rated efficacy measures and had acceptable tolerability in patients aged 2‒4 years (DAFFODIL study) and 5‒20 years (LAVENDER study) with Rett syndrome (RTT). Selection of weight-banded dosing regimens for these studies was based on population pharmacokinetic (popPK) modeling and exposure simulations. This study applied an updated popPK model to confirm steady-state trofinetide exposures achieved in DAFFODIL patients were within target range.
A popPK model was developed using data from 14 clinical studies of trofinetide in healthy volunteers and pediatric and adult patients, including the LAVENDER and DAFFODIL studies. Individual exposure measures (area under concentration–time curve over 0–12 h [AUC0–12] were generated via integration of the predicted concentration–time profile for each DAFFODIL study participant based on the popPK model and individual empiric Bayesian pharmacokinetic parameter estimates. Distributions of steady-state AUC0–12 values for each body-weight group were compared against target exposure (AUC0–12 = 800‒1200 µg·h/mL).
Distribution and box plots of simulated steady-state AUC0–12 values achieved with the weight-banded DAFFODIL dosing regimen used in younger individuals aged 2–4 years with RTT (twice daily trofinetide 5 g [≥ 9 to < 12 kg] or 6 g [≥ 12 to < 20 kg]) indicated good overlap with the target exposure range. Median steady-state AUC0–12 values for both body-weight bands fell within the target exposure range.
PopPK model-based simulations confirm that the weight-banded dosing regimen used in DAFFODIL is adequate to achieve target trofinetide exposure in 2- to 4-year-olds with RTT.
... Over the past few decades, there has been considerable interest in understanding the function of methyl-CpG-binding protein 2 (MeCP2). Mutations in MeCP2 cause Rett syndrome (RTT), a devastating neurological disorder that primarily affects female children after birth and is characterized by abnormal brain development, progressive loss of motor coordination, intellectual disability, loss of communication and other severe conditions [1][2][3] . Understanding the mechanisms according to which MeCP2 functions will provide insights into how dysregulation of MeCP2 leads to RTT. ...
Mutations in methyl-CpG-binding protein 2 (MeCP2) cause Rett syndrome. MeCP2 is thought to regulate gene transcription by binding to methylated DNA broadly across the genome. Here, using cleavage under target and release under nuclease (CUT&RUN) assays in the adult mouse cortex, we show that MeCP2 strongly binds to specific gene enhancers that we call MeCP2-binding hotspots (MBHs). Unexpectedly, we find that MeCP2 binding to MBHs occurs in a DNA methylation-independent manner at MBHs. Multiple MBH sites surrounding genes mediate the transcriptional repression of genes enriched for neuronal functions. We show that MBHs regulate genes irrespective of genic methylation levels, suggesting that MeCP2 controls transcription via an intragenic methylation-independent mechanism. Hence, disruption of intragenic methylation-independent gene regulation by MeCP2 may in part underlie Rett syndrome.
... It is linked to the X chromosome and affects females mainly, with a frequency of 1:15,000 live births [1,2]. In 95% of cases, the classical form of RTT is associated with mutations in the mecp2 gene [3], codifying for the methyl-CpG-binding protein 2 (MeCP2), which has the role of an epigenetic modulator. Most patients with RTT develop normally up to 6-18 months of age [1]. ...
Background: Rett syndrome (RTT) is an early-onset neurological disorder primarily affecting females, leading to severe cognitive and physical disabilities. Recent studies indicate that an imbalance of redox homeostasis and exacerbated inflammatory responses are key players in the clinical manifestations of the disease. Emerging evidence highlights that the p75 neurotrophin receptor (p75NTR) is implicated in the regulation of oxidative stress (OS) and inflammation. Thus, this study is aimed at investigating the effects of p75NTR modulation by LM11A-31 on fibroblasts derived from RTT donors. Methods: RTT cells were treated with 0.1 µM of LM11A-31 for 24 h, and results were obtained using qPCR, immunofluorescence, ELISA, and Western blot techniques. Results: Our findings demonstrate that LM11A-31 reduces OS markers in RTT fibroblasts. Specifically, p75NTR modulation by LM11A-31 restores protein glutathionylation and reduces the expression of the pro-oxidant enzyme NOX4. Additionally, LM11A-31 significantly decreases the expression of the pro-inflammatory mediators interleukin-6 and interleukin-8. Additionally, LM11A-31 normalizes the expression levels of transcription factors involved in the regulation of the antioxidant response and inflammation. Conclusions: Collectively, these data suggest that p75NTR modulation may represent an effective therapeutic target to improve redox balance and reduce inflammation in RTT.
... RTT is caused by mutations in the gene encoding the MeCP2 protein (Amir et al., 1999). Remarkably, MeCP2 has been shown to undergo LLPS and disease-related mutations significantly alter this process (Fan et al., 2020;Jiang et al., 2020;Wang et al., 2020). ...
Mutations of the cyclin-dependent kinase-like 5 (CDKL5) gene, which encodes a serine/threonine protein kinase, can cause the CDKL5 deficiency disorder (CDD), a severe neurodevelopmental disease characterized by epileptic encephalopathy and neurocognitive impairment. The CDKL5 kinase consists of a catalytic N-terminal domain (NTD) and a less characterized C-terminal domain (CTD). Numerous disease-related mutations truncate CDKL5, leaving the NTD intact while variably shortening the CTD, which highlights the importance of the CTD for CDKL5 function. By systematically analyzing CDKL5 compositional features and evolutionary dynamics, we found that the CTD is a low-complexity region (LCR) highly enriched in serine residues and with a high propensity to undergo liquid-liquid phase separation (LLPS), a biophysical process of condensation controlling protein localization and function. Using a combination of super-resolution imaging, electron microscopy, and molecular and cellular approaches, including optogenetic LLPS induction, we discovered that CDKL5 undergoes LLPS, predominantly driven by its CTD, forming membraneless condensates in neuronal and non-neuronal cells. A CTD internal fragment (CTIF) plays a pivotal LLPS-promoting role, along with the distal portion of the protein. Indeed, two disease-related truncating mutations (S726X and R781X), eliding variable portions of the CTIF, significantly impair LLPS. This impairment is paralleled at the functional level by a reduction in the CDKL5-dependent phosphorylation of EB2, a known CDKL5 target. These findings demonstrate that CDKL5 undergoes LLPS, driven by a CTD region elided by most disease-related truncating mutations. Its loss, through the impairment of CDKL5 LLPS and functional activity, may play a key role in the molecular pathogenesis of CDD.
