ABSTRACT
Genes encoding the KDM5 family of transcriptional regulators are disrupted in individuals with intellectual disability (ID). To understand the link between KDM5 and ID, we characterized five Drosophila strains harboring missense alleles analogous to those observed in patients. These alleles disrupted neuroanatomical development, cognition and other behaviors, and displayed a transcriptional signature characterized by the downregulation of many ribosomal protein genes. A similar transcriptional profile was observed in KDM5C knockout iPSC-induced human glutamatergic neurons, suggesting an evolutionarily conserved role for KDM5 proteins in regulating this class of gene. In Drosophila, reducing KDM5 changed neuronal ribosome composition, lowered the translation efficiency of mRNAs required for mitochondrial function, and altered mitochondrial metabolism. These data highlight the cellular consequences of altered KDM5-regulated transcriptional programs that could contribute to cognitive and behavioral phenotypes. Moreover, they suggest that KDM5 may be part of a broader network of proteins that influence cognition by regulating protein synthesis.
Subject(s)
Drosophila Proteins , Neurons , Ribosomal Proteins , Animals , Humans , Drosophila/genetics , Drosophila/metabolism , Drosophila melanogaster/genetics , Drosophila melanogaster/metabolism , Drosophila Proteins/genetics , Drosophila Proteins/metabolism , Histone Demethylases/metabolism , Histone Demethylases/genetics , Intellectual Disability/genetics , Intellectual Disability/metabolism , Mitochondria/metabolism , Mitochondria/genetics , Neurons/metabolism , Protein Biosynthesis , Ribosomal Proteins/genetics , Ribosomal Proteins/metabolism , Ribosomes/metabolism , Ribosomes/genetics , Transcriptional ActivationABSTRACT
22q11.2 deletion is one of the strongest known genetic risk factors for schizophrenia. Recent whole-genome sequencing of schizophrenia cases and controls with this deletion provided an unprecedented opportunity to identify risk modifying genetic variants and investigate their contribution to the pathogenesis of schizophrenia in 22q11.2 deletion syndrome. Here, we apply a novel analytic framework that integrates gene network and phenotype data to investigate the aggregate effects of rare coding variants and identified modifier genes in this etiologically homogenous cohort (223 schizophrenia cases and 233 controls of European descent). Our analyses revealed significant additive genetic components of rare nonsynonymous variants in 110 modifier genes (adjusted P = 9.4E-04) that overall accounted for 4.6% of the variance in schizophrenia status in this cohort, of which 4.0% was independent of the common polygenic risk for schizophrenia. The modifier genes affected by rare coding variants were enriched with genes involved in synaptic function and developmental disorders. Spatiotemporal transcriptomic analyses identified an enrichment of coexpression between modifier and 22q11.2 genes in cortical brain regions from late infancy to young adulthood. Corresponding gene coexpression modules are enriched with brain-specific protein-protein interactions of SLC25A1, COMT, and PI4KA in the 22q11.2 deletion region. Overall, our study highlights the contribution of rare coding variants to the SCZ risk. They not only complement common variants in disease genetics but also pinpoint brain regions and developmental stages critical to the etiology of syndromic schizophrenia.
Subject(s)
DiGeorge Syndrome , Schizophrenia , Humans , Young Adult , Adult , Schizophrenia/genetics , DiGeorge Syndrome/genetics , Brain , Gene Expression Profiling , Whole Genome SequencingABSTRACT
Postinfectious neuroinflammation has been implicated in multiple models of acute-onset obsessive-compulsive disorder including Sydenham chorea (SC), pediatric acute-onset neuropsychiatric syndrome (PANS), and pediatric autoimmune neuropsychiatric disorders associated with streptococcal infection (PANDAS). These conditions are associated with a range of autoantibodies which are thought to be triggered by infections, most notably group A streptococci (GAS). Based on animal models using huma sera, these autoantibodies are thought to cross-react with neural antigens in the basal ganglia and modulate neuronal activity and behavior. As is true for many childhood neuroinflammatory diseases and rheumatological diseases, SC, PANS, and PANDAS lack clinically available, rigorous diagnostic biomarkers and randomized clinical trials. In this review article, we outline the accumulating evidence supporting the role neuroinflammation plays in these disorders. We describe work with animal models including patient-derived anti-neuronal autoantibodies, and we outline imaging studies that show alterations in the basal ganglia. In addition, we present research on metabolites, which are helpful in deciphering functional phenotypes, and on the implication of sleep in these disorders. Finally, we encourage future researchers to collaborate across medical specialties (e.g., pediatrics, psychiatry, rheumatology, immunology, and infectious disease) in order to further research on clinical syndromes presenting with neuropsychiatric manifestations.
