Mutations In methyl-CpG Binding Protein Research Articles

Genome-wide redistribution of MeCP2 in dorsal root ganglia after peripheral nerve injury.

BackgroundMethyl-CpG-binding protein 2 (MeCP2), a protein with affinity for methylated cytosines, is crucial for neuronal development and function. MeCP2 regulates gene expression through activation, repression and chromatin remodeling. Mutations in MeCP2 cause Rett syndrome, and these patients display impaired nociception. We observed an increase in MeCP2 expression in mouse dorsal root ganglia (DRG) after peripheral nerve injury. The functional implication of increased MeCP2 is largely unknown. To identify regions of the genome bound by MeCP2 in the DRG and the changes induced by nerve injury, a chromatin immunoprecipitation of MeCP2 followed by sequencing (ChIP-seq) was performed 4 weeks after spared nerve injury (SNI).ResultsWhile the number of binding sites across the genome remained similar in the SNI model and sham control, SNI induced the redistribution of MeCP2 to transcriptionally relevant regions. To determine how differential binding of MeCP2 can affect gene expression in the DRG, we investigated mmu-miR-126, a microRNA locus that had enriched MeCP2 binding in the SNI model. Enriched MeCP2 binding to miR-126 locus after nerve injury repressed miR-126 expression, and this was not mediated by alterations in methylation pattern at the miR-126 locus. Downregulation of miR-126 resulted in the upregulation of its two target genes Dnmt1 and Vegfa in Neuro 2A cells and in SNI model compared to control. These target genes were significantly downregulated in Mecp2-null mice compared to wild-type littermates, indicating a regulatory role for MeCP2 in activating Dnmt1 and Vegfa expression. Intrathecal delivery of miR-126 was not sufficient to reverse nerve injury-induced mechanical and thermal hypersensitivity, but decreased Dnmt1 and Vegfa expression in the DRG.ConclusionsOur study shows a regulatory role for MeCP2 in that changes in global redistribution can result in direct and indirect modulation of gene expression in the DRG. Alterations in genome-wide binding of MeCP2 therefore provide a molecular basis for a better understanding of epigenetic regulation-induced molecular changes underlying nerve injury.Electronic supplementary materialThe online version of this article (doi:10.1186/s13072-016-0073-5) contains supplementary material, which is available to authorized users.

Determinants of sleep disturbances in Rett syndrome: Novel findings in relation to genotype.

Rett syndrome is a rare but severe neurological disorder associated with a mutation in the methyl CpG binding protein 2 (MECP2) gene. Sleep problems and epilepsy are two of many comorbidities associated with this disorder. This study investigated the prevalence and determinants of sleep problems in Rett syndrome using an international sample. Families with a child with a confirmed Rett syndrome diagnosis and a MECP2 mutation registered in the International Rett Syndrome Phenotype Database (InterRett) were invited to participate. Questionnaires were returned by 364/461 (78.9%) either in web-based or paper format. Families completed the Sleep Disturbance Scale for Children and provided information on the presence, nature, and frequency of their child's sleep problems. Multivariate multinomial regression was used to investigate the relationships between selected sleep problems, age group, and genotype and linear regression for the relationships between sleep disturbance scales and a range of covariates. Night waking was the most prevalent sleep problem affecting over 80% with nearly half (48.3%) currently waking often at night. Initiating and maintaining sleep was most disturbed for younger children and those with a p.Arg294* mutation. Severe seizure activity was associated with poor sleep after adjusting for age group, mutation type, and mobility. We were surprised to find associations between the p.Arg294* mutation and some sleep disturbances given that other aspects of its phenotype are milder. These findings highlight the complexities of aberrant MECP2 function in Rett syndrome and explain some of the variation in manifestation of sleep disturbances. © 2016 Wiley Periodicals, Inc.

Progressive Changes in a Distributed Neural Circuit Underlie Breathing Abnormalities in Mice Lacking MeCP2.

