Dlx Transcription Factors Research Articles

Dissecting the impact of regional identity and the oncogenic role of human-specific NOTCH2NL in an hESC model of H3.3G34R-mutant glioma

H3.3G34R-mutant gliomas are lethal tumors of the cerebral hemispheres with unknown mechanisms of regional specificity and tumorigenicity. We developed a human embryonic stem cell (hESC)-based model of H3.3G34R-mutant glioma that recapitulates the key features of the tumors with cell-type specificity to forebrain interneuronal progenitors but not hindbrain precursors. We show that H3.3G34R, ATRX, and TP53 mutations cooperatively impact alternative RNA splicing events, particularly suppression of intron retention. This leads to increased expression of components of the Notch pathway, notably NOTCH2NL, a human-specific gene family. We also uncover a parallel mechanism of enhanced NOTCH2NL expression via genomic amplification of its locus in some H3.3G34R-mutant tumors. These findings demonstrate a novel mechanism whereby evolutionary pathways that lead to larger brain size in humans are co-opted to drive tumor growth.

Genomic Resolution of DLX-Orchestrated Transcriptional Circuits Driving Development of Forebrain GABAergic Neurons

DLX transcription factors (TFs) are master regulators of the developing vertebrate brain, driving forebrain GABAergic neuronal differentiation. Ablation of Dlx1&2 alters expression of genes that are critical for forebrain GABAergic development. We integrated epigenomic and transcriptomic analyses, complemented with in situ hybridization (ISH), and in vivo and in vitro studies of regulatory element (RE) function. This elucidated the DLX-organized gene regulatory network at genomic, cellular, and spatial level in mouse embryonic basal ganglia. DLX TFs perform dual activating and repressing functions; the consequences of their binding were determined by the sequence and genomic context of target loci. Our results reveal and, in part, explain the paradox of widespread DLX binding contrasted with a limited subset of target loci that are sensitive at the epigenomic and transcriptomic level to Dlx1&2 ablation. The regulatory properties identified here for DLX TFs suggest general mechanisms by which TFs orchestrate dynamic expression programs underlying neurodevelopment.

09 Could DLX2 regulation of neural progenitor cell fate contribute to differentiation of diffuse intrinsic pontine glioma (DIPG)?

Introduction: Diffuse intrinsic pontine glioma (DIPG) is refractory to therapy. The identification of histone H3.1/H3.3 K27M mutations in most DIPG has provided new insights. The DLX homeobox genes are expressed in the developing forebrain. The Dlx1/Dlx2 double knockout (DKO) mouse loses tangential GABAergic interneuron migration to the neocortex. We have identified genes that encode glutamic acid decarboxylase (GAD) enzymes as direct targets of DLX1/DLX2. In DIPG patients with H3.3 K27M mutations there is decreased Dlx2 and increased expression of the myelin transcription factor, Myt1. Methods and Results: We used bioinformatics approaches and chromatin immunoprecipitation (ChIP) assays to identify Olig2, Nkx2.2 and Myt1 promoter sequences as candidate DLX2 targets in vivo. DNA binding specificity was confirmed. The functional consequences of Dlx2 co-expression with reporter constructs of ChIP-isolated promoter fragments of Olig2 and Nkx2.2 demonstrated repression of gene targets in vitro. qPCR showed increased Olig2 and Nkx2.2 expression in the DKO forebrain. Stable transfection of a murine DIPG cell line with Dlx2 resulted in increased Gad1 and Gad2 and decreased Olig2 and Nkx2.2 expression. Of significance, we demonstrated decreased expression of H3.3 K27M and restoration of H3.3 K27 tri-methylation (me3). Conclusions: DLX transcription factors promote GABAergic interneuron and concomitant inhibition of oligodendroglial differentiation in neural progenitors by repression of a suite of genes including Olig2 and Nkx2.2. Restoration of H3 K27me3 expression in DIPG provides a promising lead towards exploration of differentiation as a therapeutic strategy for DIPG.