Subject(s)
Chorea , Obsessive-Compulsive Disorder , Streptococcal Infections , Animals , Child , Humans , Autoimmunity , Chorea/diagnosis , Chorea/complications , Neuroinflammatory Diseases , Streptococcal Infections/complications , Streptococcal Infections/diagnosis , Streptococcal Infections/drug therapy , Obsessive-Compulsive Disorder/diagnosis , Obsessive-Compulsive Disorder/psychology , Autoantibodies/therapeutic use , InflammationABSTRACT
Jansen-de Vries syndrome (JdVS) is a neurodevelopmental condition attributed to pathogenic variants in Exons 5 and 6 of PPM1D. As the full phenotypic spectrum and natural history remain to be defined, we describe a large cohort of children and adults with JdVS. This is a retrospective cohort study of 37 individuals from 34 families with disease-causing variants in PPM1D leading to JdVS. Clinical data were provided by treating physicians and/or families. Of the 37 individuals, 27 were male and 10 female, with median age 8.75 years (range 8 months to 62 years). Four families document autosomal dominant transmission, and 32/34 probands were diagnosed via exome sequencing. The facial gestalt, including a broad forehead and broad mouth with a thin and tented upper lip, was most recognizable between 18 and 48 months of age. Common manifestations included global developmental delay (35/36, 97%), hypotonia (25/34, 74%), short stature (14/33, 42%), constipation (22/31, 71%), and cyclic vomiting (6/35, 17%). Distinctive personality traits include a hypersocial affect (21/31, 68%) and moderate-to-severe anxiety (18/28, 64%). In conclusion, JdVS is a clinically recognizable neurodevelopmental syndrome with a characteristic personality and distinctive facial features. The association of pathogenic variants in PPM1D with cyclic vomiting bears not only medical attention but also further pathogenic and mechanistic evaluation.
Subject(s)
Intellectual Disability , Neurodevelopmental Disorders , Adult , Child , Female , Humans , Infant , Male , Developmental Disabilities/diagnosis , Developmental Disabilities/genetics , Intellectual Disability/diagnosis , Intellectual Disability/genetics , Intellectual Disability/pathology , Neurodevelopmental Disorders/diagnosis , Neurodevelopmental Disorders/epidemiology , Neurodevelopmental Disorders/genetics , Phenotype , Protein Phosphatase 2C/genetics , Retrospective Studies , Vomiting , Child, Preschool , Adolescent , Young Adult , Middle AgedABSTRACT
BACKGROUND: Monoallelic expression of autosomal genes has been implicated in human psychiatric disorders. However, there is a paucity of allelic expression studies in human brain cells at the single cell and genome wide levels. RESULTS: In this report, we reanalyzed a previously published single-cell RNA-seq dataset from several postmortem human brains and observed pervasive monoallelic expression in individual cells, largely in a random manner. Examining single nucleotide variants with a predicted functional disruption, we found that the "damaged" alleles were overall expressed in fewer brain cells than their counterparts, and at a lower level in cells where their expression was detected. We also identified many brain cell type-specific monoallelically expressed genes. Interestingly, many of these cell type-specific monoallelically expressed genes were enriched for functions important for those brain cell types. In addition, function analysis showed that genes displaying monoallelic expression and correlated expression across neuronal cells from different individual brains were implicated in the regulation of synaptic function. CONCLUSIONS: Our findings suggest that monoallelic gene expression is prevalent in human brain cells, which may play a role in generating cellular identity and neuronal diversity and thus increasing the complexity and diversity of brain cell functions.
Subject(s)
Alleles , Brain/cytology , Gene Expression Profiling , Sequence Analysis, RNA , Single-Cell Analysis , Humans , Polymorphism, Single NucleotideABSTRACT
Induced pluripotent stem cell (iPSC)-derived neurons and neural progenitors are great resources for studying neural development and differentiation and their disruptions in disease conditions, and hold the promise of future cell therapy. In general, iPSC lines can be established either specifically from patients with neuropsychiatric disorders or from healthy subjects. The iPSCs can then be induced to differentiate into neural lineages and the iPSC-derived neurons are valuable for various types of cell-based assays that seek to understand disease mechanisms and identify and test novel therapies. In addition, it is an ideal system for gene expression profiling (i.e., transcriptomic analysis), an efficient and cost-effective way to explore the genetic programs regulating neurodevelopment. Moreover, transcriptomic comparison, which can be performed between patient-derived samples and controls, or in control lines in which the expression of specific genes has been disrupted, can uncover convergent gene targets and pathways that are downstream of the hundreds of candidate genes that have been associated with neuropsychiatric disorders. The results, especially after integration with spatiotemporal transcriptomic profiles of normal human brain development, have indeed helped to uncover gene networks, molecular pathways, and cellular signaling that likely play critical roles in disease development and progression. On the other hand, despite the great promise, many challenges remain in the usage of iPSC-derived neurons for modeling neuropsychiatric disorders, for example, how to generate relatively homogenous populations of specific neuronal subtypes that are affected in a particular disorder and how to better address the genetic heterogeneity that exists in the patient population.
