Publications

Abstract

Retinoic acid-related orphan receptor alpha gene (RORa) and the microRNA MIR137 have both recently been identified as novel candidate genes for neuropsychiatric disorders. RORa encodes a ligand-dependent orphan nuclear receptor that acts as a transcriptional regulator and miR-137 is a brain enriched small non-coding RNA that interacts with gene transcripts to control protein levels. Given the mounting evidence for RORa in autism spectrum disorders (ASD) and MIR137 in schizophrenia and ASD, we investigated if there was a functional biological relationship between these two genes. Herein, we demonstrate that miR-137 targets the 3'UTR of RORa in a site specific manner. We also provide further support for MIR137 as an autism candidate by showing that a large number of previously implicated autism genes are also putatively targeted by miR-137. This work supports the role of MIR137 as an ASD candidate and demonstrates a direct biological link between these previously unrelated autism candidate genes

Abstract

FOXP2 was the first gene shown to cause a Mendelian form of speech and language disorder. Although developmentally expressed in many organs, loss of a single copy of FOXP2 leads to a phenotype that is largely restricted to orofacial impairment during articulation and linguistic processing deficits. Why perturbed FOXP2 function affects specific aspects of the developing brain remains elusive. We investigated the role of FOXP2 in neuronal differentiation and found that FOXP2 drives molecular changes consistent with neuronal differentiation in a human model system. We identified a network of FOXP2 regulated genes related to retinoic acid signaling and neuronal differentiation. FOXP2 also produced phenotypic changes associated with neuronal differentiation including increased neurite outgrowth and reduced migration. Crucially, cells expressing FOXP2 displayed increased sensitivity to retinoic acid exposure. This suggests a mechanism by which FOXP2 may be able to increase the cellular differentiation response to environmental retinoic acid cues for specific subsets of neurons in the brain. These data demonstrate that FOXP2 promotes neuronal differentiation by interacting with the retinoic acid signaling pathway and regulates key processes required for normal circuit formation such as neuronal migration and neurite outgrowth. In this way, FOXP2, which is found only in specific subpopulations of neurons in the brain, may drive precise neuronal differentiation patterns and/or control localization and connectivity of these FOXP2 positive cells

Abstract

The genetic basis of complex neurological disorders involving language are poorly understood, partly due to the multiple additive genetic risk factors that are thought to be responsible. Furthermore, these conditions are often syndromic in that they have a range of endophenotypes that may be associated with the disorder and that may be present in different combinations in patients. However, the emergence of individual genes implicated across multiple disorders has suggested that they might share similar underlying genetic mechanisms. The CNTNAP2 gene is an excellent example of this, as it has recently been implicated in a broad range of phenotypes including autism spectrum disorder (ASD), schizophrenia, intellectual disability, dyslexia and language impairment. This review considers the evidence implicating CNTNAP2 in these conditions, the genetic risk factors and mutations that have been identified in patient and population studies and how these relate to patient phenotypes. The role of CNTNAP2 is examined in the context of larger neurogenetic networks during development and disorder, given what is known regarding the regulation and function of this gene. Understanding the role of CNTNAP2 in diverse neurological disorders will further our understanding of how combinations of individual genetic risk factors can contribute to complex conditions

Abstract

The fruitless gene (fru) encodes a set of transcription factors (Fru) that display sexually dimorphic gene expression in the brain of the fruit-fly;Drosophila melanogaster . Behavioural studies have demonstrated that fru isessentialforcourtshipbehaviour inthemale flyandisthoughttoact bydirectingthe development of sex-specific neural circuitry that encodes this innate behavioural response. This study reports the identification of direct regulatory targets of the sexually dimorphic isoforms of the Fru protein using an in vitro model system. Genome wide binding sites were identified for each of the isoforms using Chromatin Immunoprecipitation coupled to deep sequencing (ChIP-Seq). Putative target genes were found to be involved in processes such as neurotransmission, ion-channel signalling and neuron development. All isoforms showed asignificant bias towards genes located on the X-chromosome,which may reflect a specific role for Fru in regulating x-linked genes. Taken together with expression analysis carried out in Fru positive neurons specifically isolated from the male fly brain, it appears that the Fru protein acts as a transcriptional activator. Understanding the regulatory cascades induced by Fru will help to shed light on the molecular mechanisms that are important for specification of neural circuitry underlying complex behaviour