... Mammalian MBD proteins perform important functions. Mutations in the human MeCP2 gene cause a progressive neurological disorder, namely, Rett syndrome (RTT) [11]. Mammals are not the only species containing MBD motifs. ...
Methyl-CpG-binding domain (MBD) proteins play vital roles in epigenetic gene regulation, and they have diverse molecular, cellular, and biological functions in plants. MBD proteins have been functionally characterized in a few plant species. However, the structure and function of MBD proteins in Arabidopsis halleri and Arabidopsis lyrata remain unknown. In this study, 12 A. halleri MBD (AhMBD) and 13 A. lyrata MBD (AlMBD) genes were identified. A phylogenetic analysis of the Arabidopsis genus showed that the MBD proteins of three species (Arabidopsis thaliana, A. helleri, and A. lyrata) could be classified into eight classes. Expression patterns suggested that the AtMBD genes were expressed in different tissues. We characterized the function of AtMBD3 and found that it was constitutively localized to the nucleus and interacted with several AtMBD protein members. Our results reveal that AtMBD3 is involved in the development of A. thaliana, which may be helpful in further studies on these genes in A. helleri and A. lyrata.
Human puberty is a dynamic biological process determined by the increase in the pulsatile secretion of GnRH triggered by distinct factors not fully understood. Current knowledge reveals fine tuning between an increase in stimulatory factors and a decrease in inhibitory factors, where genetic and epigenetic factors have been indicated as key players in the regulation of puberty onset by distinct lines of evidence. Central precocious puberty (CPP) results from the premature reactivation of pulsatile secretion of GnRH. In the past decade, the identification of genetic causes of CPP has largely expanded, revealing hypothalamic regulatory factors of pubertal timing. Among them, 3 genes associated with CPP are linked to mechanisms involving DNA methylation, reinforcing the strong role of epigenetics underlying this disorder. Loss-of-function mutations in Makorin Ring-Finger Protein 3 (MKRN3) and Delta-Like Non-Canonical Notch Ligand 1 (DLK1), 2 autosomal maternally imprinted genes, have been described as relevant monogenic causes of CPP with the phenotype exclusively associated with paternal transmission. MKRN3 has proven to be a key component of the hypothalamic inhibitory input on GnRH neurons through different mechanisms. Additionally, rare heterozygous variants in the Methyl-CpG-Binding Protein 2 (MECP2), an X-linked gene that is a key factor of DNA methylation machinery, were identified in girls with sporadic CPP with or without neurodevelopmental disorders. In this mini-review, we focus on how the identification of genetic causes of CPP has revealed epigenetic regulators of human pubertal timing, summarizing the latest knowledge on the associations of puberty with MKRN3, DLK1, and MECP2.
The cases of head and neck cancer among persons with intellectual disability (PWID) are infrequently reported and therefore poorly understood. PWID often face increased barriers of access to healthcare, which can be further compounded when faced with a cancer diagnosis. This report presents the case of a 34‐year‐old Chinese female patient with Rett syndrome and intellectual disability, presenting with two primary cancers of the tongue and the trachea. The case highlights the importance of regular dental review in cancer surveillance, post‐operative supportive dental care, and challenges in behavioral management in cancer diagnosis and treatment.
The brain plays a vital role in maintaining homeostasis and effective interaction with the environment, shaped by genetic and environmental factors throughout neurodevelopment and maturity. While genetic components dictate initial neurodevelopment stages, epigenetics—specifically neuroepigenetics—modulates gene expression in response to environmental influences, allowing for brain adaptability and plasticity. This interplay is particularly evident in neuropathologies like Rett syndrome and CDKL5 deficiency syndrome, where disruptions in neuroepigenetic processes underline significant cognitive and motor impairments. The environmental enrichment paradigm, introduced by Donald Hebb in the late 1940s, demonstrates how enriching stimuli—such as complex sensory, social, and cognitive inputs—affect brain structure and function. Despite methodological variability, evidence reveals that enriched environments catalyze beneficial changes in behavior and neuroanatomy, including increased synaptic plasticity, enhanced motor coordination, and improved cognitive performance in rodent models. Additionally, environmental enrichment induces epigenetic modifications that facilitate these outcomes, highlighting the necessity of understanding the mechanisms driving gene expression changes within the context of enriched experiences. Ultimately, this manifold relationship between environment, neuroepigenetic modulation, and brain function highlights the brain’s capacity for change, reinforcing the importance of considering environmental factors in studies of neurodevelopment and therapy for neurological disorders.
During the last decade, the study of behavioral phenotypes has generated a great deal of interest that has focused on genetically determined syndromes associated with mental retardation. As Hodapp and Dykens (1) report, there has been a 10-fold or greater increase in the number of articles published on the etiologies of genetic syndromes associated with mental retardation in the 1990s compared to the 1980s. The increase does not merely reflect advances in molecular genetics, but also includes a growing awareness that behavioral disorders are not a nonspecific outcome of mental retardation. It is well known that mental retardation is associated with an increased risk of behavioral disorder: epidemiological work carried out a quarter of a century ago indicated that 30% of mentally retarded children had some form of psychopathology, compared to 6% of controls (2).Part of the increased risk is due to the presence of behavioral phenotypes that are relatively specific and characteristic of individual genetic syndromes.