Subject(s)
Brain Diseases/metabolism , Cell Culture Techniques/methods , Induced Pluripotent Stem Cells/metabolism , Mental Disorders/metabolism , Neurons/metabolism , Transcriptome , Brain Diseases/genetics , Brain Diseases/pathology , Humans , Induced Pluripotent Stem Cells/cytology , Mental Disorders/genetics , Mental Disorders/pathology , Models, Biological , Neurons/cytology , Polymorphism, GeneticABSTRACT
Recent studies have shown that the transcriptional functions of REST are much broader than repressing neuronal genes in non-neuronal systems. Whether REST occupies similar chromatin regions in different cell types and how it interacts with other transcriptional regulators to execute its functions in a context-dependent manner has not been adequately investigated. We have applied ChIP-seq analysis to identify the REST cistrome in human CD4+ T cells and compared it with published data from 15 other cell types. We found that REST cistromes were distinct among cell types, with REST binding to several tumor suppressors specifically in cancer cells, whereas 7% of the REST peaks in non-neuronal cells were ubiquitously called and <25% were identified for ≥ 5 cell types. Nevertheless, using a quantitative metric directly comparing raw ChIP-seq signals, we found the majority (â¼80%) was shared by ≥ 2 cell types. Integration with RNA-seq data showed that REST binding was generally correlated with low gene expression. Close examination revealed that multiple contexts were correlated with reduced expression of REST targets, e.g., the presence of a cognate RE1 motif and cellular specificity of REST binding. These contexts were shown to play a role in differential corepressor recruitment. Furthermore, transcriptional outcome was highly influenced by REST cofactors, e.g., SIN3 and EZH2 co-occupancy marked higher and lower expression of REST targets, respectively. Unexpectedly, the REST cistrome in differentiated neurons exhibited unique features not observed in non-neuronal cells, e.g., the lack of RE1 motifs and an association with active gene expression. Finally, our analysis demonstrated how REST could differentially regulate a transcription network constituted of miRNAs, REST complex and neuronal factors. Overall, our findings of contexts playing critical roles in REST occupancy and regulatory outcome provide insights into the molecular interactions underlying REST's diverse functions, and point to novel roles of REST in differentiated neurons.
Subject(s)
CD4-Positive T-Lymphocytes/metabolism , Gene Expression Regulation/genetics , Genomics/methods , Neurons/metabolism , Repressor Proteins/genetics , Animals , Cell Line , Chromatin Immunoprecipitation , Gene Silencing , Histones/genetics , Humans , Mice , MicroRNAs/geneticsABSTRACT
More than 100 genes have been associated with significantly increased risks of autism spectrum disorders (ASD) with an estimate of â¼1000 genes that may contribute. The new challenge is to investigate the molecular and cellular functions of these genes during neural and brain development, and then even more challenging, to link the altered molecular and cellular phenotypes to the ASD clinical manifestations. In this study, we used single-cell RNA-seq analysis to study one of the top risk genes, CHD8, in cerebral organoids, which models early neural development. We identified 21 cell clusters in the organoid samples, representing non-neuronal cells, neural progenitors, and early differentiating neurons at the start of neural cell fate commitment. Comparisons of the cells with one copy of a CHD8 knockout allele, generated by CRISPR/Cas9 editing, and their isogenic controls uncovered thousands of differentially expressed genes, which were enriched with functions related to neural and brain development, cilium organization, and extracellular matrix organization. The affected genes were also enriched with genes and pathways previously implicated in ASD, but surprisingly not for schizophrenia and intellectual disability risk genes. The comparisons also uncovered cell composition changes, indicating potentially altered neural differential trajectories upon CHD8 reduction. Moreover, we found that cell-cell communications were affected in the CHD8 knockout organoids, including the interactions between neural and glial cells. Taken together, our results provide new data and information for understanding CHD8 functions in the early stages of neural lineage development and interaction.
