RESUMEN
The remarkable similarity in the profile of genetic activity and the frequent association of developmental defects of limb and craniofacial structures in mouse mutant and hereditary disorders point to the possibility that the development of the head and limb involves common morphogenetic mechanisms. Our recent studies on the impact of the loss of Twist function has highlighted the essential role of the basic helix-loop-helix transcription factor encoded by this gene on the development of both body parts. We have summarized in this review our findings on the molecular pathways that are disrupted in Twist mutant mouse embryos. Our results revealed an evolutionarily conserved function for Twist in mesodermal differentiation, and previously unrecognised effects of the loss-of-function mutation of this gene in the outgrowth and patterning of the limb and branchial arches,and neural crest cell migration. An important outcome of our study is the demonstration of a differential requirement for Twist in forelimb versus hindlimb development, and its functional interaction with Gli3 in specifying anterior digit formation. Further evidence of the conservation of the function of Twist in different species is highlighted by similarity in the spectrum of potential downstream targets and interacting genes of Twist that have been identified by genetic, functional and microarray analysis.
Asunto(s)
Proteínas Nucleares/metabolismo , Proteínas Nucleares/fisiología , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico , Tipificación del Cuerpo , Diferenciación Celular , División Celular , Movimiento Celular , Proteínas de Unión al ADN/metabolismo , Relación Dosis-Respuesta a Droga , Regulación hacia Abajo , Extremidades/embriología , Regulación del Desarrollo de la Expresión Génica , Ratones , Mutación , Cresta Neural/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Fenotipo , Transducción de Señal , Somitos/metabolismo , Factores de Transcripción/metabolismo , Proteína 1 Relacionada con TwistRESUMEN
In a screen for potential targets of regulation by TWIST in mouse embryos we isolated a fragment with homology to type II early transposon (ETn) and type D endogenous provirus (MusD) elements. Whole-mount in situ hybridization to E7.5-E13.5 mouse embryos reveals a tissue- and stage-specific expression pattern that contrasts with the previously reported lack of expression of ETn elements in mouse embryos beyond late gastrulation. Transcripts were detected in the epiblast at E7.5 and in the neural tube from E8.5 to E10.5. Later expression is predominantly confined to the mesodermal tissues of craniofacial structures, limb buds and somites. The tissue specificity of expression suggests tight regulation of the activity of this early transposon element during embryogenesis.
Asunto(s)
Desarrollo Embrionario/fisiología , Retrovirus Endógenos/genética , Gástrula/fisiología , Regulación del Desarrollo de la Expresión Génica/genética , Animales , Desarrollo Embrionario/genética , Regulación del Desarrollo de la Expresión Génica/fisiología , RatonesRESUMEN
Loss of Twist gene function arrests the growth of the limb bud shortly after its formation. In the Twist(-/-) forelimb bud, Fgf10 expression is reduced, Fgf4 is not expressed, and the domain of Fgf8 and Fgfr2 expression is altered. This is accompanied by disruption of the expression of genes (Shh, Gli1, Gli2, Gli3, and Ptch) associated with SHH signalling in the limb bud mesenchyme, the down-regulation of Bmp4 in the apical ectoderm, the absence of Alx3, Alx4, Pax1, and Pax3 activity in the mesenchyme, and a reduced potency of the limb bud tissues to differentiate into osteogenic and myogenic tissues. Development of the hindlimb buds in Twist(-/-) embryos is also retarded. The overall activity of genes involved in SHH signalling is reduced.Fgf4 and Fgf8 expression is lost or reduced in the apical ectoderm, but other genes (Fgf10, Fgfr2) involved with FGF signalling are expressed in normal patterns. Twist(+/-);Gli3(+/XtJ) mice display more severe polydactyly than that seen in either Twist(+/-) or Gli3(+/XtJ) mice, suggesting that there is genetic interaction between Twist and Gli3 activity. Twist activity is therefore essential for the growth and differentiation of the limb bud tissues as well as regulation of tissue patterning via the modulation of SHH and FGF signal transduction.