Abstract

It is a challenge to identify the molecular networks contributing to the neural basis of human speech. Mutations in transcription factor FOXP2 cause difficulties mastering fluent speech (developmental verbal dyspraxia, DVD), while mutations of sushi-repeat protein SRPX2 lead to epilepsy of the rolandic (sylvian) speech areas, with DVD or with bilateral perisylvian polymicrogyria. Pathophysiological mechanisms driven by SRPX2 involve modified interaction with the plasminogen activator receptor (uPAR). Independent chromatin-immunoprecipitation microarray screening has identified the uPAR gene promoter as a potential target site bound by FOXP2. Here, we directly tested for the existence of a transcriptional regulatory network between human FOXP2 and the SRPX2/uPAR complex. In silico searches followed by gel retardation assays identified specific efficient FOXP2 binding sites in each of the promoter regions of SRPX2 and uPAR. In FOXP2-transfected cells, significant decreases were observed in the amounts of both SRPX2 (43.6%) and uPAR (38.6%) native transcripts. Luciferase reporter assays demonstrated that FOXP2 expression yielded marked inhibition of SRPX2 (80.2%) and uPAR (77.5%) promoter activity. A mutant FOXP2 that causes DVD (p.R553H) failed to bind to SRPX2 and uPAR target sites, and showed impaired down-regulation of SRPX2 and uPAR promoter activity. In a patient with polymicrogyria of the left rolandic operculum, a novel FOXP2 mutation (p.M406T) was found in the leucine-zipper (dimerization) domain. p.M406T partially impaired FOXP2 regulation of SRPX2 promoter activity, while that of the uPAR promoter remained unchanged. Together with recently described FOXP2-CNTNPA2 and SRPX2/uPAR links, the FOXP2-SRPX2/uPAR network provides exciting insights into molecular pathways underlying speech-related disorders.

Abstract

Mutations of the epidermal growth factor receptor (EGFR) gene are found at a relatively high frequency in glioma, with the most common being the de2-7 EGFR (or EGFRvIII). This mutation arises from an in-frame deletion of exons 2–7, which removes 267 amino acids from the extracellular domain of the receptor. Despite being unable to bind ligand, the de2-7 EGFR is constitutively active at a low level. Transfection of human glioma cells with the de2-7 EGFR has little effect in vitro, but when grown as tumor xenografts this mutated receptor imparts a dramatic growth advantage. We have now mapped the phosphorylation pattern of de2-7 EGFR, both in vivo and in vitro, using a panel of antibodies unique to the different phosphorylated tyrosine residues. Phosphorylation of de2-7 EGFR was detected constitutively at all tyrosine sites surveyed both in vitro and in vivo, including tyrosine 845, a known target in the wild-type EGFR for src kinase. There was a substantial upregulation of phosphorylation at every tyrosine residue of the de2-7 EGFR when cells were grown in vivo compared to the receptor isolated from cells cultured in vitro. Upregulation of phosphorylation could be mimicked in vitro by the addition of specifi c components of the ECM such as collagen via an integrin-dependent mechanism. Since this increase in in vivo phosphorylation enhances de2-7 EGFR signaling, this observation explains why the growth enhancement mediated by de2-7 EGFR is largely restricted to the in vivo environment. In a second set of experiments we analyzed the interaction between EGFRvIII and ErbB2. Co-expression of these proteins in NR6 cells, a mouse fi broblast line devoid of ErbB family members, dramatically enhanced in vivo tumorigenicity of these cells compared to cells expressing either protein alone. Detailed analysis of these xenografts demonstrated that EGFRvIII could heterodimerize and transphosphorylate the ErbB2. Since both EGFRvIII and ErbB2 are commonly expressed at gliomas, this data suggests that the co-expression of these two proteins may enhance glioma tumorigenicity.

Abstract

FOXP2, the first gene to have been implicated in a developmental communication disorder, offers a unique entry point into neuromolecular mechanisms influencing human speech and language acquisition. In multiple members of the well-studied KE family, a heterozygous missense mutation in FOXP2 causes problems in sequencing muscle movements required for articulating speech (developmental verbal dyspraxia), accompanied by wider deficits in linguistic and grammatical processing. Chromosomal rearrangements involving this locus have also been identified. Analyses of FOXP2 coding sequence in typical forms of specific language impairment (SLI), autism, and dyslexia have not uncovered any etiological variants. However, no previous study has performed mutation screening of children with a primary diagnosis of verbal dyspraxia, the most overt feature of the disorder in affected members of the KE family. Here, we report investigations of the entire coding region of FOXP2, including alternatively spliced exons, in 49 probands affected with verbal dyspraxia. We detected variants that alter FOXP2 protein sequence in three probands. One such variant is a heterozygous nonsense mutation that yields a dramatically truncated protein product and cosegregates with speech and language difficulties in the proband, his affected sibling, and their mother. Our discovery of the first nonsense mutation in FOXP2 now opens the door for detailed investigations of neurodevelopment in people carrying different etiological variants of the gene. This endeavor will be crucial for gaining insight into the role of FOXP2 in human cognition.