ABSTRACT
About 100 genes have been associated with significantly increased risks of autism spectrum disorders (ASD) with an estimate of ~1000 genes that may be involved. The new challenge now is to investigate the molecular and cellular functions of these genes during neural and brain development, and then even more challenging, to link the altered molecular and cellular phenotypes to the ASD clinical manifestations. In this study, we use single cell RNA-seq analysis to study one of the top risk gene, CHD8, in cerebral organoids, which models early neural development. We identify 21 cell clusters in the organoid samples, representing non-neuronal cells, neural progenitors, and early differentiating neurons at the start of neural cell fate commitment. Comparisons of the cells with one copy of the CHD8 knockout and their isogenic controls uncover thousands of differentially expressed genes, which are enriched with function related to neural and brain development, with genes and pathways previously implicated in ASD, but surprisingly not for Schizophrenia and intellectual disability risk genes. The comparisons also find cell composition changes, indicating potential altered neural differential trajectories upon CHD8 reduction. Moreover, we find that cell-cell communications are affected in the CHD8 knockout organoids, including the interactions between neural and glial cells. Taken together, our results provide new data for understanding CHD8 functions in the early stages of neural lineage development and interaction.
ABSTRACT
Background: Jansen de Vries Syndrome (JdVS) is a rare neurodevelopmental disorder (NDD) caused by gain-of-function (GOF) truncating mutations in PPM1D exons 5 or 6. PPM1D is a serine/threonine phosphatase that plays an important role in the DNA damage response (DDR) by negatively regulating TP53 (P53). JdVS-associated mutations lead to the formation of a truncated PPM1D protein that retains catalytic activity and has a GOF effect because of reduced degradation. Somatic PPM1D exons 5 and 6 truncating mutations are well-established factors in a number of cancers, due to excessive dephosphorylation and reduced function of P53 and other substrates involved in DDR. Children with JdVS have a variety of neurodevelopmental, psychiatric, and physical problems. In addition, a small fraction has acute neuropsychiatric decompensation apparently triggered by infection or severe non-infectious environmental stress factors. Methods: To understand the molecular basis of JdVS, we developed an induced pluripotent stem cell (iPSC) model system. iPSCs heterozygous for the truncating variant (PPM1D+/tr), were made from a patient, and control lines engineered using CRISPR-Cas9 gene editing. Proteomics and phosphoprotemics analyses were carried out on iPSC-derived glutamatergic neurons and microglia from three control and three PPM1D+/tr iPSC lines. We also analyzed the effect of the TLR4 agonist, lipopolysaccharide, to understand how activation of the innate immune system in microglia could account for acute behavioral decompensation. Results: One of the major findings was the downregulation of POGZ in unstimulated microglia. Since loss-of-function variants in the POGZ gene are well-known causes of autism spectrum disorder, the decrease in PPM1D+/tr microglia suggests this plays a role in the neurodevelopmental aspects of JdVS. In addition, neurons, baseline, and LPS-stimulated microglia show marked alterations in the expression of several E3 ubiquitin ligases, most notably UBR4, and regulators of innate immunity, chromatin structure, ErbB signaling, and splicing. In addition, pathway analysis points to overlap with neurodegenerative disorders. Limitations: Owing to the cost and labor-intensive nature of iPSC research, the sample size was small. Conclusions: Our findings provide insight into the molecular basis of JdVS and can be extrapolated to understand neuropsychiatric decompensation that occurs in subgroups of patients with ASD and other NDDs.
ABSTRACT
BACKGROUND: Autism spectrum disorder is a neurodevelopmental disorder, affecting 1-2% of children. Studies have revealed genetic and cellular abnormalities in the brains of affected individuals, leading to both regional and distal cell communication deficits. METHODS: Recent application of single-cell technologies, especially single-cell transcriptomics, has significantly expanded our understanding of brain cell heterogeneity and further demonstrated that multiple cell types and brain layers or regions are perturbed in autism. The underlying high-dimensional single-cell data provides opportunities for multilevel computational analysis that collectively can better deconvolute the molecular and cellular events altered in autism. Here, we apply advanced computation and pattern recognition approaches on single-cell RNA-seq data to infer and compare inter-cell-type signaling communications in autism brains and controls. RESULTS: Our results indicate that at a global level, there are cell-cell communication differences in autism in comparison with controls, largely involving neurons as both signaling senders and receivers, but glia also contribute to the communication disruption. Although the magnitude of changes is moderate, we find that excitatory and inhibitor neurons are involved in multiple intercellular signaling that exhibits increased strengths in autism, such as NRXN and CNTN signaling. Not all genes in the intercellular signaling pathways show differential expression, but genes in the affected pathways are enriched for axon guidance, synapse organization, neuron migration, and other critical cellular functions. Furthermore, those genes are highly connected to and enriched for genes previously associated with autism risks. CONCLUSIONS: Overall, our proof-of-principle computational study using single-cell data uncovers key intercellular signaling pathways that are potentially disrupted in the autism brains, suggesting that more studies examining cross-cell type effects can be valuable for understanding autism pathogenesis.
