MicroRNA Profiling of Neurons Generated Using Induced Pluripotent Stem Cells Derived from Patients with Schizophrenia and Schizoaffective Disorder, and 22q11.2 Del
TL;DR: Differentially expressed miRNAs previously identified using autopsy samples and peripheral cells, both of which have significant methodological problems, are indeed disrupted in neuropsychiatric disorders and likely have an underlying genetic basis.
Abstract: We are using induced pluripotent stem cell (iPSC) technology to study neuropsychiatric disorders associated with 22q112 microdeletions (del), the most common known schizophrenia (SZ)-associated genetic factor Several genes in the region have been implicated; a promising candidate is DGCR8, which codes for a protein involved in microRNA (miRNA) biogenesis We carried out miRNA expression profiling (miRNA-seq) on neurons generated from iPSCs derived from controls and SZ patients with 22q112 del Using thresholds of p<001 for nominal significance and 15-fold differences in expression, 45 differentially expressed miRNAs were detected (13 lower in SZ and 32 higher) Of these, 6 were significantly down-regulated in patients after correcting for genome wide significance (FDR<005), including 4 miRNAs that map to the 22q112 del region In addition, a nominally significant increase in the expression of several miRNAs was found in the 22q112 neurons that were previously found to be differentially expressed in autopsy samples and peripheral blood in SZ and autism spectrum disorders (eg, miR-34, miR-4449, miR-146b-3p, and miR-23a-5p) Pathway and function analysis of predicted mRNA targets of the differentially expressed miRNAs showed enrichment for genes involved in neurological disease and psychological disorders for both up and down regulated miRNAs Our findings suggest that: i neurons with 22q112 del recapitulate the miRNA expression patterns expected of 22q112 haploinsufficiency, ii differentially expressed miRNAs previously identified using autopsy samples and peripheral cells, both of which have significant methodological problems, are indeed disrupted in neuropsychiatric disorders and likely have an underlying genetic basis
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19 Nov 2015
TL;DR: The first description in the English language of the constellation of findings now known to be due to this chromosomal difference was made in the 1960s in children with DiGeorge syndrome, who presented with the clinical triad of immunodeficiency, hypoparathyroidism and congenital heart disease as mentioned in this paper.
Abstract: 22q11.2 deletion syndrome (22q11.2DS) is the most common chromosomal microdeletion disorder, estimated to result mainly from de novo non-homologous meiotic recombination events occurring in approximately 1 in every 1,000 fetuses. The first description in the English language of the constellation of findings now known to be due to this chromosomal difference was made in the 1960s in children with DiGeorge syndrome, who presented with the clinical triad of immunodeficiency, hypoparathyroidism and congenital heart disease. The syndrome is now known to have a heterogeneous presentation that includes multiple additional congenital anomalies and later-onset conditions, such as palatal, gastrointestinal and renal abnormalities, autoimmune disease, variable cognitive delays, behavioural phenotypes and psychiatric illness - all far extending the original description of DiGeorge syndrome. Management requires a multidisciplinary approach involving paediatrics, general medicine, surgery, psychiatry, psychology, interventional therapies (physical, occupational, speech, language and behavioural) and genetic counselling. Although common, lack of recognition of the condition and/or lack of familiarity with genetic testing methods, together with the wide variability of clinical presentation, delays diagnosis. Early diagnosis, preferably prenatally or neonatally, could improve outcomes, thus stressing the importance of universal screening. Equally important, 22q11.2DS has become a model for understanding rare and frequent congenital anomalies, medical conditions, psychiatric and developmental disorders, and may provide a platform to better understand these disorders while affording opportunities for translational strategies across the lifespan for both patients with 22q11.2DS and those with these associated features in the general population.
1,850 citations
Journal Article•
TL;DR: 22q11.2 deletion syndrome is now known to have a heterogeneous presentation that includes multiple additional congenital anomalies and later-onset conditions, such as palatal, gastrointestinal and renal abnormalities, autoimmune disease, variable cognitive delays, behavioural phenotypes and psychiatric illness — all far extending the original description of DiGeorge syndrome.
983 citations
01 Jan 2009
TL;DR: In this article, a review outlines the current understanding of miRNA target recognition in animals and discusses the widespread impact of miRNAs on both the expression and evolution of protein-coding genes.