BI-03 * DLX HOMEOBOX GENES REGULATE NEURONAL VS OLIGODENDROGLIAL CELL FATE DECISIONS: POTENTIAL RELEVANCE TO PEDIATRIC GLIONEURONAL TUMORS AND HIGH GRADE GLIOMAS

BACKGROUND: Pediatric high grade gliomas (pHGG) are localized to different neuroanatomic compartments with significant differences in their gene expression profiles. In Histone H3.3 G34V/R mutants, DLX2 homeobox gene expression is increased with a corresponding decrease in myelin transcription factor (MYT1) expression. DLX genes are necessary for tangential migration and differentiation of inhibitory interneurons during CNS development. Neural progenitors derived from DLX1/DLX2 double knockout (DKO) mice transplanted into a wild-type forebrain differentiate into oligodendrocytes, providing support for the DLX homeobox genes in neuronal-glial cell fate decisions. METHODS: Chromatin immunoprecipitation (ChIP) assays using a DLX2 antibody were followed by qPCR. ChIP-reChIP assays were performed using antibodies that recognize specific histone modifications. Electrophoretic mobility shift assays (EMSA) were performed using recombinant DLX2 protein and oligonucleotide probes from ChIP-specified promoter regions. Reporter gene assays provided functional assessment of protein-DNA interactions in vitro. Target gene expression was assessed comparing wild-type (WT) and DLX1/DLX2 DKO tissues. RESULTS: Homeodomain binding sites were localized to the promoters of Olig2, Myt1, Nkx2.2 and others in silico. ChIP assays confirmed promoter occupancy by DLX2. ChIPseq experiments are underway. EMSA studies demonstrated specific DLX2-promoter complexes. Luciferase assays showed repression of Olig2 and Nkx2.2 reporter gene expression, consistent with co-occupancy of H3K27me3 in WT and increased target gene expression in DLX1/DLX2 DKO embryonic tissues. We also demonstrated DLX2 expression in a cohort of gangliogliomas. CONCLUSIONS: Our results support a role for DLX transcription factors in controlling neural progenitor specification by activating GABAergic and inhibiting oligodendroglial cell fates through transcriptional repression of a suite of genes required for oligodendrocyte differentiation. Understanding how pHGG and glioneuronal tumors co-opt these neurodevelopmental programs will lead to novel pharmacologic approaches that promote glioma differentiation.

MiR-124 negatively regulates osteogenic differentiation and in vivo bone formation of mesenchymal stem cells.

MicroRNAs are novel key regulators of cellular differentiation. Dlx transcription factors play an important role in osteoblast differentiation, and Dlx5 and Dlx2 are known targets of miR-124. Therefore, in the present study, we investigated the regulatory effects of miR-124 on the osteogenic differentiation and in vivo bone formation of mesenchymal stem cells (MSCs). During osteogenic induction by BMP2, the expression levels of miR-124 were inversely correlated with those of osteogenic differentiation marker genes in human and mouse bone marrow-derived MSCs, MC3T3-E1 cells and C2C12 cells. The overexpression of a miR-124 mimic significantly decreased the expression levels of Dlx5, Dlx3, and Dlx2, whereas the silencing of miR-124 with hairpin inhibitors significantly increased the expression of these Dlx genes. Luciferase reporter assays demonstrated that miR-124 directly targets the 3'UTRs of Dlx3, Dlx5, and Dlx2. The overexpression of a miR-124 mimic suppressed the osteogenic marker gene expression levels, alkaline phosphatase activity and matrix mineralization, which were all significantly increased by the overexpression of a miR-124 inhibitor. When ectopic bone formation was induced by the subcutaneous transplantation of human bone marrow-derived MSCs in nude mice, MSCs overexpressing a miR-124 inhibitor significantly enhanced woven bone formation compared with control MSCs. However, MSCs overexpressing a miR-124 mimic exhibited increased adipocyte differentiation at the expense of ectopic bone formation. These results suggest that miR-124 is a negative regulator of osteogenic differentiation and in vivo bone formation and that the targeting of Dlx5, Dlx3, and Dlx2 genes partly contributes to this inhibitory effect exerted by miR-124.