Subject(s)
Autism Spectrum Disorder , Autistic Disorder , Autism Spectrum Disorder/pathology , Autistic Disorder/genetics , Brain/pathology , Cell Communication , Child , Humans , TranscriptomeABSTRACT
Abrupt onset of severe neuropsychiatric symptoms including obsessive-compulsive disorder, tics, anxiety, mood swings, irritability, and restricted eating is described in children with Pediatric Acute-Onset Neuropsychiatric Syndrome (PANS). Symptom onset is often temporally associated with infections, suggesting an underlying autoimmune/autoinflammatory etiology, although direct evidence is often lacking. The pathological mechanisms are likely heterogeneous, but we hypothesize convergence on one or more biological pathways. Consequently, we conducted whole exome sequencing (WES) on a U.S. cohort of 386 cases, and whole genome sequencing (WGS) on ten cases from the European Union who were selected because of severe PANS. We focused on identifying potentially deleterious genetic variants that were de novo or ultra-rare (MAF) < 0.001. Candidate mutations were found in 11 genes (PPM1D, SGCE, PLCG2, NLRC4, CACNA1B, SHANK3, CHK2, GRIN2A, RAG1, GABRG2, and SYNGAP1) in 21 cases, which included two or more unrelated subjects with ultra-rare variants in four genes. These genes converge into two broad functional categories. One regulates peripheral immune responses and microglia (PPM1D, CHK2, NLRC4, RAG1, PLCG2). The other is expressed primarily at neuronal synapses (SHANK3, SYNGAP1, GRIN2A, GABRG2, CACNA1B, SGCE). Mutations in these neuronal genes are also described in autism spectrum disorder and myoclonus-dystonia. In fact, 12/21 cases developed PANS superimposed on a preexisting neurodevelopmental disorder. Genes in both categories are also highly expressed in the enteric nervous system and the choroid plexus. Thus, genetic variation in PANS candidate genes may function by disrupting peripheral and central immune functions, neurotransmission, and/or the blood-CSF/brain barriers following stressors such as infection.
Subject(s)
Autism Spectrum Disorder , Autoimmune Diseases , Obsessive-Compulsive Disorder , Streptococcal Infections , Autism Spectrum Disorder/complications , Autism Spectrum Disorder/genetics , Autoimmune Diseases/diagnosis , Child , Exome/genetics , Homeodomain Proteins , Humans , Obsessive-Compulsive Disorder/diagnosis , Streptococcal Infections/complications , Exome Sequencing , Whole Genome SequencingABSTRACT
Induced pluripotent stem cell (iPSC) technology has the potential to transform regenerative medicine. It also offers a powerful tool for establishing in vitro models of disease, in particular, for neuropsychiatric disorders where live human neurons are essentially impossible to procure. Using iPSCs derived from three schizophrenia (SZ) patients, one of whom has 22q11.2del (velocardiofacial syndrome; VCFS), the authors developed a culture system to study SZ on a molecular and cellular level. SZ iPSCs were differentiated into functional, primarily glutamatergic neurons that were able to fire action potentials after â¼8 weeks in culture. Early differentiating neurons expressed a number of transcription factors/chromatin remodeling proteins and synaptic proteins relevant to SZ pathogenesis, including ZNF804A, RELN, CNTNAP2, CTNNA2, SMARCA2, and NRXN1. Although a small number of lines were developed in this preliminary study, the SZ line containing 22q11.2del showed a significant delay in the reduction of endogenous OCT4 and NANOG expression that normally occurs during differentiation. Constitutive expression of OCT4 has been observed in Dgcr8-deficient mouse embryonic stem cells (mESCs); DGCR8 maps to the 22q11.2-deleted region. These findings demonstrate that the method of inducing neural differentiation employed is useful for disease modeling in SZ and that the transition of iPSCs with 22q11.2 deletions towards a differentiated state may be marked by subtle changes in expression of pluripotency-associated genes.