Rett syndrome (RTT) is a neurodevelopmental disorder caused by mutations in Methyl-CpG-binding protein 2 (MECP2). Severe breathing abnormalities are common in RTT and are reproduced in mouse models of RTT. Previously, we found that removing MeCP2 from the brainstem and spinal cord in mice caused early lethality and abnormal breathing. To determine whether loss of MeCP2 in functional components of the respiratory network causes specific breathing disorders, we used the Cre/LoxP system to differentially manipulate MeCP2 expression throughout the brainstem respiratory network, specifically within HoxA4-derived tissues, which include breathing control circuitry within the nucleus tractus solitarius and the caudal part of ventral respiratory column but do not include more rostral parts of the breathing control circuitry. To determine whether respiratory phenotypes manifested in animals with MeCP2 removed from specific pons medullary respiratory circuits, we performed whole-body plethysmography and electrophysiological recordings from in vitro brainstem slices from mice lacking MeCP2 in different circuits. Our results indicate that MeCP2 expression in the medullary respiratory network is sufficient for normal respiratory rhythm and preventing apnea. However, MeCP2 expression within components of the breathing circuitry rostral to the HoxA4 domain are neither sufficient to prevent the hyperventilation nor abnormal hypoxic ventilatory response. Surprisingly, we found that MeCP2 expression in the HoxA4 domain alone is critical for survival. Our study reveals that MeCP2 is differentially required in select respiratory components for different aspects of respiratory functions, and collectively for the integrity of this network functions to maintain proper respiration. Breathing abnormalities are a significant clinical feature in Rett syndrome and are robustly reproduced in the mouse models of this disease. Previous work has established that alterations in the function of MeCP2, the protein encoded by the gene mutated in Rett syndrome, within the hindbrain are critical for control of normal breathing. Here we show that MeCP2 function plays distinct roles in specific brainstem regions in the genesis of various aspects of abnormal breathing. This provides insight into the pathogenesis of these breathing abnormalities in Rett syndrome, which could be used to target treatments to improve these symptoms. Furthermore, it provides further knowledge about the fundamental neural circuits that control breathing.

Sequential chromatin immunoprecipitation to detect SUMOylated MeCP2 in neurons

The small ubiquitin-like modifier (SUMO) is a short peptide that can be covalently linked to proteins altering their function. SUMOylation is an essential post-translational modification (PTM). Because of its dynamic nature, low abundance levels, and technical limitations, the occupation of endogenous SUMOylated transcription factors at genomic loci is challenging to detect. The chromatin regulator Methyl CpG binding protein 2 (MeCP2) is subjected to PTMs including SUMO. Mutations in MeCP2 lead to Rett syndrome, a severe neurodevelopmental disorder. Here, we present an efficient method to perform sequential chromatin immunoprecipitation (Seq-ChIP) for detecting SUMOylated MeCP2 in neurons. This Seq-ChIP technique is a useful tool to determine the occupancy of SUMOylated transcription and chromatin factors at specific genomic regions.

A novel MeCP2 acetylation site regulates interaction with ATRX and HDAC1

Methyl-CpG-binding protein-2 (MeCP2) regulates gene expression by recruiting SWI/SNF DNA helicase/ATPase (ATRX) and Histone Deacetylase-1 (HDAC1) to methylated gene regions and modulates heterochromatin association by interacting with Heterochromatin protein-1. As MeCP2 contributes to tumor suppressor gene silencing and its mutation causes Rett Syndrome, we investigated how novel post-translational-modification contributes to its function. Herein we report that upon pharmacological inhibition of SIRT1 in RKO colon and MCF-7 breast cancer cells, endogenous MeCP2 is acetylated at sites critical for binding to DNA and transcriptional regulators. We created an acetylation mimetic mutation in MeCP2 and found it to possess decreased binding to ATRX and HDAC1. Conditions inducing MeCP2 acetylation do not alter its promoter occupancy at a subset of target genes analyzed, but do cause decreased binding to ATRX and HDAC1. We also report here that a specific inhibitor of SIRT1, IV, can be used to selectively decrease H3K27me3 repressive marks on a subset of repressed target gene promoters analyzed. Lastly, we show that RKO cells over-expressing MeCP2 mutant show reduced proliferation compared to those over-expressing MeCP2-wildtype. Our study demonstrates the importance of acetylated lysine residues and suggests their key role in regulating MeCP2 function and its ability to bind transcriptional regulators.

