The role of Wnt signalling in the development of somites and neural crest

The Wnt family of secreted signalling molecules controls a wide range of developmental processes in all metazoans. In this investigation we concentrate on the role that members of this family play during the development of (1) the somites and (2) the neural crest. (3) We also isolate a novel component of the Wnt signalling pathway called Naked cuticle and investigate the role that this protein may play in both of the previously mentioned developmental processes. (1) In higher vertebrates the paraxial mesoderm undergoes a mesenchymal-to-epithelial transformation to form segmentally organised structures called somites. Experiments have shown that signals originating from the ectoderm overlying the somites or from midline structures are required for the formation of the somites, but their identity has yet to be determined. Wnt6 is a good candidate as a somite epithelialisation factor from the ectoderm since it is expressed in this tissue. In this study we show that injection of Wnt6-producing cells beneath the ectoderm at the level of the segmental plate or lateral to the segmental plate leads to the formation of numerous small epithelial somites. We show that Wnts are indeed responsible for the epithelialisation of somites by applying Wnt antagonists which result in the segmental plate being unable to form somites. These results show that Wnt6, the only member of this family to be localised to the chick paraxial ectoderm, is able to regulate the development of epithelial somites and that cellular organisation is pivotal in the execution of the differentiation programmes. (2) The neural crest is a population of multipotent progenitor cells that arise from the neural ectoderm in all vertebrate embryos and form a multitude of derivatives including the peripheral sensory neurons, the enteric nervous system, Schwann cells, pigment cells and parts of the craniofacial skeleton. The induction of the neural crest relies on an ectodermally derived signal, but the identity of the molecule performing this role in amniotes is not known. Here we show that Wnt6, a protein expressed in the ectoderm, induces neural crest production. (3) The intracellular response to Wnt signalling depends on the choice of signalling cascade activated in the responding cell. Cells can activate either the canonical pathway that modulates gene expression to control cellular differentiation and proliferation, or the non-canonical pathway that controls cell polarity and movement (Pandur et al. 2002b). Recent work has identified the protein Naked cuticle as an intracellular switch promoting the non-canonical pathway at the expense of the canonical pathway. We have cloned chick Naked cuticle-1 (cNkd1) and demonstrate that it is expressed in a dynamic manner during early embryogenesis. We show that it is expressed in the somites and in particular regions where cells are undergoing movement. Lastly our study shows that the expression of cNkd1 is regulated by Wnt expression originating from the neural tube. This study provides evidence that non-canonical Wnt signalling plays a part in somite development.

Wnt6 controls amniote neural crest induction through the non-canonical signaling pathway

The neural crest is a multipotent embryonic cell population that arises from neural ectoderm and forms derivatives essential for vertebrate function. Neural crest induction requires an ectodermal signal, thought to be a Writ ligand, but the identity of the Wnt that performs this function in amniotes is unknown. Here, we demonstrate that Wnt6, derived from the ectoderm, is necessary for chick neural crest induction. Crucially, we also show that Wnt6 acts through the non-canonical pathway and not the beta-catenin-dependant pathway. Surprisingly, we found that canonical Wnt signaling inhibited neural crest production in the chick embryo. In light of studies in anamniotes demonstrating that canonical Wnt signaling induces neural crest, these results indicate a significant and novel change in the mechanism of neural crest induction during vertebrate evolution. These data also highlight a key role for noncanonical Wnt signaling in cell type specification from a stem population during development.

Molecular evidence from ascidians for the evolutionary origin of vertebrate cranial sensory placodes

Cranial sensory placodes are specialised areas of the head ectoderm of vertebrate embryos that contribute to the formation of the cranial sense organs and associated ganglia. Placodes are often considered a vertebrate innovation, and their evolution has been hypothesised as one key adaptation underlying the evolution of active predation by primitive vertebrates. Here, we review recent molecular evidence pertinent to understanding the evolutionary origin of placodes. The development of vertebrate placodes is regulated by numerous genes, including members of the Pax, Six, Eya, Fox, Phox, Neurogenin and Pou gene families. In the sea squirt Ciona intestinalis (a basal chordate and close relative of the vertebrates), orthologues of these genes are deployed in the development of the oral and atrial siphons, structures used for filter feeding by the sessile adult. Our interpretation of these findings is that vertebrate placodes and sea squirt siphon primordia have evolved from the same patches of specialised ectoderm present in the common ancestor of the chordates.