Expression of GAD67 and Dlx5 in the Taste Buds of Mice Genetically Lacking Mash1

It has been reported that a subset of type III taste cells express glutamate decarboxylase (GAD)67, which is a molecule that synthesizes gamma-aminobutyric acid (GABA), and that Mash1 could be a potential regulator of the development of GABAnergic neurons via Dlx transcription factors in the central nervous system. In this study, we investigated the expression of GAD67 and Dlx in the embryonic taste buds of the soft palate and circumvallate papilla using Mash1 knockout (KO)/GAD67-GFP knock-in mice. In the wild-type animal, a subset of type III taste cells contained GAD67 in the taste buds of the soft palate and the developing circumvallate papilla, whereas GAD67-expressing taste bud cells were missing from Mash1 KO mice. A subset of type III cells expressed mRNA for Dlx5 in the wild-type animals, whereas Dlx5-expressing cells were not evident in the apical part of the circumvallate papilla and taste buds in the soft palate of Mash1 KO mice. Our results suggest that Mash1 is required for the expression of GAD67 and Dlx5 in taste bud cells.

BMP-Mediated Functional Cooperation between Dlx5;Dlx6 and Msx1;Msx2 during Mammalian Limb Development

The Dlx and Msx homeodomain transcription factors play important roles in the control of limb development. The combined disruption of Msx1 and Msx2, as well as that of Dlx5 and Dlx6, lead to limb patterning defects with anomalies in digit number and shape. Msx1;Msx2 double mutants are characterized by the loss of derivatives of the anterior limb mesoderm which is not observed in either of the simple mutants. Dlx5;Dlx6 double mutants exhibit hindlimb ectrodactyly. While the morphogenetic action of Msx genes seems to involve the BMP molecules, the mode of action of Dlx genes still remains elusive. Here, examining the limb phenotypes of combined Dlx and Msx mutants we reveal a new Dlx-Msx regulatory loop directly involving BMPs. In Msx1;Dlx5;Dlx6 triple mutant mice (TKO), beside the expected ectrodactyly, we also observe the hallmark morphological anomalies of Msx1;Msx2 double mutants suggesting an epistatic role of Dlx5 and Dlx6 over Msx2. In Msx2;Dlx5;Dlx6 TKO mice we only observe an aggravation of the ectrodactyly defect without changes in the number of the individual components of the limb. Using a combination of qPCR, ChIP and bioinformatic analyses, we identify two Dlx/Msx regulatory pathways: 1) in the anterior limb mesoderm a non-cell autonomous Msx-Dlx regulatory loop involves BMP molecules through the AER and 2) in AER cells and, at later stages, in the limb mesoderm the regulation of Msx2 by Dlx5 and Dlx6 occurs also cell autonomously. These data bring new elements to decipher the complex AER-mesoderm dialogue that takes place during limb development and provide clues to understanding the etiology of congenital limb malformations.

Proneural Transcription Factors Regulate Different Steps of Cortical Neuron Migration through Rnd-Mediated Inhibition of RhoA Signaling

SummaryLittle is known of the intracellular machinery that controls the motility of newborn neurons. We have previously shown that the proneural protein Neurog2 promotes the migration of nascent cortical neurons by inducing the expression of the atypical Rho GTPase Rnd2. Here, we show that another proneural factor, Ascl1, promotes neuronal migration in the cortex through direct regulation of a second Rnd family member, Rnd3. Both Rnd2 and Rnd3 promote neuronal migration by inhibiting RhoA signaling, but they control distinct steps of the migratory process, multipolar to bipolar transition in the intermediate zone and locomotion in the cortical plate, respectively. Interestingly, these divergent functions directly result from the distinct subcellular distributions of the two Rnd proteins. Because Rnd proteins also regulate progenitor divisions and neurite outgrowth, we propose that proneural factors, through spatiotemporal regulation of Rnd proteins, integrate the process of neuronal migration with other events in the neurogenic program.