Rett syndrome: A classic presentation

We report a 3-year-old girl with characteristic clinical, electroencephalographic and neuroradiological features of Rett syndrome (RS) - an X-linked dominant, neurodevelopmental disorder caused by mutation in methyl-CpG binding protein 2 (MECP2) gene. It predominantly affects females and is one of the commonest causes of mental retardation in females. It is characterized by progressive loss of intellectual functioning, fine and gross motor skills and communicative abilities, deceleration of head growth, and the development of stereotypic hand movements, occurring after a period of normal development. A 3-year-old girl was brought from our neighboring country with progressive loss of milestones. She had a normal development till one year of age when she gradually lost her motor milestones and started socially withdrawing from her surroundings. She also started excessive hand playing which were stereotypical and repetitive. She was microcephalic and had marked postural hypotonia. Her electroencephalography and magnetic resonance imaging showed the characteristic change associated with RS. These clinical features support the diagnosis of RS even though genetic analysis was not done. This is the first case report of RS from Pakistan.

Neurological, genetic and epigenetic features of Rett syndrome

Rett syndrome (RTT) is a severe neurodevelopmental disorder associated with mutations in the X-linked gene encoding methyl-CpG-binding protein 2 ( MECP2 ). Despite the fact that this disease has been studied for more than 20 years the problem of genetic and epigenetic factor involvement in the pathogenesis of RTT is not completely solved. In the present review we describe the contemporary knowledge of neurological, genetic and epigenetic features of RTT. We propose that subsequent studies of RTT should be targeted to reveal genotype-phenotype correlations taking into account the pathogenic effect of MECP2 mutations as well as X chromosome inactivation. We also suggest that additional investigations of epigenetic phenomena in cells with MECP2 mutations are needed in order to describe the complex interaction of genetic and epigenetic processes that leads to the myriad clinical manifestations of RTT. (J Pediatr Neurol 2004; 2(4): 179–190).

PP04.16 – 3030: Phenotypic variability with the same MECP2 mutations – An analysis of the Romanian Rett database

Objective Rett Syndrome (RTT) is an X-linked neurodevelopmental disorder that is mainly caused by mutations in Methyl-CpGbinding protein 2 (MECP2). The classical RTT picture is characterized by acquired microcephaly and a period of regression with loss of purposeful hand skills and of spoken language, gait abnormalities and stereotypic hand movements. Methods We exemplify the phenotypes of R168X and R255X mutations wich are the second and third most frequent MECP2 mutations found. The patients fom the national Rett database were analysed searching for the same mutation type. As of 2014 the database consists of 40 patients with clinical diagnosis of RTT, 27 of these patients have been genetically tested for MECP2 mutation, 16 of which are positive for a mutation in this gene. Results The patients are described regarding prehension, loss of ambulation, loss of speech, hand sterotypies presence, age at regression onset, presence of breathing anomalies; also association of epilepsy diagnosis and its response to treatment is discussed. Two groups of patients with the same mutation were identified – three girls with R255X and three girls with R168X mutation with ages between 13 months to 10 years. They all have clinical pictures of classical Rett with age at regession onset under 18 months for 2 of the girls in each group. Phenotypic and genotypic variability are frequent in RTT and this is reflected in the two groups. In the R255X group (which is associated with epilepsy in the literature) one girl had early onset drug-resistant epilepsy and severe clinical regression; the other kept prehension and gait, has mild-moderate mental retardation and no epilepsy. Conclusion No significant clinical-genetic correlations could be realised, due to the small number of identical mutations in our group. More factors (genetic or nongenetic) that influence RTT clinical picture and outcome may be involved.