Subject(s)
Gene Expression Regulation/physiology , Induced Pluripotent Stem Cells/physiology , Neurons/physiology , Schizophrenia/pathology , Action Potentials/drug effects , Action Potentials/genetics , Adult , Cell Differentiation/drug effects , Cell Differentiation/physiology , Cells, Cultured , Chromosome Aberrations , Chromosome Deletion , Chromosomes, Human, 21-22 and Y , Computational Biology , Embryonic Stem Cells/cytology , Embryonic Stem Cells/drug effects , Female , Gene Expression Regulation/drug effects , Humans , Induced Pluripotent Stem Cells/drug effects , Male , Microarray Analysis , Middle Aged , Nerve Growth Factors/pharmacology , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/metabolism , Patch-Clamp Techniques , RNA, Messenger/metabolism , Reelin Protein , Transcription Factors/genetics , Transcription Factors/metabolism , Wnt3A Protein/pharmacology , Young AdultABSTRACT
Several addiction susceptibility genes have been mapped by linkage and genomewide association. However, functional alleles associated with disease risk have not been identified, with a few possible exceptions. In addition, little is known about the cis- and trans-acting factors involved in regulating their expression. To address these issues, we used a ChIP-chip approach to identify regulatory elements in fetal-brain- targeting genes implicated in addiction and other neuropsychiatric conditions. Our data point to a number of putative regulatory elements, several of which, we show, are functionally significant. Many established or putative regulatory elements map near-disease-associated SNPs. These regions would be of interest to survey for patient-specific functional variants involved in disease susceptibility.
Subject(s)
Cell Adhesion Molecules, Neuronal/genetics , Gene Expression Profiling/methods , Nerve Tissue Proteins/genetics , Neurocognitive Disorders/genetics , Oligonucleotide Array Sequence Analysis/methods , Substance-Related Disorders/genetics , Brain/metabolism , Brain Chemistry/genetics , Calcium-Binding Proteins , Cell Line, Tumor , DNA Mutational Analysis/methods , Fetus/metabolism , Gene Expression Regulation/genetics , Genetic Markers/genetics , Genetic Testing/methods , Humans , Neural Cell Adhesion Molecules , Polymorphism, Single Nucleotide/genetics , Protein Structure, Tertiary/geneticsABSTRACT
Therapeutic concentrations of lithium salts inhibit glycogen synthase kinase 3 beta (GSK3ß) and phosphoinositide (PI) signaling suggesting that abnormal activation of these pathways could be a factor in the pathophysiology of bipolar disorder (BD). Involvement of these pathways is also supported by recent genome-wide association studies (GWASs). One way investigators have investigated the molecular basis of BD and the therapeutic action of lithium is by microarray expression studies, since both GSK3ß- and PI-mediated signal transduction pathways are coupled to transcriptional activation and inhibition. However, expression profiling has some limitations and investigators cannot use the approach to analyze fetal brain tissue, arguably the most relevant biological structure related to the development of genetically based psychiatric disorders. To address these shortcomings, the authors have taken a novel approach using chromatin immunoprecipitation-enriched material annealed to microarrays (ChIP-chip) targeting genes in fetal brain tissue bound by ß-catenin, a transcription factor that is directly regulated by GSK3ß. The promoters for 640 genes were found to be bound by ß-catenin, many of which are known schizophrenia (SZ), autism spectrum disorder (ASD), and BD candidates, including CACNA1B, NRNG, SNAP29, FGFR1, PCDH9, and nine others identified in recently published GWASs and genome-wide searches for copy number variants (CNVs). The findings suggest that seemingly disparate candidate genes for SZ and BD can be incorporated into a common molecular network revolving around GSK3ß/ß-catenin signaling. In addition, the finding that a putative lithium-responsive pathway may influence a subgroup of SZ and ASD candidate genes could have therapeutic implications.
