RESUMEN
Head development in vertebrates proceeds through a series of elaborate patterning mechanisms and cell-cell interactions involving cephalic neural crest cells (CNCC). These cells undergo extensive migration along stereotypical paths after their separation from the dorsal margins of the neural tube and they give rise to most of the craniofacial skeleton. Here, we report that the silencing of the LKB1 tumor suppressor affects the delamination of pre-migratory CNCC from the neural primordium as well as their polarization and survival, thus resulting in severe facial and brain defects. We further show that LKB1-mediated effects on the development of CNCC involve the sequential activation of the AMP-activated protein kinase (AMPK), the Rho-dependent kinase (ROCK) and the actin-based motor protein myosin II. Collectively, these results establish that the complex morphogenetic processes governing head formation critically depends on the activation of the LKB1 signaling network in CNCC.
Asunto(s)
Proteínas Aviares/fisiología , Cresta Neural/fisiología , Proteínas Serina-Treonina Quinasas/fisiología , Proteínas Quinasas Activadas por AMP/fisiología , Animales , Proteínas Aviares/antagonistas & inhibidores , Proteínas Aviares/genética , Embrión de Pollo , Anomalías Craneofaciales/embriología , Anomalías Craneofaciales/genética , Regulación del Desarrollo de la Expresión Génica , Silenciador del Gen , Cabeza/embriología , Ratones , Ratones Noqueados , Cadenas Ligeras de Miosina/fisiología , Cresta Neural/citología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/deficiencia , Proteínas Serina-Treonina Quinasas/genética , Transducción de Señal/genética , Transducción de Señal/fisiología , Quinasas Asociadas a rho/fisiologíaRESUMEN
During the early steps of head development, ectodermal patterning leads to the emergence of distinct non-neural and neural progenitor cells. The induction of the preplacodal ectoderm and the neural crest depends on well-studied signalling interactions between the non-neural ectoderm fated to become epidermis and the prospective neural plate. By contrast, the involvement of the non-neural ectoderm in the morphogenetic events leading to the development and patterning of the central nervous system has been studied less extensively. Here, we show that the removal of the rostral non-neural ectoderm abutting the prospective neural plate at late gastrulation stage leads, in mouse and chick embryos, to morphological defects in forebrain and craniofacial tissues. In particular, this ablation compromises the development of the telencephalon without affecting that of the diencephalon. Further investigations of ablated mouse embryos established that signalling centres crucial for forebrain regionalization, namely the axial mesendoderm and the anterior neural ridge, form normally. Moreover, changes in cell death or cell proliferation could not explain the specific loss of telencephalic tissue. Finally, we provide evidence that the removal of rostral tissues triggers misregulation of the BMP, WNT and FGF signalling pathways that may affect telencephalon development. This study opens new perspectives on the role of the neural/non-neural interface and reveals its functional relevance across higher vertebrates.
Asunto(s)
Ectodermo/embriología , Animales , Apoptosis/genética , Apoptosis/fisiología , Tipificación del Cuerpo/genética , Tipificación del Cuerpo/fisiología , Embrión de Pollo , Ectodermo/metabolismo , Femenino , Regulación del Desarrollo de la Expresión Génica/genética , Regulación del Desarrollo de la Expresión Génica/fisiología , Ratones , Cresta Neural/embriología , Cresta Neural/metabolismo , Neurogénesis/genética , Neurogénesis/fisiología , Embarazo , Prosencéfalo/embriología , Prosencéfalo/metabolismo , Telencéfalo/embriología , Telencéfalo/metabolismoRESUMEN
The combinatorial expression of Hox genes is an evolutionarily ancient program underlying body axis patterning in all Bilateria. In the head, the neural crest (NC)--a vertebrate innovation that contributes to evolutionarily novel skeletal and neural features--develops as a structure free of Hox-gene expression. The activation of Hoxa2 in the Hox-free facial NC (FNC) leads to severe craniofacial and brain defects. Here, we show that this condition unveils the requirement of three Six genes, Six1, Six2, and Six4, for brain development and morphogenesis of the maxillo-mandibular and nasofrontal skeleton. Inactivation of each of these Six genes in FNC generates diverse brain defects, ranging from plexus agenesis to mild or severe holoprosencephaly, and entails facial hypoplasia or truncation of the craniofacial skeleton. The triple silencing of these genes reveals their complementary role in face and brain morphogenesis. Furthermore, we show that the perturbation of the intrinsic genetic FNC program, by either Hoxa2 expression or Six gene inactivation, affects Bmp signaling through the downregulation of Bmp antagonists in the FNC cells. When upregulated in the FNC, Bmp antagonists suppress the adverse skeletal and cerebral effects of Hoxa2 expression. These results demonstrate that the combinatorial expression of Six1, Six2, and Six4 is required for the molecular programs governing craniofacial and cerebral development. These genes are crucial for the signaling system of FNC origin, which regulates normal growth and patterning of the cephalic neuroepithelium. Our results strongly suggest that several congenital craniofacial and cerebral malformations could be attributed to Six genes' misregulation.