713. IntraCSF Administration of AAV9/MeCP2 Extends Lifespan of MeCP2-Null Mice While Preventing Toxicity Associated With IV Administration

Rett Syndrome (RTT) is a neurodevelopmental disorder that shortens the lifespan of patients while depriving them of the ability to walk, talk, or interact with others. RTT is caused by inactivating mutations in the X chromosome-linked gene encoding methyl-CpG-binding protein 2 (MeCP2), a transcription regulator that is highly expressed in neurons. Patients with RTT express mutant MeCP2 in ~50% of their cells due to random X-chromosome inactivation in each cell. The remaining “healthy” cells have an activated chromosome encoding functional (WT) MeCP2. Although researchers have already used MeCP2 gene therapy to reverse the RTT phenotype in mosaic mice, concerns about side effects (i.e., motor and learning deficits) resulting from MeCP2 overexpression in “healthy” cells persist. Indeed, humans suffering from MeCP2 duplication syndrome have been shown to suffer from muscular spasticity and intellectual disabilities. In addition to persistent concerns about MeCP2 overexpression, IV administration of the scAAV9/hMeCP2 vector (1×1011 vg/mouse) has been shown to cause significant liver toxicity in MeCP2-null mice (Gadalla et al, Mol Ther, 2013). We tested the hypothesis that by moving from an IV route of administration to an intraCSF route, we could lower the dose of AAV vector enough to avoid the high peripheral organ transduction that generates toxic side effects. Modeling our earlier experiment with an IV route, juvenile (4-5 week old) knock-out (KO) mice and WT littermates received a single intrathecal injection of AAV9/MeCP2 vector. RTT mice dosed intrathecally with 1×1010 vg had a 48% increase in median survival compared to vehicle-injected mice (n=6 treated; n=11 vehicle; P=0.008), nearly identical to the rescue we previously observed following the IV approach but at 1/10th the dose and avoiding overexpression-related toxicity in the liver. In conclusion, by changing the route of administration of the AAV9/MeCP2 vector from IV to intraCSF, we were able to generate a nearly identical therapeutic outcome with a 10-fold lower dose, and without excessive (toxic) gene transfer to peripheral organs. As a next step to utilize the intraCSF route of administration, DNA shuffling and directed evolution were utilized to select for AAV capsid variants that would have higher efficiency and improved biodistribution in the RTT mice after intraCSF administration. Multiple clones were selected that have CNS transduction equivalent to AAV9, but importantly, all had highly reduced peripheral organ biodistribution. Preliminary studies using two clones, RTTF1.04 and RTTF1.11, were able to confer significant survival benefits to male RTT KO mice (34% and 25% prolongation of lifespan, P = 0.03 and 0.05, respectively) after delivery of the MeCP2 transgene at a dose of 1×1010 vg. Overall, our studies support the use of an intraCSF route of administration to treat RTT using AAV vectors, and the potential to develop safer and more effective AAV-based vectors tailored to the intraCSF route of administration.We acknowledge generous support from IRSF and RSRT.

Rett syndrome: a complex disorder with simple roots.

Rett syndrome (RTT) is a severe neurological disorder caused by mutations in the X-linked gene MECP2 (methyl-CpG-binding protein 2). Two decades of research have fostered the view that MeCP2 is a multifunctional chromatin protein that integrates diverse aspects of neuronal biology. More recently, studies have focused on specific RTT-associated mutations within the protein. This work has yielded molecular insights into the critical functions of MeCP2 that promise to simplify our understanding of RTT pathology.

Rett syndrome like phenotypes in the R255X Mecp2 mutant mouse are rescued by MECP2 transgene.

Rett syndrome (RTT) is a severe neurodevelopmental disorder that is usually caused by mutations in Methyl-CpG-binding Protein 2 (MECP2). Four of the eight common disease causing mutations in MECP2 are nonsense mutations and are responsible for over 35% of all cases of RTT. A strategy to overcome disease-causing nonsense mutations is treatment with nonsense mutation suppressing drugs that allow expression of full-length proteins from mutated genes with premature in-frame stop codons. To determine if this strategy is useful in RTT, we characterized a new mouse model containing a knock-in nonsense mutation (p.R255X) in the Mecp2 locus (Mecp2(R255X)). To determine whether the truncated gene product acts as a dominant negative allele and if RTT-like phenotypes could be rescued by expression of wild-type protein, we genetically introduced an extra copy of MECP2 via an MECP2 transgene. The addition of MECP2 transgene to Mecp2(R255X) mice abolished the phenotypic abnormalities and resulted in near complete rescue. Expression of MECP2 transgene Mecp2(R255X) allele also rescued mTORC1 signaling abnormalities discovered in mice with loss of function and overexpression of Mecp2. Finally, we treated Mecp2(R255X) embryonic fibroblasts with the nonsense mutation suppressing drug gentamicin and we were able to induce expression of full-length MeCP2 from the mutant p.R255X allele. These data provide proof of concept that the p.R255X mutation of MECP2 is amenable to the nonsense suppression therapeutic strategy and provide guidelines for the extent of rescue that can be expected by re-expressing MeCP2 protein.