Asunto(s)
Extremidades/embriología , Factores de Crecimiento de Fibroblastos/genética , Proteínas Nucleares/genética , Proteínas Nucleares/fisiología , Transactivadores/genética , Factores de Transcripción , Animales , Factor 10 de Crecimiento de Fibroblastos , Factor 8 de Crecimiento de Fibroblastos , Factores de Crecimiento de Fibroblastos/biosíntesis , Proteínas Hedgehog , Heterocigoto , Hibridación in Situ , Etiquetado Corte-Fin in Situ , Operón Lac , Mesodermo/metabolismo , Ratones , Ratones Endogámicos C57BL , Mutación , Polidactilia/genética , Unión Proteica , Estructura Terciaria de Proteína , ARN Mensajero/metabolismo , Proteínas Tirosina Quinasas Receptoras/biosíntesis , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos , Receptores de Factores de Crecimiento de Fibroblastos/biosíntesis , Transducción de Señal , Factores de Tiempo , Proteína 1 Relacionada con TwistRESUMEN
In the mouse, Twist is required for normal limb and craniofacial development. We show that the aristaless-like transcription factors, Alx3 and Alx4 are downregulated in the Twist(-/-) mutant and may be potential targets of Twist. By suppression subtractive hybridization we isolated 31 and 18 unique clones representing mRNAs that are putatively downregulated and upregulated respectively in Twist(-/-) forelimb buds. These included genes encoding cytoskeletal components, metabolic enzymes, hemoglobin molecules, membrane transport proteins, components of transcription and translation complexes, protein modification enzymes and proteins related to cell proliferation and apoptosis. Differential expression of selected clones was validated by whole mount in situ hybridization to E10.5 wild-type and Twist(-/-) embryos. We show that four novel clones are expressed in the Twist-expressing craniofacial tissues and paraxial mesoderm and downregulated in Twist(-/-) embryos, raising the possibility that they are, in addition to genes of the Alx family, downstream targets of Twist.
Asunto(s)
Proteínas de Unión al ADN , Regulación del Desarrollo de la Expresión Génica , Esbozos de los Miembros/embriología , Esbozos de los Miembros/metabolismo , Mutación/genética , Proteínas Nucleares/genética , Factores de Transcripción , Animales , Secuencia de Bases , Secuencia Conservada , Regulación hacia Abajo , Perfilación de la Expresión Génica , Proteínas de Homeodominio/genética , Hibridación in Situ , Ratones , Datos de Secuencia Molecular , Proteínas/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Homología de Secuencia de Ácido Nucleico , Proteína 1 Relacionada con TwistRESUMEN
Twist encodes a basic helix-loop-helix transcription factor that is required for normal craniofacial morphogenesis in the mouse. Loss of Twist activity in the cranial mesenchyme leads to aberrant migratory behaviour of the neural crest cells, whereas Twist-deficient neural crest cells are located in an inappropriate location in the first branchial arch and display defective osteogenic and odontogenic differentiation (Soo et al. [2002] Dev. Biol. 247:251-270). Results of the present study further show that loss of Twist impacts on the patterning of the cranial ganglia and nerves but not that of the peripheral ganglia and nerves in the trunk region of the body axis. Analyses of the expression of molecular markers of early differentiation of the paraxial mesoderm and the histogenetic potency of somites of Twist(-/-) embryos reveal that Twist-deficient somites can differentiate into muscles, cartilage, and bones, albeit less prolifically. Twist function, therefore, is not essential for mesoderm differentiation. The poor growth of the Twist-deficient somites after transplantation to the ectopic site may be attributed to reduced proliferative capacity and extensive apoptosis of the paraxial mesoderm, suggesting that Twist is required for maintaining cell proliferation and viability in the mesodermal progenitors.
Asunto(s)
Tipificación del Cuerpo , Nervios Craneales/embriología , Nervios Craneales/metabolismo , Mesodermo/citología , Mesodermo/metabolismo , Proteínas Nucleares/metabolismo , Factores de Transcripción/metabolismo , Animales , Apoptosis , Huesos/citología , Huesos/embriología , Huesos/metabolismo , Región Branquial/citología , Región Branquial/metabolismo , Diferenciación Celular , Movimiento Celular , Proliferación Celular , Supervivencia Celular , Nervios Craneales/citología , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Embrión de Mamíferos/citología , Embrión de Mamíferos/embriología , Embrión de Mamíferos/metabolismo , Ganglios/citología , Ganglios/embriología , Ganglios/metabolismo , Regulación del Desarrollo de la Expresión Génica , Proteínas del Grupo de Alta Movilidad/genética , Proteínas del Grupo de Alta Movilidad/metabolismo , Ratones , Ratones Noqueados , Proteínas Nucleares/deficiencia , Proteínas Nucleares/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factores de Transcripción SOXE , Somitos/citología , Somitos/metabolismo , Factores de Transcripción/deficiencia , Factores de Transcripción/genética , Proteína 1 Relacionada con TwistRESUMEN
Loss of Twist function in the cranial mesenchyme of the mouse embryo causes failure of closure of the cephalic neural tube and malformation of the branchial arches. In the Twist(-/-) embryo, the expression of molecular markers that signify dorsal forebrain tissues is either absent or reduced, but those associated with ventral tissues display expanded domains of expression. Dorsoventral organization of the mid- and hindbrain and the anterior-posterior pattern of the neural tube are not affected. In the Twist(-/-) embryo, neural crest cells stray from the subectodermal migratory path and the late-migrating subpopulation invades the cell-free zone separating streams of cells going to the first and second branchial arches. Cell transplantation studies reveal that Twist activity is required in the cranial mesenchyme for directing the migration of the neural crest cells, as well as in the neural crest cells within the first branchial arch to achieve correct localization. Twist is also required for the proper differentiation of the first arch tissues into bone, muscle, and teeth.