Asunto(s)
Tipificación del Cuerpo/genética , Huesos/metabolismo , Encéfalo/metabolismo , Regulación del Desarrollo de la Expresión Génica , Proteínas de Homeodominio/genética , Cresta Neural/metabolismo , Animales , Proteínas Morfogenéticas Óseas/antagonistas & inhibidores , Proteínas Morfogenéticas Óseas/genética , Proteínas Morfogenéticas Óseas/metabolismo , Huesos/embriología , Encéfalo/embriología , Embrión de Pollo , Electroporación , Embrión no Mamífero , Cabeza/embriología , Proteínas de Homeodominio/antagonistas & inhibidores , Proteínas de Homeodominio/metabolismo , Cresta Neural/embriología , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Transducción de SeñalRESUMEN
Cubilin (Cubn) is a multiligand endocytic receptor critical for the intestinal absorption of vitamin B12 and renal protein reabsorption. During mouse development, Cubn is expressed in both embryonic and extra-embryonic tissues, and Cubn gene inactivation results in early embryo lethality most likely due to the impairment of the function of extra-embryonic Cubn. Here, we focus on the developmental role of Cubn expressed in the embryonic head. We report that Cubn is a novel, interspecies-conserved Fgf receptor. Epiblast-specific inactivation of Cubn in the mouse embryo as well as Cubn silencing in the anterior head of frog or the cephalic neural crest of chick embryos show that Cubn is required during early somite stages to convey survival signals in the developing vertebrate head. Surface plasmon resonance analysis reveals that fibroblast growth factor 8 (Fgf8), a key mediator of cell survival, migration, proliferation, and patterning in the developing head, is a high affinity ligand for Cubn. Cell uptake studies show that binding to Cubn is necessary for the phosphorylation of the Fgf signaling mediators MAPK and Smad1. Although Cubn may not form stable ternary complexes with Fgf receptors (FgfRs), it acts together with and/or is necessary for optimal FgfR activity. We propose that plasma membrane binding of Fgf8, and most likely of the Fgf8 family members Fgf17 and Fgf18, to Cubn improves Fgf ligand endocytosis and availability to FgfRs, thus modulating Fgf signaling activity.
Asunto(s)
Factor 8 de Crecimiento de Fibroblastos/metabolismo , Cabeza/embriología , Sistema de Señalización de MAP Quinasas/fisiología , Cresta Neural/embriología , Receptores de Superficie Celular/metabolismo , Receptores de Factores de Crecimiento de Fibroblastos/metabolismo , Animales , Supervivencia Celular/fisiología , Endocitosis/fisiología , Quinasas MAP Reguladas por Señal Extracelular/genética , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Factor 8 de Crecimiento de Fibroblastos/genética , Factores de Crecimiento de Fibroblastos/genética , Factores de Crecimiento de Fibroblastos/metabolismo , Silenciador del Gen , Ligandos , Ratones , Ratones Transgénicos , Cresta Neural/citología , Unión Proteica , Receptores de Superficie Celular/genética , Receptores de Factores de Crecimiento de Fibroblastos/genéticaRESUMEN
The role of the neural crest (NC) in the construction of the vertebrate head was demonstrated when cell tracing techniques became available to follow the cells exiting from the cephalic neural folds in embryos of various vertebrate species. Experiments carried out in the avian embryo, using the quail/chick chimera system, were critical in showing that the entire facial skeleton and most of the skull (except for he occipital region) were derived from the NC domain of the posterior diencephalon, mesencephalon and rhombomeres 1 and 2 (r1, r2). This region of the NC was designated FSNC (for Facial Skeletogenic NC). One characteristic of this part of the head including the neural anlage is that it remains free of expression of the homeotic genes of the Hox-clusters. In an attempt to see whether this rostral Hox-negative domain of the NC has a specific role in the development of the skeleton, we have surgically removed it in chick embryos at 5-6 somite stages (5-6 ss). The operated embryos showed a complete absence of facial and skull cartilages and bones showing that the Hox expressing domain of the NC caudally located to the excision did not regenerate to replace the anterior NC. In addition to the deficit in skeletal structures, the operated embryos exhibited severe brain defects resulting in anencephaly. Experiments described here have shown that the neural crest cells regulate the amount of Fgf8 produced by the two brain organizers, the Anterior Neural Ridge (ANR) and the isthmus. This regulation is exerted via the secretion of anti-BMP signaling molecules (e.g. Gremlin and Noggin), which decrease BMP production hence enhancing the amount of Fgf8 synthesized in the ANR (also called "Prosencephalic organizer") and the isthmus. In addition to its role in building up the face and skull, the NC is therefore an important signaling center for brain development.