Molecular evidence from Ciona intestinalis for the evolutionary origin of vertebrate sensory placodes

Cranial sensory placodes are focused areas of the head ectoderm of vertebrates that contribute to the development of the cranial sense organs and their associated ganglia. Placodes have long been considered a key character of vertebrates, and their evolution is proposed to have been essential for the evolution of an active predatory lifestyle by early vertebrates. Despite their importance for understanding vertebrate origins, the evolutionary origin of placodes has remained obscure. Here, we use a panel of molecular markers from the Six, Eya, Pax, Dach, FoxI, COE and POUIV gene families to examine the tunicate Ciona intestinalis for evidence of structures homologous to vertebrate placodes. Our results identify two domains of Ciona ectoderm that are marked by the genetic cascade that regulates vertebrate placode formation. The first is just anterior to the brain, and we suggest this territory is equivalent to the olfactoty/adenohypophyseal placodes of vertebrates. The second is a bilateral domain adjacent to the posterior brain and includes cells fated to form the atrium and atrial siphon of adult Ciona. We show this bares most similarity to placodes fated to form the vertebrate acoustico-lateralis system. We interpret these data as support for the hypothesis that sensory placodes did not arise de novo in vertebrates, but evolved froth pre-existing specialised areas of ectoderm that contributed to sensory organs in the common ancestor of vertebrate and tunicates. Published by Elsevier Inc.

Expression of AmphiCoe, an amphioxus COE/EBF gene, in the developing central nervous system and epidermal sensory neurons

The COE/EBF gene family marks a subset of prospective neurons in the vertebrate central and peripheral. nervous system; including neurons deriving from some ectodermal placodes. Since placodes are often considered unique to vertebrates, we have characterised an amphioxus COE/EBF gene with the aim of using it as a marker to examine the timing and location of peripheral neuron differentiation. A single COE/EBF family member, AmphiCoe, was isolated from the amphioxus Branchiostoma floridae: AmphiCoe lies basal to the vertebrate COE/EBF genes in molecular phylogenetic analysis, suggesting that the duplications that formed the vertebrate COE/EBF family were specific to the vertebrate lineage. AmphiCoe is expressed in the central nervous system and in a small number of scattered ectodermal cells on the flanks of neurulae stage embryos. These cells become at least largely recessed beneath the ectoderm. Scanning electron microscopy was used to examine embryos in which the ectoderm had been partially peeled away. This revealed that these cells have neuronal morphology, and we infer that they are the precursors of epidermal primary sensory neurons. These characters lead us to suggest that differentiation of some ectodermal cells into sensory neurons with a tendency to sink beneath the embryonic surface represents a primitive feature that has become incorporated into placodes during vertebrate evolution. (C) 2004 Wiley-Liss, Inc.

Protochordate Zic genes define primitive somite compartments and highlight molecular changes underlying neural crest evolution

The vertebrate Zic gene family encodes C2H2 zinc finger transcription factors closely related to the Gli proteins. Zic genes are expressed in multiple areas of developing vertebrate embryos, including the dorsal neural tube where they act as potent neural crest inducers. Here we describe the characterization of a Zic ortholog from the amphioxus Branchiostoma floridae and further describe the expression of a Zic ortholog from the ascidian Ciona intestinalis. Molecular phylogenetic analysis and sequence comparisons suggest the gene duplications that formed the vertebrate Zic family were specific to the vertebrate lineage. In Ciona maternal CiZic/Ci-macho1 transcripts are localized during cleavage stages by asymmetric cell division, whereas zygotic expression by neural plate cells commences during neurulation. The amphioxus Zic ortholog AmphiZic is expressed in dorsal mesoderm and ectoderm during gastrulation, before being eliminated first from midline cells and then from all neurectoderm during neurulation. After neurulation, expression is reactivated in the dorsal neural tube and dorsolateral somite. Comparison of CiZic and AmphiZic expression with vertebrate Zic expression leads to two main conclusions. First, Zic expression allows us to define homologous compartments between vertebrate and amphioxus somites, showing primitive subdivision of vertebrate segmented mesoderm. Second, we show that neural Zic expression is a chordate synapomorphy, whereas the precise pattern of neural expression has evolved differently on the different chordate lineages. Based on these observations we suggest that a change in Zic regulation, specifically the evolution of a dorsal neural expression domain in vertebrate neurulae, was an important step in the evolution of the neural crest.

Adult craniofacial stem cells: sources and relation to the neural crest

During the process of development, neural crest cells migrate out from their niche between the newly formed ectoderm and the neural tube. Thereafter, they give rise not only to ectodermal cell types, but also to mesodermal cell types. Cell types with neural crest ancestry consequently comprise a number of specialized varieties, such as ectodermal neurons, melanocytes and Schwann cells, as well as mesodermal osteoblasts, adipocytes and smooth muscle cells. Numerous recent studies suggest that stem cells with a neural crest origin persist into adulthood, especially within the mammalian craniofacial compartment. This review discusses the sources of adult neural crest-derived stem cells (NCSCs) derived from the cranium, as well as their differentiation potential and expression of key stem cell markers. Furthermore, the expression of marker genes associated with embryonic stem cells and the issue of multi- versus pluripotency of adult NCSCs is reviewed. Stringent tests are proposed, which, if performed, are anticipated to clarify the issue of adult NCSC potency. Finally, current pre-clinical and clinical data are discussed in light of the clinical impact of adult NCSCs.