Subject(s)
Bipolar Disorder/genetics , Chromatin Immunoprecipitation/methods , Gene Regulatory Networks , Promoter Regions, Genetic , Schizophrenia/genetics , beta Catenin/genetics , Bipolar Disorder/drug therapy , Bipolar Disorder/metabolism , Brain , Cadherins/genetics , Calcium Channels, N-Type/genetics , Child , Child Development Disorders, Pervasive/genetics , Female , Fetus , Genome-Wide Association Study , Glycogen Synthase Kinase 3/antagonists & inhibitors , Glycogen Synthase Kinase 3/metabolism , Glycogen Synthase Kinase 3 beta , Humans , Inositol Phosphates/antagonists & inhibitors , Inositol Phosphates/metabolism , Lithium/metabolism , Lithium/pharmacology , Oligonucleotide Array Sequence Analysis , Protocadherins , Qb-SNARE Proteins/genetics , Qc-SNARE Proteins/genetics , Receptor, Fibroblast Growth Factor, Type 1/genetics , Schizophrenia/drug therapy , Schizophrenia/mortality , Signal Transduction/genetics , Transcriptional Activation , beta Catenin/metabolismABSTRACT
BACKGROUND: Lowe syndrome (LS) is caused by loss-of-function mutations in the X-linked gene OCRL, which codes for an inositol polyphosphate 5-phosphatase that plays a key role in endosome recycling, clathrin-coated pit formation, and actin polymerization. It is characterized by congenital cataracts, intellectual and developmental disability, and renal proximal tubular dysfunction. Patients are also at high risk for developing glaucoma and seizures. We recently developed induced pluripotent stem cell (iPSC) lines from three patients with LS who have hypomorphic variants affecting the 3' end of the gene, and their neurotypical brothers to serve as controls. METHODS: In this study, we used RNA sequencing (RNA-seq) to obtain transcriptome profiles in LS and control neural progenitor cells (NPCs). RESULTS: In a comparison of the patient and control NPCs (n = 3), we found 16 differentially expressed genes (DEGs) at the multiple test adjusted p value (padj) < 0.1, with nine at padj < 0.05. Using nominal p value < 0.05, 319 DEGs were detected. The relatively small number of DEGs could be due to the fact that OCRL is not a transcription factor per se, although it could have secondary effects on gene expression through several different mechanisms. Although the number of DEGs passing multiple test correction was small, those that were found are quite consistent with some of the known molecular effects of OCRL protein, and the clinical manifestations of LS. Furthermore, using gene set enrichment analysis (GSEA), we found that genes increased expression in the patient NPCs showed enrichments of several gene ontology (GO) terms (false discovery rate < 0.25): telencephalon development, pallium development, NPC proliferation, and cortex development, which are consistent with a condition characterized by intellectual disabilities and psychiatric manifestations. In addition, a significant enrichment among the nominal DEGs for genes implicated in autism spectrum disorder (ASD) was found (e.g., AFF2, DNER, DPP6, DPP10, RELN, CACNA1C), as well as several that are strong candidate genes for the development of eye problems found in LS, including glaucoma. The most notable example is EFEMP1, a well-known candidate gene for glaucoma and other eye pathologies. CONCLUSION: Overall, the RNA-seq findings present several candidate genes that could help explain the underlying basis for the neurodevelopmental and eye problems seen in boys with LS.
Subject(s)
Eye Diseases/genetics , Gene Expression Profiling , Induced Pluripotent Stem Cells/metabolism , Neural Stem Cells/metabolism , Oculocerebrorenal Syndrome/genetics , Adolescent , Adult , Cataract/genetics , Cells, Cultured , Child , Endosomes/metabolism , Extracellular Matrix Proteins/genetics , Glaucoma/genetics , Humans , Male , Mutation , Oculocerebrorenal Syndrome/metabolism , Oculocerebrorenal Syndrome/physiopathology , Phosphoric Monoester Hydrolases/genetics , Phosphoric Monoester Hydrolases/metabolism , Reelin Protein , Sequence Analysis, RNA , Young AdultABSTRACT
It has been difficult to identify disease-causing alleles in schizophrenia (SZ) and bipolar disorder (BD) candidate genes. One reason is that responsible functional variants may exist in unidentified regulatory domains. With the advent of microarray technology and high throughput sequencing, however, it is now feasible to screen genes for such regulatory domains relatively easily by using chromatin immunoprecipitation-based methodologies, such as ChIP-chip and ChIP-seq. In ChIP-chip, regulatory sequences can be captured from chromatin immunoprecipitates prepared with antibodies against covalently modified histones that mark certain regulatory domains; DNA extracted from such immunoprecipitates can then be used as microarray probes. As a first step toward demonstrating the feasibility of this approach in psychiatric genetics, we used ChIP-chip to identify regulatory domains in several candidate genes: NRG1, DTNBP1, DISC1, DAO, DAOA, PDE4B, and COMT. Immunoprecipitates were generated with antibodies to histone H3 acetylated at lysine 9 (H3K9Ac) and histone H3 monomethylated at lysine 4 (H3K4me1), which mark promoters and some enhancers, using fetal brain chromatin as a substrate. Several novel putative regulatory elements, as well as the core and proximal promoters for each gene, were enriched in the immunoprecipitates. Genetic variants within these regions would be of interest to study as potential disease-associated alleles.