Mental retardation (MR) is defined as an overall “intelligence quotient” (IQ) <70, associated with functional deficits in adaptive behaviour (such as daily-living skills, social skills and communication), with an onset before 18 years (1). Approximately 2–3% of the population have an intelligence quotient (IQ) <70 (2,3) and at least 0.3% of individuals are severely handicapped (IQ <50) (Table 1), yet a cause for mental retardation is established in less than half of all cases (4). The underlying causes of MR are extremely heterogeneous (Table 2). In addition to multiple nongenetic factors that act prenatally or during early infancy and cause brain injury, chromosomal anomalies, such as aneuploidy syndromes, for example, Down syndrome, the microdeletion syndromes, for example, Prader—Willi, Angelman, Miller—Dieker, Smith—Magenis, and Williams syndromes, represent an important genetic cause of MR. Recent studies suggest that chromosomal rearrangements that affect the telomeric regions of autosomes, not detectable by conventional cytogenetic analysis, may account for up to 7% of moderate to severe MR (5,6). Genetic causes may be involved in one half of severely retarded patients (7). Some disorders for which the gene is identified affect relatively significant numbers of patients and families, such as the fragile X syndrome (which affects approx 1/4000–6000 males) (8,9) and Rett syndrome (1/10,000–15,000 girls) (10),but our knowledge of these monogenic causes is still far from complete.
Rett syndrome (RS) was first recognized in the early 1960s as a developmental disorder affecting young females only. Andreas Rett, a Viennese developmental pediatrician, reported the initial accounts of this unique syndrome, but none was widely circulated (1). Bengt Hagberg identified girls in Sweden with similar clinical features and together with colleagues from France and Portugal presented the first English language publication on RS in 1983 (2). As a result, RS was soon recognized in the United States, Japan, and throughout western Europe (3–6), and has now been reported in all ethnic groups. The prevalence of RS (Table 1) ranges from 1/10,000 in Sweden (7) to 1/22,000 in Texas (8). In the United States, more than 3000 females meeting the clinical criteria for RS have been identified. As described in the following paragraphs, mutations in the gene MECP2, which encodes methyl-CpG-binding protein 2, have been found in most girls or women (and some boys) with RS. Surprisingly, mutations in this gene can also lead to a wide variety of clinical phenotypes ranging from normal females to fatal encephalopathy in males.
Rett syndrome, a neurodevelopmental disorder primarily affecting females, presents unique challenges in managing associated scoliosis. This study aims to evaluate the efficacy and challenges of posterior spinal fusion (PSF) in Rett syndrome patients by analyzing postoperative complications. A retrospective cohort study was conducted using a large national database. We included Rett syndrome patients aged 10–18 years who underwent PSF between 2010 and 2020. Outcomes such as medical and surgical complications, emergency department visits, readmissions, mortality, and reoperation rates up to 5 years were compared with a matched neuromuscular scoliosis (NMS) group. The study identified 195 Rett syndrome patients and 973 NMS patients. Post-surgery, Rett syndrome patients showed a significantly higher incidence of pneumothorax (56.9%, P < 0.001), respiratory failure (24.6%, P = 0.013), and pneumonia (26.2%, P < 0.001). Additionally, ileus (7.2%, P = 0.041), acute kidney injury (14.9%, P = 0.029), and urinary tract infections (14.9%, P < 0.001) were also significantly more frequent in the Rett syndrome group. Rett syndrome group also had higher rates of transfusion (11.3%, P = 0.004). Interestingly, the incidence of pseudarthrosis, implant complications, junctional failures, and the necessity for reoperation did not significantly differ at postoperative year 2. Mid-term follow-up showed that the reoperation rates over a 5-year period did not significantly differ between the Rett syndrome and NMS groups. Rett syndrome is associated with increased immediate postoperative complications, necessitating tailored preoperative planning, and intensive postoperative care. Despite these challenges, the mid-term surgical outcomes are comparable to those in NMS patients.
Background: Neurodevelopmental disorders (NDDs) affect approximately 15% of children and adolescents worldwide. This group of disorders is often polygenic with varying risk factors, with many associated genes converging on shared molecular pathways, including chromatin regulation and transcriptional control. Understanding how NDD-associated chromatin regulators and protein complexes orchestrate these regulatory pathways is crucial for elucidating NDD pathogenesis and developing targeted therapeutic strategies. Recently, the TCF20/PHF14 chromatin complex was identified in the mammalian brain, expanding the list of chromatin regulatory remodelers implicated in NDDs. This complex—which includes MeCP2, RAI1, TCF20, PHF14, and HMG20A—plays a vital role in epigenetic and transcriptional regulation. Methods: We review and summarize current research and clinical reports pertaining to the different components of the MeCP2-interacting TCF20/PHF14 complex. We examine the NDDs associated with the TCF20/PHF14 complex, explore the molecular and neuronal functions of its components, and discuss emerging therapeutic strategies targeting this complex to mitigate symptoms, with broader applicability to other NDDs. Results: Mutations in the genes encoding the components of the MeCP2-interacting TCF20/PHF14 complex have been linked to various NDDs, underscoring its critical contribution to brain development and NDD pathogenesis. Conclusions: The MeCP2-interacting TCF20/PHF14 complex and its associated NDDs could serve as a model system to provide insight into the interplay between epigenetic regulation and NDD pathogenesis.
Post-transcriptional mechanisms, such as alternative splicing and polyadenylation, are recognized as critical regulatory processes that increase transcriptomic and proteomic diversity. The advent of next-generation sequencing and whole-genome analyses has revealed that numerous transcription and epigenetic regulators, including transcription factors and histone-modifying enzymes, undergo alternative splicing, most notably in the nervous system. Given the complexity of regulatory processes in the brain, it is conceivable that many of these splice variants control different aspects of neuronal development. Mutations or dysregulation of splicing and transcription regulatory proteins are frequently linked to various neurodevelopmental disorders, highlighting the importance of understanding the role of neuron-specific alternative splicing in maintaining proper transcriptional regulation in the brain. This review consolidates current insights into the role of alternative splicing in influencing transcriptional and chromatin regulatory programs in neuronal development.