The increase in body weight induced by lack of methyl CpG binding protein‐2 is associated with altered leptin signalling in the hypothalamus

Methyl CpG binding protein-2 (MECP2) is a chromatin-remodelling factor with a dual role in gene expression. Evidence from patients carrying MECP2 mutations and from transgenic mouse models demonstrates that this protein is involved in the control of body weight. However, the mechanism for this has not been fully elucidated. To address this, we used a previously characterized Mecp2-null mouse model and found that the increase in body weight is associated with an increased amount of adipose tissue and high leptin levels. Appropriate body weight control requires the proper expression of pro-opiomelanocortin (Pomc) and agouti-related peptide (Agrp), two neuropeptides essential for satiety and appetite signals, respectively. Our results show that in the absence of Mecp2, Pomc and Agrp mRNA expression are altered, and the mice are leptin resistant. To determine the mechanism underlying the defective leptin sensing, we evaluated the expression of genes and the post-translational modifications associated with leptin signalling, which are fundamental to Pomc and Agrp transcriptional control and proper leptin response. We found a decrease in the phosphorylation level of Akt and its target protein Foxo1, which indicate an alteration in leptin-induced signal transduction. Our results demonstrate that the absence of Mecp2 disrupted body weight balance by altering post-translational modifications in leptin-signalling components that regulate Pomc and Agrp expression.

Rett Syndrome and MeCP2

Rett syndrome (RTT) is a severe and progressive neurological disorder, which mainly affects young females. Mutations of the methyl-CpG binding protein 2 (MECP2) gene are the most prevalent cause of classical RTT cases. MECP2 mutations or altered expression are also associated with a spectrum of neurodevelopmental disorders such as autism spectrum disorders with recent links to fetal alcohol spectrum disorders. Collectively, MeCP2 relation to these neurodevelopmental disorders highlights the importance of understanding the molecular mechanisms by which MeCP2 impacts brain development, mental conditions, and compromised brain function. Since MECP2 mutations were discovered to be the primary cause of RTT, a significant progress has been made in the MeCP2 research, with respect to the expression, function and regulation of MeCP2 in the brain and its contribution in RTT pathogenesis. To date, there have been intensive efforts in designing effective therapeutic strategies for RTT benefiting from mouse models and cells collected from RTT patients. Despite significant progress in MeCP2 research over the last few decades, there is still a knowledge gap between the in vitro and in vivo research findings and translating these findings into effective therapeutic interventions in human RTT patients. In this review, we will provide a synopsis of Rett syndrome as a severe neurological disorder and will discuss the role of MeCP2 in RTT pathophysiology.

Respiratory phenotypes are distinctly affected in mice with common Rett syndrome mutations MeCP2 T158A and R168X

Respiratory disturbances are a primary phenotype of the neurological disorder, Rett syndrome (RTT), caused by mutations in the X-linked gene encoding methyl-CpG-binding protein 2 (MeCP2). Mouse models generated with null mutations in Mecp2 mimic respiratory abnormalities in RTT girls. Large deletions, however, are seen in only ∼10% of affected human individuals. Here we characterized respiration in heterozygous females from two mouse models that genetically mimic common RTT point mutations, a missense mutation T158A (Mecp2T158A/+) or a nonsense mutation R168X (Mecp2R168X/+). MeCP2 T158A shows decreased binding to methylated DNA, while MeCP2 R168X retains the capacity to bind methylated DNA but lacks the ability to recruit complexes required for transcriptional repression. We found that both Mecp2T158A/+ and Mecp2R168X/+ heterozygotes display augmented hypoxic ventilatory responses and depressed hypercapnic responses, compared to wild-type controls. Interestingly, the incidence of apnea was much greater in Mecp2R168X/+ heterozygotes, 189 per hour, than Mecp2T158A/+ heterozygotes, 41 per hour. These results demonstrate that different RTT mutations lead to distinct respiratory phenotypes, suggesting that characterization of the respiratory phenotype may reveal functional differences between MeCP2 mutations and provide insights into the pathophysiology of RTT.