Subject(s)
Bipolar Disorder/genetics , Chromatin Immunoprecipitation/methods , Genetic Predisposition to Disease , Microarray Analysis/methods , Schizophrenia/genetics , Brain/metabolism , Brain/pathology , Carrier Proteins/genetics , Catechol O-Methyltransferase/genetics , Cyclic Nucleotide Phosphodiesterases, Type 4/genetics , D-Amino-Acid Oxidase/genetics , Dysbindin , Dystrophin-Associated Proteins , Fetus , Histones/genetics , Humans , Intracellular Signaling Peptides and Proteins , Lysine/genetics , Nerve Tissue Proteins/genetics , Neuregulin-1/geneticsABSTRACT
BACKGROUND/AIMS: Neuregulin 1 (NRG1) is a positional candidate gene in schizophrenia (SZ). Two major susceptibility loci in the NRG1 gene approximately one million nucleotides apart have been identified in genetic studies. Several candidate functional allelic variants have been described that might be involved in disease susceptibility. However, the findings are still preliminary. We recently mapped active promoters and other regulatory domains in several SZ and bipolar disorder (BD) candidate genes using ChIP-chip (chromatin immunoprecipitation hybridized to microarrays). One was the promoter for the NRG1 isoform, SMDF, which maps to the 3' end of the gene complex. Analysis of the SNP database revealed several polymorphisms within the approximate borders of the region immunoprecipitated in our ChIP-chip experiments, one of which is rs7825588. METHODS: This SNP was analyzed in patients with SZ and BD and its effect on promoter function was assessed by electromobility gel shift assays and luciferase reporter constructs. RESULTS: A significant increase in homozygosity for the minor allele was found in patients with SZ (genotype distribution chi(2) = 7.32, p = 0.03) but not in BD (genotype distribution chi(2) = 0.52, p = 0.77). Molecular studies demonstrated modest, but statistically significant allele-specific differences in protein binding and promoter function. CONCLUSION: The findings suggest that homozygosity for rs725588 could be a risk genotype for SZ.
Subject(s)
Nerve Tissue Proteins/genetics , Polymorphism, Single Nucleotide , Promoter Regions, Genetic , Schizophrenia/genetics , Adult , Cell Line, Tumor , Electrophoretic Mobility Shift Assay , Female , Gene Frequency , Genetic Predisposition to Disease , Genotype , Humans , Male , Middle Aged , Nerve Tissue Proteins/metabolism , Neuregulin-1 , Protein Isoforms/genetics , Sequence Analysis, DNA , TransfectionABSTRACT
Cannabis has been reported as a likely risk factor for the development of psychosis, and a gene × environment interaction with the catechol-O-methyltransferase (COMT) gene has been proposed. Moreover, COMT has been separately linked to affective symptoms in psychosis. Despite a high rate of cannabis abuse and affective symptoms in African Americans, no studies exploring a relationship between COMT and psychosis in this group have been reported. An existing database of psychotic patients with and without adolescent cannabis use/affective symptoms was examined, and chi-square analyses for independence were applied separately for both Caucasians and African-Americans to examine genotype associations with adolescent cannabis use and affective symptoms (past or present). The two subject groups did not differ with respect to the prevalence of adolescent cannabis abuse or presence of affective symptoms. Further study is needed, with non-psychotic controls and larger samples.
Subject(s)
Catechol O-Methyltransferase/genetics , Cross-Cultural Comparison , Marijuana Abuse/genetics , Mental Disorders/genetics , Polymorphism, Genetic/genetics , Adolescent , Black or African American , Chi-Square Distribution , Databases, Bibliographic/statistics & numerical data , Female , Gene Frequency , Genotype , Humans , Male , Marijuana Abuse/epidemiology , Mental Disorders/epidemiology , White PeopleABSTRACT
Schizophrenia (SZ) is a highly heterogeneous disorder in both its symptoms and risk factors. One of the most prevalent genetic risk factors for SZ is the hemizygous microdeletion at chromosome 22q11.2 (22q11DS) that confers a 25-fold increased risk. Six of the genes directly disrupted in 22qDS encode for mitochondrial-localizing proteins. Here, we test the hypothesis that stem cell-derived neurons from subjects with the 22q11DS and SZ have mitochondrial deficits relative to typically developing controls. Human iPSCs from four lines of affected subjects and five lines of controls were differentiated into forebrain-like excitatory neurons. In the patient group, we find significant reductions of ATP levels that appear to be secondary to reduced activity in oxidative phosphorylation complexes I and IV. Protein products of mitochondrial-encoded genes are also reduced. As one of the genes deleted in the 22q11.2 region is MRPL40, a component of the mitochondrial ribosome, we generated a heterozygous mutation of MRPL40 in a healthy control iPSC line. Relative to its isogenic control, this line shows similar deficits in mitochondrial DNA-encoded proteins, ATP level, and complex I and IV activity. These results suggest that in the 22q11DS MRPL40 heterozygosity leads to reduced mitochondria ATP production secondary to altered mitochondrial protein levels. Such defects could have profound effects on neuronal function in vivo.