Sleep problems are common in Rett syndrome and other neurogenetic syndromes. Actigraphy is a cost-effective, objective method for measuring sleep. Current guidelines require caregiver-reported bed and wake times to facilitate actigraphy data scoring. The current study examined missingness and consistency of caregiver-reported bed and wake times from paper sleep diaries and actigraphy event mark button presses in a sample of 38 individuals with Rett and related syndromes (aged 2–36 years, mean = 13.1) across two 14-day collection time points. Rates of missingness and discrepancy between the 2 sources were relatively high and correlated with clinical severity and quality of life. Overall, the results suggest a need for alternative actigraphy scoring methods that do not rely on caregiver report in this population.
Bone is a unique organ crucial for locomotion, mineral metabolism, and hematopoiesis. It maintains homeostasis through a balance between bone formation by osteoblasts and bone resorption by osteoclasts, which is regulated by the basic multicellular unit (BMU). Abnormal bone metabolism arises from an imbalance in the BMU. Osteoclasts, derived from the monocyte‐macrophage lineage, are regulated by the RANKL‐RANK‐OPG system, which is a key factor in osteoclast differentiation. RANKL activates osteoclasts through its receptor RANK, while OPG acts as a decoy receptor that inhibits RANKL. In trabecular bone, high turnover involves rapid bone formation and resorption, influenced by conditions such as malignancy and inflammatory cytokines that increase RANKL expression. Cortical bone remodeling, regulated by aged osteocytes expressing RANKL, is less understood, despite ongoing research into how Rett syndrome, characterized by MeCP2 abnormalities, affects RANKL expression. Balancing trabecular and cortical bone involves mechanisms that preserve cortical bone, despite overall bone mass reduction due to aging or oxidative stress. Research into genes like sFRP4, which modulates bone mass, highlights the complex regulation by BMUs. The roles of the RANKL‐RANK‐OPG system extend beyond bone, affecting processes such as aortic valve formation and temperature regulation, which highlight the interconnected nature of biological research.
The 2024 International Rett Syndrome Foundation (IRSF) Rett Syndrome Scientific Meeting, held in Westminster, Colorado, gathered over 200 researchers and clinicians to discuss advancements in understanding and treating Rett syndrome (RTT). Key topics included MeCP2 biology, neuronal circuitry, therapeutic development, and clinical trial outcomes. The meeting reinforced the importance of collaborative research in unraveling RTT’s complex pathology and advancing treatment approaches. With promising therapeutic candidates in development, the conference underscored a growing hope for effective treatments, offering a path toward improving the quality of life for individuals with Rett syndrome and their families.
Both genetic and environmental factors shape the process of neurodevelopment. At the interface of nature and nurture lies neuronal activity-dependent transcription, which, by internalizing experiences, sculpts and optimizes the primitive neural circuits laid down by a hardwired genetic program. In this chapter, we will review the important roles played by activity-dependent transcription in the key steps of the development of functional neural circuits: neurite development, synapse formation, and synaptic plasticity. We’ll also discuss how activity-dependent transcription is relevant to understanding the mechanisms underlying neurodevelopmental disorders.
Alternative polyadenylation (APA) is a pervasive regulatory mechanism in the human brain that controls the stability and cellular localization of mRNA transcripts. Single-nucleotide polymorphisms associated with psychiatric disorders may exert their deleterious effects by altering 3’ untranslated site usage, which may change the stability and processing of mRNA transcripts. The authors previously performed a 3’APA transcriptomic-wide association study using the DePars2 framework and the GTEx v8, PsychENCODE, and ROS/MAP datasets to identify APA-linked genes associated with eleven brain disorders. Here we focus on 3’APA-linked genes associated with the major psychiatric conditions: schizophrenia, bipolar disorder, and depression. There are 286 APA-linked genes associated with these psychiatric disorders, and 60%–65% of these genes have not been associated with the major psychiatric disorders through their expression and/or splicing. Protein–protein interaction networks indicate that APA-linked genes associated with schizophrenia are involved in intracellular transport and cellular localization pathways. Future research is needed to elucidate the role of alternative 3’ untranslated region usage of APA-linked genes on neuronal function and phenotypic expression in psychiatric disorders.
Background : Methyl CpG binding protein 2 (MECP2) is vital for neuronal function as it is an essential global modulator of transcription, and mutations in MECP2 are the most common cause of Rett syndrome, an X-linked neurodevelopmental disorder. Patients diagnosed with Rett syndrome have increased risk for epilepsy as well as problems with anxiety and social communication. Using the zebrafish mecp2 Q63X line, this study aimed to increase our understanding of the role of mecp2 in regulation of seizures and general locomotion, ontogeny of social behaviour, and adult socialization and anxiety behaviour. To determine responses of mecp2 -/- zebrafish to a stimulating convulsant, general locomotor activity was measured at 5 days post-fertilization (dpf) in sibling mecp2 +/+ , mecp2 +/- , and mecp2 -/- fish after treatment with a GABA A receptor antagonist pentylenetetrazol (PTZ) at varying concentrations. Responses to social stimulus were investigated in larval (21 dpf) and adult mecp2 -/- and mecp2 +/+ fish. Anxiety responses to a novel tank and whole-body cortisol levels were also measured in adult mecp2 -/- and control mecp2 +/+ zebrafish. Results : The behavioural tests showed that mecp2 -/- zebrafish displayed hypolocomotion at the larval stage, along with increased freezing time and thigmotaxis, and higher whole-body cortisol levels in adulthood. However, lack of functional Mecp2 did not change the hyper-locomotion response to PTZ at 5 dpf or affect the social preference for visual social stimulus at 21 dpf and in adulthood. Conclusions: Functional mecp2 modulated larval locomotion and behavioural anxiety at different ages and adult cortisol levels, but lacking mecp2 did not alter adult locomotion or socialization, and developmental sociability and PTZ-induced hyperlocomotion in zebrafish. Given the variability reported in humans and rodent Mecp2 models, studies using zebrafish can explore vital elements of MECP2’s role across development and improve our understanding of neural mechanisms underlying neurodevelopmental disorders.