The Rb/E2F pathway modulates neurogenesis through direct regulation of the Dlx transcription factors

The Rb/E2F pathway modulates neurogenesis through direct regulation of the Dlx transcription factors

Role for TGF-β superfamily signaling in telencephalic GABAergic neuron development

Signaling mechanisms mediated by the Transforming Growth Factor-β (TGF-β) superfamily regulate a variety of developmental processes. Here we show that components of both bone morphogenetic protein/growth differentiation factor and TGF-β/activin/Nodal branches of TGF-β superfamily signaling are expressed in the developing subpallium. Furthermore, Smad proteins, transcriptional effectors of TGF-β signaling, are co-expressed and physically interact in the basal ganglia with Dlx homeodomain transcription factors, which are critical regulators of the differentiation, migration and survival of telencephalic GABAergic neurons. We also show that Dlx and Smad proteins localize to promoters/enhancers of a number of common telencephalic genes in vivo and that Smad proteins co-activate transcription with Dlx family members, except with certain mutated human DLX proteins identified in autistic individuals. In agreement with these observations, expression of dominant-negative Smads in the developing basal ganglia phenocopies the cell migration defects observed in Dlx1/2-deficient mice. Together, these results suggest that TGF-β superfamily signaling plays a role in telencephalic GABAergic neuron development through functional interactions with Dlx transcription factors.Electronic supplementary materialThe online version of this article (doi:10.1007/s11689-009-9035-6) contains supplementary material, which is available to authorized users.

Balanced gene regulation by an embryonic brain ncRNA is critical for adult hippocampal GABA circuitry

SummaryGenomic studies demonstrate that while the majority of the mammalian genome is transcribed, only about 2% of these transcripts are protein coding. We have been investigating how the long, polyadenylated Evf2 non-coding RNA regulates transcription of homeodomain transcription factors DLX5 and DLX6 in the developing mouse forebrain. Here we show that in developing ventral forebrain, Evf2 recruits DLX and MECP2 transcription factors to key DNA regulatory elements in the Dlx 5/6 intergenic region and controls Dlx5, Dlx6, and GAD67 expression through trans and cis-acting mechanisms. Evf2 mouse mutants have reduced numbers of GABAergic interneurons in early post-natal hippocampus and dentate gyrus. Although the numbers of GABAergic interneurons and GAD67 RNA levels return to normal in Evf2 mutant adult hippocampus, reduced synaptic inhibition occurs. These results suggest that non-coding RNA-dependent balanced gene regulation in embryonic brain is critical for proper formation of GABA-dependent neuronal circuitry in adult brain.

Dlx transcription factors regulate the Brn3b homeobox gene during vertebrate retina development

Dlx transcription factors regulate the Brn3b homeobox gene during vertebrate retina development

