Molecular characterization of wdr68 gene in embryonic development of Xenopus laevis.

WDR68, also known as DCAF7, is a WD40 repeated domain protein highly conserved in eukaryotic organisms in both plants and animals. This protein participates in numerous cellular processes and exerts its function through interaction with other proteins. In the present work, we isolated, sequenced and characterized cDNA corresponding to the wdr68 gene in embryos of the amphibian Xenopus laevis. Syntenic analysis revealed high conservation of the genomic region containing the WDR68 locus in amniotes. Nevertheless, in fishes and amphibians, we observed that the tandem genes surrounding wdr68 undergoes certain rearrangements with respect to the organization found in amniotes. We also defined the temporal and spatial expression pattern of the wdr68 gene in the development of Xenopus laevis through whole mount in situ hybridization and RT-PCR techniques. We observed that wdr68 is ubiquitously expressed during early embryonic development but, during the neurula stage, it undergoes a strong expression in the neural tube and in the migratory cephalic streams of the neural crest. At the tailbud stages, it is strongly expressed in the cephalic region, particularly in otic and optic vesicles, in addition to branchial arches. In contrast, wdr68 transcripts are localized in the somitic mesoderm in the trunk. The expression area that includes the migratory neural crest of the head and the branchial arches suggest that this gene would be involved in jaws formation, probably through a hierarchical relationship with the component genes of the endothelin-1/endothelin receptor type A cell signaling pathway.

Functional analysis of Hairy genes in Xenopus neural crest initial specification and cell migration.

BACKGROUND: Neural crest formation is one of the fundamental processes in the early stages of embryonic development in vertebrates. This transient and multipotent embryonic cell population is able to generate a variety of tissues and cell types in the adult body. hairy genes are transcription factors that contain a basic helix-loop-helix domain which binds to DNA. In Xenopus three hairy genes are known: hairy1, hairy2a, and hairy2b. The requirement of hairy genes was explored in early neural crest development although the late requirements of these genes during neural crest maintenance, migration and derivatives formation are still unknown. RESULTS: In this work, we extended the analysis of Xenopus hairy genes expression patterns and described new domains of expression. Functional analysis showed that hairy genes are required for the induction and migration of the neural crest and for the control of apoptosis. Moreover, we showed that hairy genes function as transcriptional repressors and that they are down-regulated by bone morphogenetic protein-Smad signaling and positively regulated by the Notch/Delta-Su(h) pathway. CONCLUSIONS: Our results indicate that hairy genes have a functional equivalence between them and that they are required for multiple processes during neural crest development.

A new role for the Endothelin-1/Endothelin-A receptor signaling during early neural crest specification.

The neural crest is induced at the border of the neural plate in a multistep process by signals emanated from the epidermis, neural plate and mesoderm. In this work we show for the first time the existence of a neural crest maintenance step which is dependent on signals released from the mesoderm. We identified Endothelin-1 (Edn1) and its receptor (Ednra) as key players of this signal and we show that Edn1/Ednra signaling is required for maintenance of the neural crest by a dual mechanism of cell specification and cell survival. We show that: (i) Ednra is expressed in prospective neural crest; (ii) loss-of-function experiments with antisense morpholino or with specific chemical inhibitor suppress the expression of early neural crest markers; (iii) gain-of-function experiments expand the neural crest territory; (iv) epistatic experiments show that Ednra/Edn1 is downstream of the early neural crest gene Msx1 and upstream of the late genes Sox9 and Sox10; and (v) Edn1/Ednra signaling inhibits apoptosis and controls cell specification of the neural crest. Together, our results provide insight on a new role of Edn1/Ednra cell signaling pathway during early neural crest development.