Abstract: MicroRNAs (miRNAs) are endogenous ∼23 nt RNAs that play important gene-regulatory roles in animals and plants by pairing to the mRNAs of protein-coding genes to direct their posttranscriptional repression. This review outlines the current understanding of miRNA target recognition in animals and discusses the widespread impact of miRNAs on both the expression and evolution of protein-coding genes.
646 citations
TL;DR: The findings show that distinct ASD, SZ, and BD candidate genes converge on common molecular targets—an important consideration for developing novel therapeutics in genetically heterogeneous complex traits.
Abstract: CHD8 (chromodomain helicase DNA-binding protein 8), which codes for a member of the CHD family of ATP-dependent chromatin-remodeling factors, is one of the most commonly mutated genes in autism spectrum disorders (ASD) identified in exome-sequencing studies. Loss of function mutations in the gene have also been found in schizophrenia (SZ) and intellectual disabilities and influence cancer cell proliferation. We previously reported an RNA-seq analysis carried out on neural progenitor cells (NPCs) and monolayer neurons derived from induced pluripotent stem (iPS) cells that were heterozygous for CHD8 knockout (KO) alleles generated using CRISPR-Cas9 gene editing. A significant number of ASD and SZ candidate genes were among those that were differentially expressed in a comparison of heterozygous KO lines (CHD8
+/−) vs isogenic controls (CHD8
+/−), including the SZ and bipolar disorder (BD) candidate gene TCF4, which was markedly upregulated in CHD8
+/− neuronal cells. In the current study, RNA-seq was carried out on CHD8
+/− and isogenic control (CHD8
+/+) cerebral organoids, which are 3-dimensional structures derived from iPS cells that model the developing human telencephalon. TCF4 expression was, again, significantly upregulated. Pathway analysis carried out on differentially expressed genes (DEGs) revealed an enrichment of genes involved in neurogenesis, neuronal differentiation, forebrain development, Wnt/β-catenin signaling, and axonal guidance, similar to our previous study on NPCs and monolayer neurons. There was also significant overlap in our CHD8
+/− DEGs with those found in a transcriptome analysis carried out by another group using cerebral organoids derived from a family with idiopathic ASD. Remarkably, the top DEG in our respective studies was the non-coding RNA DLX6-AS1, which was markedly upregulated in both studies; DLX6-AS1 regulates the expression of members of the DLX (distal-less homeobox) gene family. DLX1 was also upregulated in both studies. DLX genes code for transcription factors that play a key role in GABAergic interneuron differentiation. Significant overlap was also found in a transcriptome study carried out by another group using iPS cell-derived neurons from patients with BD, a condition characterized by dysregulated WNT/β-catenin signaling in a subgroup of affected individuals. Overall, the findings show that distinct ASD, SZ, and BD candidate genes converge on common molecular targets—an important consideration for developing novel therapeutics in genetically heterogeneous complex traits.
198 citations
Cites background from "MicroRNA Profiling of Neurons Gener..."
...2 del diagnosed with SZ or schizoaffective disorder [44]....
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TL;DR: Transcriptome profiling revealed that CHD8 hemizygosity (CHD8+/−) affected the expression of several thousands of genes in neural progenitors and early differentiating neurons, and seven of the twelve genes associated with human brain volume or head size by genome-wide association studies were dysregulated in CHD 8+/+ neural progensitors or neurons.
Abstract: Disruptive mutation in the CHD8 gene is one of the top genetic risk factors in autism spectrum disorders (ASDs). Previous analyses of genome-wide CHD8 occupancy and reduced expression of CHD8 by shRNA knockdown in committed neural cells showed that CHD8 regulates multiple cell processes critical for neural functions, and its targets are enriched with ASD-associated genes. To further understand the molecular links between CHD8 functions and ASD, we have applied the CRISPR/Cas9 technology to knockout one copy of CHD8 in induced pluripotent stem cells (iPSCs) to better mimic the loss-of-function status that would exist in the developing human embryo prior to neuronal differentiation. We then carried out transcriptomic and bioinformatic analyses of neural progenitors and neurons derived from the CHD8 mutant iPSCs. Transcriptome profiling revealed that CHD8 hemizygosity (CHD8
+/−) affected the expression of several thousands of genes in neural progenitors and early differentiating neurons. The differentially expressed genes were enriched for functions of neural development, β-catenin/Wnt signaling, extracellular matrix, and skeletal system development. They also exhibited significant overlap with genes previously associated with autism and schizophrenia, as well as the downstream transcriptional targets of multiple genes implicated in autism. Providing important insight into how CHD8 mutations might give rise to macrocephaly, we found that seven of the twelve genes associated with human brain volume or head size by genome-wide association studies (e.g., HGMA2) were dysregulated in CHD8
+/− neural progenitors or neurons. We have established a renewable source of CHD8
+/− iPSC lines that would be valuable for investigating the molecular and cellular functions of CHD8. Transcriptomic profiling showed that CHD8 regulates multiple genes implicated in ASD pathogenesis and genes associated with brain volume.