TALEN-Mediated Gene Mutagenesis in Rhesus and Cynomolgus Monkeys

Recent advances in gene editing technology have introduced the potential for application of mutagenesis approaches in nonhuman primates to model human development and disease. Here we report successful TALEN-mediated mutagenesis of an X-linked, Rett syndrome (RTT) gene, methyl-CpG binding protein 2 (MECP2), in both rhesus and cynomolgus monkeys. Microinjection of MECP2-targeting TALEN plasmids into rhesus and cynomolgus zygotes leads to effective gene editing of MECP2 with no detected off-target mutagenesis. Male rhesus (2) and cynomolgous (1) fetuses carrying MECP2 mutations in various tissues including testes were miscarried during midgestation, consistent with RTT-linked male embryonic lethality in humans. One live delivery of a female cynomolgus monkey occurred after 162days of gestation, with abundant MECP2 mutations in peripheral tissues. We conclude that TALEN-mediated mutagenesis can be an effective tool for genetic modeling of human disease in nonhuman primates.

MeCP2 is required for activity-dependent refinement of olfactory circuits

Methyl CpG binding protein 2 (MeCP2) is a structural chromosomal protein involved in the regulation of gene expression. Alterations in the levels of MeCP2 have been related to neurodevelopmental disorders. Studies in mouse models of MeCP2 deficiency have demonstrated that this protein is important for neuronal maturation, neurite complexity, synaptogenesis, and synaptic plasticity. However, the mechanisms by which MeCP2 dysfunction leads to neurodevelopmental defects, and the role of activity, remain unclear, as most studies examine the adult nervous system, which may obfuscate the primary consequences of MeCP2 mutation. We hypothesize that MeCP2 plays a role during the formation and activity-driven maturation of neural circuits at early postnatal stages. To test this hypothesis, we use the olfactory system as a neurodevelopmental model. This system undergoes postnatal neurogenesis; axons from olfactory neurons form highly stereotyped projections to higher-order neurons, facilitating the detection of possible defects in the establishment of connectivity. In vivo olfactory stimulation paradigms were used to produce physiological synaptic activity in gene-targeted mice in which specific olfactory circuits are visualized. Our results reveal defective postnatal refinement of olfactory circuits in Mecp2 knock out (KO) mice after sensory (odorant) stimulation. This failure in refinement was associated with deficits in the normal responses to odorants, including brain-derived neurotrophic factor (BDNF) production, as well as changes in adhesion molecules known to regulate axonal convergence. The defective refinement observed in Mecp2 KO mice was prevented by daily treatment with ampakine beginning after the first postnatal week. These observations indicate that increasing synaptic activity at early postnatal stage might circumvent the detrimental effect of MeCP2 deficiency on circuitry maturation. The present results provide in vivo evidence in real time for the role of MeCP2 in activity-dependent maturation of olfactory circuitry, with implications for understanding the mechanism of MeCP2 mutations in the development of neural connectivity.

Methyl-CpG-binding protein 2 (MECP2) mutation type is associated with disease severity in Rett syndrome

BackgroundRett syndrome (RTT), a neurodevelopmental disorder that primarily affects girls, is characterised by a period of apparently normal development until 6–18 months of age when motor and communication abilities regress....