Rett syndrome is a severe neurodevelopmental disorder in girls, underpinned by mutations in the X-linked gene MECP2. In their recent work (Frasca et al, 2024), Frasca and colleagues identified a novel pathway involving interferon-gamma (IFNγ) that could pave the way to potential therapies.
MeCP2 is a chromosomal protein which binds to DNA that is methylated at CpG. In situ immunofluorescence in mouse cells has shown that the protein is most concentrated in pericentromeric heterochromatin, suggesting that MeCP2 may play a role in the formation of inert chromatin. Here we have isolated a minimal methyl-CpG binding domain (MBD) from MeCP2. MBD is 85 amino acids in length, and binds exclusively to DNA that contains one or more symmetrically methylated CpGs. MBD has negligable non-specific affinity for DNA, confirming that non-specific and methyl-CpG specific binding domains of MeCP2 are distinct. In vitro footprinting indicates that MBD binding can protect a 12 nucleotide region surrounding a methyl-CpG pair, with an approximate dissociation constant of 10(-9) M.
MeCP2 is an abundant mammalian protein that binds to methylated CpG. We have found that native and recombinant MeCP2 repress transcription in vitro from methylated promoters but do not repress nonmethylated promoters. Repression is nonlinearly dependent on the local density of methylation, becoming significant at the density found in bulk vertebrate genomic DNA. Transient transfection using fusions with the GAL4 DNA binding domain identified a region of MeCP2 that is capable of long-range repression in vivo. Moreover, MeCP2 is able to displace histone H1 from preassembled chromatin that contains methyl-CpG. These properties, together with the abundance of MeCP2 and the high frequency of its 2 bp binding site, suggest a role as a global transcriptional repressor in vertebrate genomes.
Although familial recurrences of Rett syndrome (RTT) comprise only approximately 1% of the reported cases, it is these cases that hold the key for the understanding of the genetic basis of the disorder. Families in which RTT occurs in mother and daughter, aunt and niece, and half sisters are consistent with dominant inheritance and variable expressivity of the phenotype. Recurrence of RTT in sisters is likely due to germ-line mosaicism in one of the parents, rather than to recessive inheritance. The exclusive occurrence of classic RTT in females led to the hypothesis that it is X-linked and may be lethal in males. In an X-linked dominant disorder, unaffected obligate-carrier females would be expected to show nonrandom or skewed inactivation of the X chromosome bearing the mutant allele. We investigated the X chromosome inactivation (XCI) patterns in the female members of a newly identified family with recurrence of RTT in a maternal aunt and a niece. Skewing of XCI is present in the obligate carrier in this family, supporting the hypothesis that RTT is an X-linked disorder. However, evaluation of the XCI pattern in the mother of affected half sisters shows random XCI, suggesting germ-line mosaicism as the cause of repeated transmission in this family. To determine which regions of the X chromosome were inherited concordantly/discordantly by the probands, we genotyped the individuals in the aunt-niece family and two previously reported pairs of half sisters. These combined exclusion-mapping data allow us to exclude the RTT locus from the interval between DXS1053 in Xp22.2 and DXS1222 in Xq22.3. This represents an extension of the previous exclusion map.
CpG methylation in vertebrates correlates with alterations in chromatin structure and gene silencing. Differences in DNA-methylation status are associated with imprinting phenomena and carcinogenesis. In Xenopus laevis oocytes, DNA methylation dominantly silences transcription through the assembly of a repressive nucleosomal array. Methylated DNA assembled into chromatin binds the transcriptional repressor MeCP2 which cofractionates with Sin3 and histone deacetylase. Silencing conferred by MeCP2 and methylated DNA can be relieved by inhibition of histone deacetylase, facilitating the remodelling of chromatin and transcriptional activation. These results establish a direct causal relationship between DNA methylation-dependent transcriptional silencing and the modification of chromatin.
Methylation at the DNA sequence 5'-CpG is required for mouse development. MeCP2 and MBD1 (formerly PCM1) are two known proteins that bind specifically to methylated DNA via a related amino acid motif and that can repress transcription. We describe here three novel human and mouse proteins (MBD2, MBD3, and MBD4) that contain the methyl-CpG binding domain. MBD2 and MBD4 bind specifically to methylated DNA in vitro. Expression of MBD2 and MBD4 tagged with green fluorescent protein in mouse cells shows that both proteins colocalize with foci of heavily methylated satellite DNA. Localization is disrupted in cells that have greatly reduced levels of CpG methylation. MBD3 does not bind methylated DNA in vivo or in vitro. MBD1, MBD2, MBD3, and MBD4 are expressed in somatic tissues, but MBD1 and MBD2 expression is reduced or absent in embryonic stem cells which are known to be deficient in MeCP1 activity. The data demonstrate that MBD2 and MBD4 bind specifically to methyl-CpG in vitro and in vivo and are therefore likely to be mediators of the biological consequences of the methylation signal.