Asunto(s)
Encéfalo/embriología , Regulación del Desarrollo de la Expresión Génica , Morfogénesis/genética , Cresta Neural , Animales , Encéfalo/citología , Encéfalo/fisiología , Embrión de Pollo , Quimera , Embrión no Mamífero/embriología , Factor 8 de Crecimiento de Fibroblastos/genética , Cabeza/embriología , Cresta Neural/citología , Cresta Neural/embriología , Cresta Neural/fisiología , Codorniz , Cráneo/embriologíaRESUMEN
Emergence of the neural crest (NC) is considered an essential asset in the evolution of the chordate phylum, as specific vertebrate traits such as peripheral nervous system, cephalic skeletal tissues, and head development are linked to the NC and its derivatives. It has been proposed that the emergence of the NC was responsible for the formation of a "new head" characterized by the spectacular development of the forebrain and associated sense organs. It was previously shown that removal of the cephalic NC (CNC) prevents the formation of the facial structures but also results in anencephaly. This article reports on the molecular mechanisms whereby the CNC controls cephalic neurulation and brain morphogenesis. This study demonstrates that molecular variations of Gremlin and Noggin level in CNC account for morphological changes in brain size and development. CNC cells act in these processes through a multi-step control and exert cumulative effects counteracting bone morphogenetic protein signaling produced by the neighboring tissues (e.g., adjacent neuroepithelium, ventro-medial mesoderm, superficial ectoderm). These data provide an explanation for the fact that acquisition of the NC during the protochordate-to-vertebrate transition has coincided with a large increase of brain vesicles.
Asunto(s)
Encéfalo/embriología , Cresta Neural/embriología , Animales , Proteína Morfogenética Ósea 4/genética , Proteína Morfogenética Ósea 7/genética , Encéfalo/metabolismo , Proteínas Portadoras/metabolismo , Embrión de Pollo , Embrión no Mamífero/embriología , Embrión no Mamífero/metabolismo , Desarrollo Embrionario/genética , Factor 8 de Crecimiento de Fibroblastos/antagonistas & inhibidores , Factor 8 de Crecimiento de Fibroblastos/genética , Regulación del Desarrollo de la Expresión Génica , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Modelos Neurológicos , Cresta Neural/metabolismo , Cresta Neural/cirugía , Codorniz , Interferencia de ARNRESUMEN
The neural crest (NC), a defining feature of vertebrate embryo, generates most of the skeletal tissues encasing the developing forebrain and provides the prosencephalon with functional vasculature and meninges. Recent findings show that early in development, the cephalic NC is also essential for the pre-otic neural tube closure and promotes the development of the prosencephalic alar plate by regulating the morphogenetic activities of forebrain organizers.
Asunto(s)
Tipificación del Cuerpo , Movimiento Celular/fisiología , Factor 8 de Crecimiento de Fibroblastos/metabolismo , Cresta Neural/embriología , Prosencéfalo/embriología , Animales , Diferenciación Celular/fisiología , Humanos , RatonesRESUMEN
For decades, the quail-chick system has been a gold standard approach to track cells and their progenies over complex morphogenetic movements and long-range migrations as well as to unravel their dialogue and interplays in varied processes of cell induction. More specifically, this model became decisive for the systematic explorations of the neural crest and its lineages and allowed a tremendous stride in understanding the wealth and complexity of this fascinating cell population. Much of our knowledge on craniofacial morphogenesis and vertebrate organogenesis was first gained in avian chimeras and later extended to mammalian models and humans. In addition, this system permits tissue and gene manipulations to be performed at once in the same cell population. Through the use of in ovo electroporation, this model became tractable for functional genomics, hence being even more resourceful for functional studies. Due to the ease of access and the possibility to combine micromanipulation of tissue anlagen and gene expression, this model offers the prospect of decrypting instructive versus permissive tissue interactions, to identify and crack the molecular codes underlying cell positioning and differentiation, with an unparalleled spatiotemporal accuracy.
Asunto(s)
Cresta Neural/citología , Animales , Pollos , Electroporación , Desarrollo Embrionario , Genómica , CodornizRESUMEN
Encephalisation is the most important characteristic in the evolutionary transition leading from protochordates to vertebrates. This event has coincided with the emergence of a transient and pluripotent structure, the neural crest (NC), which is absent in protochordates. In vertebrates, NC provides the rostral cephalic vesicles with skeletal protection and functional vascularization. The surgical extirpation of the cephalic NC, which is responsible for building up the craniofacial skeleton, results in the absence of facial skeleton together with severe defects of preotic brain development, leading to exencephaly. Here, we have analyzed the role of the NC in forebrain and midbrain development. We show that (i) NC cells (NCC) control Fgf8 expression in the anterior neural ridge, which is considered the prosencephalic organizer; (ii) the cephalic NCC are necessary for the closure of the neural tube; and (iii) NCC contribute to the proper patterning of genes that are expressed in the prosencephalic and mesencephalic alar plate. Along with the development of the roof plate, NCC also concur to the patterning of the pallial and subpallial structures. We show that the NC-dependent production of FGF8 in anterior neural ridge is able to restrict Shh expression to the ventral prosencephalon. All together, these findings support the notion that the cephalic NC controls the formation of craniofacial structures and the development of preotic brain.