126 citations
Cites background from "MicroRNA Profiling of Neurons Gener..."
...2 deletion and diagnosis of a psychotic disorder [31]....
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References
More filters
TL;DR: This work presents DESeq2, a method for differential analysis of count data, using shrinkage estimation for dispersions and fold changes to improve stability and interpretability of estimates, which enables a more quantitative analysis focused on the strength rather than the mere presence of differential expression.
Abstract: In comparative high-throughput sequencing assays, a fundamental task is the analysis of count data, such as read counts per gene in RNA-seq, for evidence of systematic changes across experimental conditions. Small replicate numbers, discreteness, large dynamic range and the presence of outliers require a suitable statistical approach. We present DESeq2, a method for differential analysis of count data, using shrinkage estimation for dispersions and fold changes to improve stability and interpretability of estimates. This enables a more quantitative analysis focused on the strength rather than the mere presence of differential expression. The DESeq2 package is available at http://www.bioconductor.org/packages/release/bioc/html/DESeq2.html
.
47,038 citations
"MicroRNA Profiling of Neurons Gener..." refers methods in this paper
...To identify differentially expressed (DE) miRNAs, we applied DESeq2 to analyze the read counts of all microRNAs [77]....
[...]
TL;DR: By following this protocol, investigators are able to gain an in-depth understanding of the biological themes in lists of genes that are enriched in genome-scale studies.
Abstract: DAVID bioinformatics resources consists of an integrated biological knowledgebase and analytic tools aimed at systematically extracting biological meaning from large gene/protein lists. This protocol explains how to use DAVID, a high-throughput and integrated data-mining environment, to analyze gene lists derived from high-throughput genomic experiments. The procedure first requires uploading a gene list containing any number of common gene identifiers followed by analysis using one or more text and pathway-mining tools such as gene functional classification, functional annotation chart or clustering and functional annotation table. By following this protocol, investigators are able to gain an in-depth understanding of the biological themes in lists of genes that are enriched in genome-scale studies.
31,015 citations
"MicroRNA Profiling of Neurons Gener..." refers methods in this paper
...Then, IPA Core Analysis and the software DAVID were used for function analysis of the target genes [84,85]....
[...]
TL;DR: Induction of pluripotent stem cells from mouse embryonic or adult fibroblasts by introducing four factors, Oct3/4, Sox2, c-Myc, and Klf4, under ES cell culture conditions is demonstrated and iPS cells, designated iPS, exhibit the morphology and growth properties of ES cells and express ES cell marker genes.
23,959 citations
TL;DR: Bowtie extends previous Burrows-Wheeler techniques with a novel quality-aware backtracking algorithm that permits mismatches and can be used simultaneously to achieve even greater alignment speeds.
Abstract: Bowtie is an ultrafast, memory-efficient alignment program for aligning short DNA sequence reads to large genomes. For the human genome, Burrows-Wheeler indexing allows Bowtie to align more than 25 million reads per CPU hour with a memory footprint of approximately 1.3 gigabytes. Bowtie extends previous Burrows-Wheeler techniques with a novel quality-aware backtracking algorithm that permits mismatches. Multiple processor cores can be used simultaneously to achieve even greater alignment speeds. Bowtie is open source http://bowtie.cbcb.umd.edu.
20,335 citations
"MicroRNA Profiling of Neurons Gener..." refers methods in this paper
...org/), allowing for one mismatch at most using bowtie ([75]....
[...]
TL;DR: It is demonstrated that iPS cells can be generated from adult human fibroblasts with the same four factors: Oct3/4, Sox2, Klf4, and c-Myc.
18,175 citations