A systematic search for expressed sequences in the human Xq28 region resulted in the isolation of 8.5 kb large contigs of human and murine cDNAs with no apparent conserved open reading frames. These cDNAs were found to be derived from the 3"-untranslated region (3"-UTR) of the methyl-CpG-binding protein 2 gene ( MeCP2 ). This long 3"-UTR is part of an alternatively polyadenylated, 10.1 kb MeCP2 transcript which is differentially expressed in human brain and other tissues. RNA in situ hybridization to sections of mouse embryo and adult tissues of an Mecp2 3"-UTR probe showed ubiquitous low level expression in early organogenesis and enhanced expression in the hippocampus during formation of the differentiated brain. Sequence comparison between the human and mouse homologues revealed several blocks of very high conservation separated by less conserved sequences. Additional support for a domain-like conservation pattern of the long 3"-UTR of the MeCP2 gene was obtained by examining conservation in the chimpanzee, orangutan, macaque, hamster, rat and kangaroo. The minimum free energy distribution for the predicted RNA secondary structure was very similar in human and mouse sequences. In particular, the conserved blocks were predicted to be of high minimum free energy, which suggests weak secondary structure with respect to RNA folding. The fact that both the sequence and predicted secondary structure have been highly conserved during evolution suggests that both the primary sequence and the three-dimensional structure of the 3"-UTR may be important for its function in post-transcriptional regulation of MeCP2 expression.
To date over 1,000 cases of Rett syndrome have been described in females exclusively. Some of these cases, less than 2 in 100, are familial. The inheritance through maternal lines in the familial cases suggests that Rett syndrome is an X-linked disorder lethal in males. Hypotheses about the genetic mechanisms involved in this syndrome along with suggestions to approach the molecular basis of this disorder are presented. (J Child Neurol 1988;3(Suppl):S76-S78).
The most likely cause of the Rett syndrome (RTT) is an X-linked dominant mutation lethal in hemizygous males. Previous exclusion mapping studies have identified putative regions for the RTT gene on the X chromosome. In the present study, we evaluated two candidate genes, glutamate dehydrogenase-2 (GLUD2 and rab GDP-dissociation inhibitor (GDI1/XAP-4), chosen because of their expression patterns and functions in the central nervous system and their location in the nonexcluded region of Xq. The intronless gene GLUD2, located in Xq25 and expressed in neuronal and testicular tissues, is involved in the metabolism of glutamate, a neurotransmitter reported to be elevated in the spinal fluid of RTT individuals. The GLUD2 gene was screened for mutations by Southern hybridization and by direct sequencing of polymerase chain reaction (PCR) products. The GDI1 gene in Xq28, also known as RABGDIA or XAP-4, encodes a human GDI that is expressed predominantly in neuronal and sensory tissues. All 11 exons and splice junctions of the GDI1 gene were PCR-amplified and sequenced directly or screened by single-strand conformation analysis. No mutation in either of these two genes was found in 22 RTT patients. Therefore, GLUD2 and GDI1 can be excluded as candidate genes for this syndrome. Am. J. Med. Genet. 78:169–172, 1998. © 1998 Wiley-Liss, Inc.
Rett syndrome (RTT) is an X-linked dominant neurodevelopmental disorder that affects females. Exclusion mapping studies using a new family with maternal inheritance of RTT defined Xq28 as the candidate region for the RTT gene. Six candidate genes were selected for mutation analysis based on their established expression patterns and known functions in the CNS. These are: Glutamate receptor subunit 3 (GLUR3), GABA receptor subunit alpha 3 (GABRA3), GABA receptor subunit e1 (GABRE1), Vacuolar ATPase subunit 1 (VATPS1, XAP3), the human homologue of plexin 3-SEX (XAP6) and the Synaptobrevin-like protein (SYBL1). Major rearrangements involving these genes were excluded by Southern analysis. No disease-causing mutations were found, but several single-nucleotide polymorphisms (SNPs) were detected. These SNPs will be useful in future linkage analysis and whole-genome association studies for other diseases. The genomic characterization of GLUR3 and GABRA3 will allow mutational analysis of these genes as candidates for other X-linked neurological disorders mapping to Xq25-Xq26 and Xq28. Am. J. Med. Genet. 90:69–71, 2000. © 2000 Wiley-Liss, Inc.
DNA methylation is essential for murine development and is implicated in the control of gene expression. MeCP2, MBD1, MBD2,
MBD3, and MBD4 comprise a family of mammalian, nuclear proteins related by the presence in each of an amino acid motif called
the methyl-CpG binding domain (MBD). Each of these proteins, with the exception of MBD3, is capable of binding specifically
to methylated DNA. MeCP2, MBD1 and MBD2 can also repress transcription. We describe the genomic structure and chromosomal
localization of the human and murine Mbd1, Mbd2, Mbd3, and Mbd4 genes. We find that the highly similar MBD2 and MBD3 proteins are encoded by genes that map to different chromosomes in humans
and mice but show a similar genomic structure. The Mbd1 and Mbd2 genes, in contrast, map together to murine and human Chromosomes (Chrs)18. The Mbd3 and Mbd4 genes map to murine Chrs 10 and 6, respectively, while the human MBD3 and MBD4 genes map to Chrs 19 and 3, respectively.
The human androgen-receptor gene (HUMARA; GenBank) contains a highly polymorphic trinucleotide repeat in the first exon. We have found that the methylation of HpaII and HhaI sites less than 100 bp away from this polymorphic short tandem repeat (STR) correlates with X inactivation. The close proximity of the restriction-enzyme sites to the STR allows the development of a PCR assay that distinguishes between the maternal and paternal alleles and identifies their methylation status. The accuracy of this assay was tested on (a) DNA from hamster/human hybrid cell lines containing either an active or inactive human X chromosome; (b) DNA from normal males and females; and (c) DNA from females showing nonrandom patterns of X inactivation. Data obtained using this assay correlated substantially with those obtained using the PGK, HPRT, and M27 beta probes, which detect X inactivation patterns by Southern blot analysis. In order to demonstrate one application of this assay, we examined X inactivation patterns in the B lymphocytes of potential and obligate carriers of X-linked agammaglobulinemia.
