RESUMO
BACKGROUND: The homeobox containing transcription factor Uncx4.1 is, amongst others, expressed in the mouse midbrain. The early expression of this transcription factor in the mouse, as well as in the chick midbrain, points to a conserved function of Uncx4.1, but so far a functional analysis in this brain territory is missing. The goal of the current study was to analyze in which midbrain neuronal subgroups Uncx4.1 is expressed and to examine whether this factor plays a role in the early development of these neuronal subgroups. RESULTS: We have shown that Uncx4.1 is expressed in GABAergic, glutamatergic and dopaminergic neurons in the mouse midbrain. In midbrain dopaminergic (mDA) neurons Uncx4.1 expression is particularly high around E11.5 and strongly diminished already at E17.5. The analysis of knockout mice revealed that the loss of Uncx4.1 is accompanied with a 25% decrease in the population of mDA neurons, as marked by tyrosine hydroxylase (TH), dopamine transporter (DAT), Pitx3 and Ngn2. In contrast, the number of glutamatergic Pax6-positive cells was augmented, while the GABAergic neuron population appears not affected in Uncx4.1-deficient embryos. CONCLUSION: We conclude that Uncx4.1 is implicated in the development of mDA neurons where it displays a unique temporal expression profile in the early postmitotic stage. Our data indicate that the mechanism underlying the role of Uncx4.1 in mDA development is likely related to differentiation processes in postmitotic stages, and where Ngn2 is engaged. Moreover, Uncx4.1 might play an important role during glutamatergic neuronal differentiation in the mouse midbrain.
Assuntos
Diferenciação Celular/genética , Neurônios Dopaminérgicos/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/genética , Proteínas de Homeodomínio/metabolismo , Mesencéfalo , Fatores Etários , Animais , Animais Recém-Nascidos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Bromodesoxiuridina/metabolismo , Contagem de Células , Morte Celular/genética , Proteínas da Membrana Plasmática de Transporte de Dopamina/metabolismo , Embrião de Mamíferos , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Proteínas de Homeodomínio/genética , Imunoprecipitação , Marcação In Situ das Extremidades Cortadas , Mesencéfalo/citologia , Mesencéfalo/embriologia , Mesencéfalo/crescimento & desenvolvimento , Camundongos , Camundongos Transgênicos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismoRESUMO
This article reports the identification and characterization of a DBL-like guanine nucleotide exchange factor (GEF) in Drosophila, called GEFmeso, as a novel binding target of the Ras-like GTPase Ral. Previous studies suggested that some aspects of Ral activity, which is involved in multiple cellular processes, are mediated through regulation of Rho GTPases. Here we show in vitro association of GEFmeso with the GTP-bound active form of Ral and the nucleotide-free form of the Rho GTPase Cdc42. GEFmeso fails to bind to other Rho GTPases, showing that Cdc42 is a specific interaction partner of this GEF. Unlike Ral and Cdc42, which are ubiquitously expressed, GEFmeso exerts distinct spatio-temporal expression patterns during embryonic development, suggesting a tissue-restricted function of the GEF in vivo. Based on previous observations that mutations in Cdc42 or overexpression of mutant alleles of Cdc42 lead to distinct effects on wing development, the effects of overexpression of dominant-negative and activated versions of Ral on wing development were analyzed. In addition, GEFmeso overexpression studies as well as RNAi experiments were performed. The results suggest that Ral, GEFmeso and Cdc42 act in the same developmental pathway and that GEFmeso mediates activation of Cdc42 in response to activated Ral in the context of Drosophila wing development.
Assuntos
Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Proteínas ral de Ligação ao GTP/metabolismo , Animais , Mapeamento Cromossômico , Proteínas de Drosophila/química , Drosophila melanogaster/embriologia , Regulação da Expressão Gênica no Desenvolvimento , Fatores de Troca do Nucleotídeo Guanina/química , Ligação Proteica , Asas de Animais/metabolismoRESUMO
Translational control plays a key role in many biological processes including pattern formation during early Drosophila embryogenesis. In this process, the anterior determinant Bicoid (BCD) acts not only as a transcriptional activator of segmentation genes but also causes specific translational repression of ubiquitously distributed caudal (cad) mRNA in the anterior region of the embryo. We show that translational repression of cad mRNA is dependent on a functional eIF4E-binding motif. The results suggest a novel mode of translational repression, which combines the strategy of target-specific binding to 3'-untranslated sequences and interference with 5'-cap-dependent translation initiation in one protein.
Assuntos
Regiões 5' não Traduzidas/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Proteínas de Insetos/metabolismo , Biossíntese de Proteínas , Capuzes de RNA/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas Repressoras/metabolismo , Transativadores/metabolismo , Sequência de Aminoácidos , Animais , Animais Geneticamente Modificados , Proteínas de Drosophila , Drosophila melanogaster , Fator de Iniciação 4E em Eucariotos/metabolismo , Humanos , Proteínas de Insetos/genética , Dados de Sequência Molecular , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas Repressoras/genética , Transativadores/genética , Fatores de TranscriçãoRESUMO
We report the identification of Ima, a novel Drosophila MAGUK-like protein, which contains two WW and four PDZ protein interaction domains and interacts with the small GTPase dRal in the yeast two-hybrid system and pull-down assays. The gene is expressed in distinct spatiotemporal patterns throughout embryonic development. Overexpression of Ima interferes with normal Drosophila development, indicating that the gene functions in a tissue specific manner.
Assuntos
Proteínas de Transporte/genética , Proteínas de Drosophila/genética , Drosophila/genética , Núcleosídeo-Fosfato Quinase , Sequência de Aminoácidos , Animais , Animais Geneticamente Modificados , Sítios de Ligação , Proteínas de Transporte/biossíntese , Proteínas de Transporte/metabolismo , Drosophila/metabolismo , Proteínas de Drosophila/biossíntese , Proteínas de Drosophila/metabolismo , Perfilação da Expressão Gênica , Dados de Sequência Molecular , Especificidade de Órgãos , Análise de Sequência de Proteína , Transgenes , Técnicas do Sistema de Duplo-HíbridoRESUMO
We report the identification of Ima, a novel Drosophila MAGUK-like protein, which contains two WW and four PDZ protein interaction domains and interacts with the small GTPase dRal in the yeast two-hybrid system and pull-down assays. The gene is expressed in distinct spatiotemporal patterns throughout embryonic development. Overexpression of Ima interferes with normal Drosophila development, indicating that the gene functions in a tissue specific manner.