The speech vocalization gene, FoxP2, regulates interneuron fate decisions

Event Abstract Back to Event The speech vocalization gene, FoxP2, regulates interneuron fate decisions Ian C. Weaver1*, T. Sakurai2, J. D. Buxbaum2, D. R. Kaplan3, 4 and Freda D. Miller1, 4 1 University of Toronto, Developmental and Stem Cell Biology, Canada 2 Mount Sinai School of Medicine, Department of Psychiatry, Seaver Autism Center, United States 3 University of Toronto, Cell Biology, MaRS Center, Hospital for Sick Children, Canada 4 University of Toronto, Departments of Molecular Genetics and Physiology, Canada The transcription factor FoxP2 is expressed in the developing brain and is involved in speech development and susceptibility to autism. To determine the role of FoxP2 in the developing brain, we focused upon the developing cerebral cortex. Immunocytochemistry demonstrated that FoxP2 is expressed in a subset of nestin-positive precursors, and in newly born interneurons in both the cortex and the ganglionic eminence. In utero electroporation revealed that only a small subset of these cortical interneurons were generated from cortical precursors, consistent with the fact that most cortical interneurons migrate into the cortex from the ganglionic eminence. To ask about the potential role of FoxP2 in these cells, we ectopically-expressed FoxP2 in E12 cortical precursor cultures and E13/14 cortical precursors in vivo. In both cases, FoxP2 expression was sufficient to enhance the number of neurons expressing interneuron markers. Thus, FoxP2 is sufficient to promote interneuron genesis from precursors that normally make small numbers of interneurons. To ask if FoxP2 is also necessary for genesis of interneurons, we cultured E12 precursors from the medial ganglionic eminence (MGE), where approximately 40% of the precursors express FoxP2. ShRNA-mediated knockdown of FoxP2 in these cultures and MGE precursors in vivo decreased the number of interneurons generated. Thus, FoxP2 is also necessary for genesis of at least some MGE interneurons, raising the possibility that it might transcriptionally activate genes necessary for the interneuron phenotype, such as Dlx transcription factors. Consistent with this hypothesis, chromatin immunoprecipitation assays demonstrated that FoxP2 is associated with the Dlx-1 promoter in the embryonic brain, and heterologous cotransfection assays demonstrated that FoxP2 could transcriptionally activate the Dlx-1 promoter. Together these studies suggest that FoxP2 is essential for the normal genesis of cortical interneurons, and that perturbations in this activity might underlie the behavioral deficits in vocalization seen in individuals with FoxP2 mutations. Acknowledgements: This work was supported by grants from the Canadian Institutes of Health Research (CIHR) to F.D.M. and D.R.K. I.C.G.W. was initially supported by a Restracomp Postdoctoral Research Fellowship from the Hospital for Sick Children Foundation and presently by a Postdoctoral Research Fellowship from the CIHR. Conference: B.R.A.I.N. platform in Physiology poster day 2009, Toronto, ON, Canada, 16 Dec - 16 Dec, 2009. Presentation Type: Poster Presentation Topic: Poster presentations Citation: Weaver IC, Sakurai T, Buxbaum JD, Kaplan DR and Miller FD (2009). The speech vocalization gene, FoxP2, regulates interneuron fate decisions. Front. Neurosci. Conference Abstract: B.R.A.I.N. platform in Physiology poster day 2009. doi: 10.3389/conf.neuro.03.2009.17.058 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 18 Dec 2009; Published Online: 18 Dec 2009. * Correspondence: Ian C Weaver, University of Toronto, Developmental and Stem Cell Biology, Toronto, Canada, ianweaver01@hotmail.com Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Ian C Weaver T. Sakurai J. D Buxbaum D. R Kaplan Freda D Miller Google Ian C Weaver T. Sakurai J. D Buxbaum D. R Kaplan Freda D Miller Google Scholar Ian C Weaver T. Sakurai J. D Buxbaum D. R Kaplan Freda D Miller PubMed Ian C Weaver T. Sakurai J. D Buxbaum D. R Kaplan Freda D Miller Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Functional differentiation of a clone resembling embryonic cortical interneuron progenitors

We have generated clones (L2.3 and RG3.6) of neural progenitors with radial glial properties from rat E14.5 cortex that differentiate into astrocytes, neurons, and oligodendrocytes. Here, we describe a different clone (L2.2) that gives rise exclusively to neurons, but not to glia. Neuronal differentiation of L2.2 cells was inhibited by bone morphogenic protein 2 (BMP2) and enhanced by Sonic Hedgehog (SHH) similar to cortical interneuron progenitors. Compared with L2.3, differentiating L2.2 cells expressed significantly higher levels of mRNAs for glutamate decarboxylases (GADs), DLX transcription factors, calretinin, calbindin, neuropeptide Y (NPY), and somatostatin. Increased levels of DLX-2, GADs, and calretinin proteins were confirmed upon differentiation. L2.2 cells differentiated into neurons that fired action potentials in vitro, and their electrophysiological differentiation was accelerated and more complete when cocultured with developing astroglial cells but not with conditioned medium from these cells. The combined results suggest that clone L2.2 resembles GABAergic interneuron progenitors in the developing forebrain.