Methylation of mammalian DNA can lead to repression of transcription and alteration of chromatin structure. Recent evidence suggests that both effects are the result of an interaction between the methylated sites and methyl-CpG-binding proteins (MeCPs). MeCP1 has previously been detected in crude nuclear extracts. Here we report the identification, purification, and cDNA cloning of a novel MeCP called MeCP2. Unlike MeCP1, the new protein is able to bind to DNA that contains a single methyl-CpG pair. By staining with an antibody, we show that the distribution of MeCP2 along the chromosomes parallels that of methyl-CpG. In mouse, for example, MeCP2 is concentrated in pericentromeric heterochromatin, which contains a large fraction (about 40%) of all genomic 5-methylcytosine.
The hypothesis of germinal mosaicism in the unaffected mother of two half-sisters affected with Rett syndrome is postulated to explain the unusual recurrence of this genetic disorder affecting only females (1/10,000); it might be caused by new X-linked mutations with lethality in male fetuses. The analysis of 34 X-linked restriction fragment length polymorphisms (RFLPs) in these two affected females and in their unaffected mother and half-brother, together with the reconstruction of phase for 15 informative RFLPs in somatic cell hybrids retaining a single X chromosome from each female, has made it possible to exclude some regions of the X chromosome as possible sites of the mutation(s) causing Rett syndrome.
Rett syndrome is a neurologic disorder characterized by early normal development followed by regression, acquired deceleration of head growth, autism, ataxia, and stereotypic hand movements. The exclusive occurrence of the syndrome in females and the occurrence of a few familial cases with inheritance through maternal lines suggest that this disorder is most likely secondary to a mutation on the X chromosome. To address this hypothesis and to identify candidate regions for the Rett syndrome gene locus, genotypic analysis was performed in two families with maternally related affected half-sisters by using 63 DNA markers from the X chromosome. Maternal and paternal X chromosomes from the affected sisters were separated in somatic cell hybrids and were examined for concordance/discordance of maternal alleles at the tested loci. Thirty-six markers were informative in at least one of the two families, and 25 markers were informative in both families. Twenty loci were excluded as candidates for the Rett syndrome gene, on the basis of discordance for maternal alleles in the half-sisters. Nineteen of the loci studied were chosen for multipoint linkage analysis because they have been previously genetically mapped using a large number of meioses from reference families. Using the exclusion criterion of a lod score less than -2, we were able to exclude the region between the Duchenne muscular dystrophy locus and the DXS456 locus. This region extends from Xp21.2 to Xq21-q23. The use of the multipoint linkage analysis approach outlined in this study should allow the exclusion of additional regions of the X chromosome as new markers are analyzed. This in turn will result in a defined region of the X chromosome that should be searched for candidate sequences for the Rett syndrome gene in both familial and sporadic cases.
The Rett syndrome (RS) is a degenerative neurological disorder occurring exclusively in young females. The disorder is sporadic in the majority of the cases, however a few familial cases with inheritance through maternal lines have been identified. Based on these observations the condition could be due to an X chromosome mutation which is lethal in males. To explain the familial cases, a hypothesis of possible non-random X inactivation is proposed. To investigate the possibility of non-random X chromosome inactivation in RS, we carried out analysis using restriction fragment length polymorphisms (RFLPs) and methylation sensitive enzymes at the PGK and HPRT loci. The results show that there is increased incidence of non-random X chromosome inactivation in peripheral blood leukocytes in sporadic RS patient (36%), as compared to healthy controls (8%). Using brain tissue from three patients, only a random pattern was detected, although varying degrees of skewing were detected in the peripheral tissues of these patients. Analysis of leukocyte DNA from a mother of two affected half-sisters revealed non-random X chromosome inactivation suggesting a possible selection against RS allele. Additional familial cases of RS should be evaluated to determine if this observation is common to all female carriers. If non-random X chromosome inactivation occurs in all the putative "carriers," this would be the first evidence to support the hypothesis of an X linked mutation which is lethal in males.
Diagnostic ultrasound has been extensively used for neurologic evaluation of cranial, vascular and spinal diseases. This study presents the techniques, methodology, and procedures for a new diagnostic application of ultrasound for evaluation of the neuromuscular system. In order to determine the optimum sonographic characterization of neuromuscular disease, normal anatomy has to be studied with ultrasound to develop a reproducible standardized methodology for routine use of ultrasound in neuromuscular imaging. To fulfill this objective, 30 subjects between the ages of 1 day and 59 years were studied with computerized real-time sonography. The results of this study are presented with illustration of the sonographic images of transverse and longitudinal planes of the upper and lower limbs. Computerization of optimum sonographic techniques, gradation of normal sonographic muscle anatomy by age, and elimination of operator variability were goals that were accomplished, thereby setting the stage for reliable, reproducible muscle imaging by ultrasound. ( J Child Neurol 1988;3:69-74).
The prevalence of Rett's syndrome was studied in a part of southwestern Sweden comprising five counties and the city of Gothenburg. In a population of 315469 children and adolescents, 6-17 years of age, 10 cases were detected, all girls. The corresponding prevalence was 0.65/10 000 girls, i.e. about twice that of phenylketonuria (PKU) in the same area. As progressive brain disorders/metabolic diseases together constitute 5-6% (1.5-2.0/10 000 children) of the aetiologies among severely mentally retarded persons of this age group in central Sweden, it can be concluded that within this group Rett's syndrome should be considered as an aetiological factor to think of in females. This syndrome might well be responsible for one-fourth to one-third of such cases among girls.