Genetic Effects on Cerebellar Structure Across Mouse Models of Autism Using a Magnetic Resonance Imaging Atlas

Magnetic resonance imaging (MRI) of autism populations is confounded by the inherent heterogeneity in the individuals' genetics and environment, two factors difficult to control for. Imaging genetic animal models that recapitulate a mutation associated with autism quantify the impact of genetics on brain morphology and mitigate the confounding factors in human studies. Here, we used MRI to image three genetic mouse models with single mutations implicated in autism: Neuroligin-3 R451C knock-in, Methyl-CpG binding protein-2 (MECP2) 308-truncation and integrin β3 homozygous knockout. This study identified the morphological differences specific to the cerebellum, a structure repeatedly linked to autism in human neuroimaging and postmortem studies. To accomplish a comparative analysis, a segmented cerebellum template was created and used to segment each study image. This template delineated 39 different cerebellar structures. For Neuroligin-3 R451C male mutants, the gray (effect size (ES) = 1.94, FDR q = 0.03) and white (ES = 1.84, q = 0.037) matter of crus II lobule and the gray matter of the paraflocculus (ES = 1.45, q = 0.045) were larger in volume. The MECP2 mutant mice had cerebellar volume changes that increased in scope depending on the genotype: hemizygous males to homozygous females. The integrin β3 mutant mouse had a drastically smaller cerebellum than controls with 28 out of 39 cerebellar structures smaller. These imaging results are discussed in relation to repetitive behaviors, sociability, and learning in the context of autism. This work further illuminates the cerebellum's role in autism.

Transcriptional regulation in pluripotent stem cells by methyl CpG-binding protein 2 (MeCP2)

Rett syndrome (RTT) is one of the most prevalent female mental disorders. De novo mutations in methyl CpG-binding protein 2 (MeCP2) are a major cause of RTT. MeCP2 regulates gene expression as a transcription regulator as well as through long-range chromatin interaction. Because MeCP2 is present on the X chromosome, RTT is manifested in an X-linked dominant manner. Investigation using murine MeCP2 null models and post-mortem human brain tissues has contributed to understanding the molecular and physiological function of MeCP2. In addition, RTT models using human induced pluripotent stem cells derived from RTT patients (RTT-iPSCs) provide novel resources to elucidate the regulatory mechanism of MeCP2. Previously, we obtained clones of female RTT-iPSCs that express either wild-type or mutant MECP2 due to the inactivation of one X chromosome. Reactivation of the X chromosome also allowed us to have RTT-iPSCs that express both wild-type and mutant MECP2. Using these unique pluripotent stem cells, we investigated the regulation of gene expression by MeCP2 in pluripotent stem cells by transcriptome analysis. We found that MeCP2 regulates genes encoding mitochondrial membrane proteins. In addition, loss of function in MeCP2 results in de-repression of genes on the inactive X chromosome. Furthermore, we showed that each mutation in MECP2 affects a partly different set of genes. These studies suggest that fundamental cellular physiology is affected by mutations in MECP2 from early development, and that a therapeutic approach targeting to unique forms of mutant MeCP2 is needed.

Transcriptional regulation of the MET receptor tyrosine kinase gene by MeCP2 and sex-specific expression in autism and Rett syndrome

Single nucleotide variants (SNV) in the gene encoding the MET receptor tyrosine kinase have been associated with an increased risk for autism spectrum disorders (ASD). The MET promoter SNV rs1858830 C ‘low activity' allele is enriched in ASD, associated with reduced protein expression, and impacts functional and structural circuit connectivity in humans. To gain insight into the transcriptional regulation of MET on ASD-risk etiology, we examined an interaction between the methyl CpG-binding protein 2 (MeCP2) and the MET 5′ promoter region. Mutations in MeCP2 cause Rett syndrome (RTT), a predominantly female neurodevelopmental disorder sharing some ASD clinical symptoms. MeCP2 binds to a region of the MET promoter containing the ASD-risk SNV, and displays rs1858830 genotype-specific binding in human neural progenitor cells derived from the olfactory neuroepithelium. MeCP2 binding enhances MET expression in the presence of the rs1858830 C allele, but MET transcription is attenuated by RTT-specific mutations in MeCP2. In the postmortem temporal cortex, a region normally enriched in MET, gene expression is reduced dramatically in females with RTT, although not due to enrichment of the rs1858830 C ‘low activity' allele. We newly identified a sex-based reduction in MET expression, with male ASD cases, but not female ASD cases compared with sex-matched controls. The experimental data reveal a prominent allele-specific regulation of MET transcription by MeCP2. The mechanisms underlying the pronounced reduction of MET in ASD and RTT temporal cortex are distinct and likely related to factors unique to each disorder, including a noted sex bias.