Msx and Dlx Homeogene Expression in Epithelial Odontogenic Tumors

Epithelial odontogenic tumors are rare jaw pathologies that raise clinical diagnosis and prognosis dilemmas notably between ameloblastomas and clear cell odontogenic carcinomas (CCOCs). In line with previous studies, the molecular determinants of tooth development-amelogenin, Msx1, Msx2, Dlx2, Dlx3, Bmp2, and Bmp4-were analyzed by RT-PCR, ISH, and immunolabeling in 12 recurrent ameloblastomas and in one case of CCOC. Although Msx1 expression imitates normal cell differentiation in these tumors, other genes showed a distinct pattern depending on the type of tumor and the tissue involved. In benign ameloblastomas, ISH localized Dlx3 transcripts and inconstantly detected Msx2 transcripts in epithelial cells. In the CCOC, ISH established a lack of both Dlx3 and Msx2 transcripts but allowed identification of the antisense transcript of Msx1, which imitates the same scheme of distribution between mesenchyme and epithelium as in the cup stage of tooth development. Furthermore, while exploring the expression pattern of signal molecules by RT-PCR, Bmp2 was shown to be completely inactivated in the CCOC and irregularly noticeable in ameloblastomas. Bmp4 was always expressed in all the tumors. Based on the established roles of Msx and Dlx transcription factors in dental cell fates, these data suggest that their altered expression is a proposed trail to explain the genesis and/or the progression of odontogenic tumors.

Dlx genes pattern mammalian jaw primordium by regulating both lower jaw-specific and upper jaw-specific genetic programs

Dlx transcription factors are implicated in patterning the mammalian jaw, based on their nested expression patterns in the first branchial arch (primordium for jaw) and mutant phenotypes; inactivation of Dlx1 and Dlx2 (Dlx1/2-/-) causes defects in the upper jaw, whereas Dlx5/6(-/-) results in homeotic transformation of the lower jaw into upper jaw. Therefore, the 'Dlx codes' appear to regionalize the jaw primordium such that Dlx1/2 regulate upper jaw development, while Dlx5/6 confer the lower jaw fate. Towards identifying the genetic pathways downstream of Dlx5/6, we compared the gene expression profiles of the wild-type and Dlx5/6(-/-) mouse mandibular arch (prospective lower jaw). We identified 20 previously unrecognized Dlx5/6-downstream genes, of which 12 were downregulated and 8 upregulated in the mutant. We found a Dlx-regulated transcriptional enhancer in close proximity to Gbx2, one of the Dlx5/6-downstream genes, strongly suggesting that Gbx2 is a direct target of Dlx5/6. We also showed that Pou3f3 is normally expressed in the maxillary (prospective upper jaw) but not mandibular arch, is upregulated in the mandibular arch of Dlx5/6(-/-), and is essential for formation of some of the maxillary arch-derived skeleton. A comparative analysis of the morphological and molecular phenotypes of various Dlx single and double mutants revealed that Dlx1, 2, 5 and 6 act both partially redundantly and antagonistically to direct differential expression of downstream genes in each domain of the first branchial arch. We propose a new model for Dlx-mediated mammalian jaw patterning.