Thirty-five patients, exclusively girls, from three countries had a uniform and striking progressive encephalopathy. After normal general and psychomotor development up to the age of 7 to 18 months, developmental stagnation occurred, followed by rapid deterioration of higher brain functions. Within one-and-a-half years this deterioration led to severe dementia, autism, loss of purposeful use of the hands, jerky truncal ataxia, and acquired microcephaly. The destructive stage was followed by apparent stability lasting through decades. Additional insidious neurological abnormalities supervened, mainly spastic parapareses, vasomotor disturbances of the lower limbs, and epilepsy. Prior extensive laboratory investigations have not revealed the cause. The condition is similar to a virtually overlooked syndrome described by Rett in the German literature. The exclusive involvement of females, correlated with findings in family data analyses, suggests a dominant mutation on one X chromosome that results in affected girls and nonviable male hemizygous conceptuses.
Four families, each with two individuals affected by Rett Syndrome (RS), were analysed using restriction fragment length polymorphisms and microsatellite markers from the X chromosome. In two of the families, X-linked dominant inheritance of the RS defect from a germinally mosaic mother could be assumed. Therefore, maternal X chromosome markers showing discordant inheritance were used to exclude regions of the X chromosome as locations of the RS gene. Much of the short arm could be excluded, including regions containing three candidate genes, OTC, synapsin 1 and synaptophysin. Although most of the long arm was inherited in common it was possible to exclude a centromeric region. Inheritance of X chromosome markers is also presented for two families with affected aunt-niece pairs, one of which has not been previously studied at the DNA level.
Several techniques have recently been developed to detect single-base mismatches in DNA heteroduplexes that contain one strand of wild-type and one strand of mutated DNA. Here we tested the hypothesis that an appropriate system of mildly denaturing solvents can amplify the tendency of single-base mismatches to produce conformational changes, such as bends in the double helix, and thereby increase the differential migration of DNA heteroduplexes and homoduplexes during gel electrophoresis. The best separations of heteroduplexes and homoduplexes were obtained with a standard 6% polyacrylamide gel polymerized in 10% ethylene glycol/15% formamide/Tris-taurine buffer. As predicted by the hypothesis of solvent-induced bends, when the concentration of either ethylene glycol or formamide was increased, the differential migration decreased. Also, single-base mismatches within 50 bp of one end of a heteroduplex did not produce differential migration. Sixty of 68 single-base mismatches in a series of PCR products were detected in some 59 different sequence contexts. The eight mismatches not detected were either within 50 bp of the nearest end of the PCR product or in isolated high-melting-temperature domains. Therefore, it was possible to predict in advance the end regions and sequence contexts in which mismatches may be difficult to detect. The procedure can be applied to any PCR products of 200-800 bp and requires no special equipment or preparation of samples.
Vertebrate genomes are heavily methylated at cytosines in the sequence CpG. The biological role of this modification is probably mediated by DNA binding proteins that are either attracted to or repelled by methyl-CpG. MeCP2 is an abundant chromosomal protein that binds specifically to methylated DNA in vitro, and depends upon methyl-CpG for its chromosomal distribution in vivo. To assess the functional significance of MeCP2, the X-linked gene was mutated in male mouse embryonic stem (ES) cells using a promoterless gene-targeting construct containing a lacZ reporter gene. Mutant ES cells lacking MeCP2 grew with the same vigour as the parental line and were capable of considerable differentiation. Chimaeric embryos derived from several independent mutant lines, however, exhibited developmental defects whose severity was positively correlated with the contribution of mutant cells. The results demonstrate that MeCP2, like DNA methyltransferase, is dispensable in stem cells, but essential for embryonic development.
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The most likely cause of the Rett syndrome (RTT) is an X-linked dominant mutation lethal in hemizygous males. Previous exclusion mapping studies have identified putative regions for the RTT gene on the X chromosome. In the present study, we evaluated two candidate genes, glutamate dehydrogenase-2 (GLUD2) and rab GDP-dissociation inhibitor (GDI1/XAP-4), chosen because of their expression patterns and functions in the central nervous system and their location in the nonexcluded region of Xq. The intronless gene GLUD2, located in Xq25 and expressed in neuronal and testicular tissues, is involved in the metabolism of glutamate, a neurotransmitter reported to be elevated in the spinal fluid of RTT individuals. The GLUD2 gene was screened for mutations by Southern hybridization and by direct sequencing of polymerase chain reaction (PCR) products. The GDI1 gene in Xq28, also known as RABGDIA or XAP-4, encodes a human GDI that is expressed predominantly in neuronal and sensory tissues. All 11 exons and splice junctions of the GDI1 gene were PCR-amplified and sequenced directly or screened by single-strand conformation analysis. No mutation in either of these two genes was found in 22 RTT patients. Therefore, GLUD2 and GDI1 can be excluded as candidate genes for this syndrome.
S.N. and E.P.H. are supported by grants from the National Institutes of Health (NIH). N.S. is supported by NIH Biotechnology Training Grant IT32 GM08540 to the University of Pittsburgh. The authors thank Mark Lanasa, for assistance with the X-inactivation studies, and Paco Murillo-Martinez, for help with linkage analyses. E.P.H. is an Established Investigator of the American Heart Association.
Rett syndrome (RTT) is an X-linked dominant neurodevelopmental disorder that affects females. Exclusion mapping studies using a new family with maternal inheritance of RTT defined Xq28 as the candidate region for the RTT gene. Six candidate genes were selected for mutation analysis based on their established expression patterns and known functions in the CNS. These are: Glutamate receptor subunit 3 (GLUR3), GABA receptor subunit alpha 3 (GABRA3), GABA receptor subunit e1 (GABRE1), Vacuolar ATPase subunit 1 (VATPS1, XAP3), the human homologue of plexin 3-SEX (XAP6) and the Synaptobrevin-like protein (SYBL1). Major rearrangements involving these genes were excluded by Southern analysis. No disease-causing mutations were found, but several single-nucleotide polymorphisms (SNPs) were detected. These SNPs will be useful in future linkage analysis and whole-genome association studies for other diseases. The genomic characterization of GLUR3 and GABRA3 will allow mutational analysis of these genes as candidates for other X-linked neurological disorders mapping to Xq25-Xq26 and Xq28.
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