Dlx Transcription Factors Promote Migration through Repression of Axon and Dendrite Growth

In the mouse telencephalon, Dlx homeobox transcription factors are essential for the tangential migration of subpallial-derived GABAergic interneurons to neocortex. However, the mechanisms underlying this process are poorly understood. Here, we demonstrate that Dlx1/2 has a central role in restraining neurite growth of subpallial-derived immature interneurons at a stage when they migrate tangentially to cortex. In Dlx1-/-;Dlx2-/- mutants, neurite length is increased and cells fail to migrate. In Dlx1-/-;Dlx2+/- mutants, while the tangential migration of immature interneurons appears normal, they develop dendritic and axonal processes with increased length and decreased branching, and have deficits in their neocortical laminar positions. Thus, Dlx1/2 is required for coordinating programs of neurite maturation and migration. In this regard, we provide genetic evidence that in immature interneurons Dlx1/2 repression of the p21-activated serine/threonine kinase PAK3, a downstream effector of the Rho family of GTPases, is critical in restraining neurite growth and promoting tangential migration.

Cortical Interneurons Refuse to Follow the PAK

The tangential migration of GABAergic interneurons to the cortex requires the activity of several transcription factors, but the mechanisms involved have remained elusive. In this issue of Neuron, Rubenstein and colleagues demonstrate that the homeobox proteins Dlx1 and Dlx2 promote interneuron migration in part by preventing the premature expression of PAK3, a serine/threonine kinase that is later involved in neurite outgrowth and other aspects of interneuron maturation.

Emerging epilepsy models: insights from mice, flies, worms and fish.

Animal models provide a means to investigate fundamental mechanisms of abnormal electrical discharge (i.e., seizures). Understanding the pathogenesis of epilepsy and therapy development have greatly benefited from these models. Here we review recent mouse mutants featuring spontaneous seizures and simpler organisms. New genetically engineered mice provide additional insights to cellular mechanisms underlying seizure generation (BK calcium-activated potassium channels and interneuron-expressed sodium channels), genetic interactions that exacerbate seizure phenotype (Scn2a, Kcnq2 and background) and neurodevelopmental influences (Dlx transcription factors). Mutants for neuronal nicotinic acetylcholine receptors, Glut-1 deficiency and aquaporin channels highlight additional seizure phenotypes in mice. Additional models in Caenorhabditis elegans (Lis-1) and Danio rerio (pentylenetetrazole) highlight a reductionist approach. Taking further advantage of 'simple' organisms, antiepileptic drugs and genetic modifiers of seizure activity are being uncovered in Drosophila. Studies of epilepsy in mutant mice provide a framework for understanding critical features of the brain that regulate excitability. These, and as yet undiscovered, mouse mutants will continue to serve as the foundation for basic epilepsy research. Interestingly, an even greater potential for analyzing epileptic phenotypes may lie in the more widespread use of genetically tractable organisms such as worms, flies and zebrafish.

The vertebrate ortholog of Aristaless is regulated by Dlx genes in the developing forebrain

The Dlx transcription factors have a central role in controlling the development of gamma-aminobutyric acid (GABA)-ergic neurons in the forebrain. However, little is known about how they control the properties of GABAergic neurons. One candidate is the Aristaless (Arx) homeobox gene, which lies genetically downstream of the fly Dlx gene (Distal-less, Dll). The expression of Arx in the mouse forebrain includes Dlx-expressing territories, such us the ventral thalamus, parts of the hypothalamus, and the ganglionic eminences and their derivatives in the subpallial telencephalon, and is expressed, as with the Dlx genes, in cortical GABAergic neurons. By using gain-of-function and loss-of-function assays in mouse and chicken embryos, we show that the Dlx genes have a conserved role in regulating the expression of Arx in the forebrain of vertebrates. Ectopic expression of Dlx genes with electroporation in brain slices from mouse embryos and in the neural tube of chick embryos shows that Dlx genes are sufficient to induce Arx ectopically. Moreover, we provide evidence that the Dlx genes exert a functionally relevant role in regulating Arx in vivo, as shown by the severe reduction in the expression of Arx in Dlx1/2 double-knockout mice. Therefore, our results suggest evolutionarily conserved functions of Dlx genes in regulating Arx expression between Drosophila and vertebrates.