Neural development


Early neural tissue development and neural crest formation

The central nervous system forms in vertebrate embryos as a result of inductive interactions between competent ectoderm and a special dorsal signaling center, known in amphibians as the Spemann organizer. Organizer cells, corresponding to dorsal mesoderm, secrete a large number of factors, which can activate neural tissue specific genes in responding ectoderm. Complex interactions between the products encoded by these genes result in the differentiation of neurons and glial cells, organized in a specific mediolateral and anteroposterior pattern. Several signaling pathways, triggered by bone morphogenetic proteins (BMP), fibroblast growth factors (FGF) and Wnt proteins were implicated in early neural development in vertebrates, however, underlying molecular mechanisms are not understood in sufficient detail.

Our studies identified Frodo/Dapper/DACT, a new signaling protein that is a founding member of a family of signaling proteins (reviewed by Brott and Sokol, 2005). Using antisense morpholino oligonucleotides (MO), an efficient in vivo method of inactivating a gene product, we have demonstrated an essential role for Frodo in neural development (Hikasa and Sokol, 2004). Depletion of Frodo resulted in a selective defect in early organizer genes, such as Chordin, Xenopus nodal-related 3 and Cerberus. The involvement of Frodo in organizer formation and organizer-specific gene expression is supported by deficient head development in Frodo-depleted embryos (Hikasa and Sokol, 2004). Targeted depletion of Frodo in neuroectoderm prevented activation of the neural markers Sox2 and Nrp1 at the earliest stages of neural plate formation, demonstrating a requirement for this signaling protein in vertebrate neural induction (Hikasa and Sokol, 2004).

A new project in our group concerns the development of neural crest (NC) cells, which form at the neural plate border as multipotent progenitors, undergo epithelial-mesenchymal transition and migrate to diverse locations in vertebrate embryos to give rise to many cell types. Multiple signaling factors, including Wnt proteins, operate during early embryonic development to induce the NC cell fate. Whereas the requirement for the Wnt/-catenin pathway in NC specification has been well established, a similar role for Wnt proteins that do not stabilize beta-catenin has remained unclear. Our gain- and loss-of-function experiments implicate Wnt11-like proteins in NC specification in Xenopus embryos. In support of this conclusion, modulation of beta-catenin-independent signaling through Dishevelled and Ror2 causes predictable changes in premigratory NC.  Morpholino-mediated depletion experiments suggest that Wnt11R, a Wnt protein that is expressed in neuroectoderm adjacent to the NC territory, is required for NC formation. Wnt11-like signals might specify NC by altering the localization and activity of the serine/threonine polarity kinase PAR-1 (also known as microtubule-associated regulatory kinase or MARK), which itself plays an essential role in NC formation. Consistent with this model, PAR-1 RNA rescues NC markers in embryos, in which noncanonical Wnt signaling has been blocked. These experiments identify novel roles for Wnt11R and PAR-1 in NC specification and reveal an unexpected connection between morphogenesis and cell fate.

Asymmetric division of neural stem / progenitor cells

In Drosophila neuroblasts, cell fates are regulated by asymmetric distribution of molecular determinants and the direction of the mitotic spindle. In vertebrate embryos and mammalian cells the significance of asymmetric division and spindle orientation for cell type specification is less well studied. We would like to understand the role for mitotic spindle orientation and asymmetric cell division of neural progenitor cells (NPCs) in neuronal cell fate determination in vertebrates.

Both Lgl and the PAR proteins function to regulate cell polarity in many embryonic tissues. Lgl has been shown to control asymmetric cell division and differentiation of Drosophila neuroblasts and mouse neural tube. We are studying the subcellular localization of Lgl and PAR proteins in cells undergoing neural differentiation and assess how modulation of their levels or localization would influence neural tissue differentiation. The knowledge of molecular mechanisms regulating neuronal differentiation should have implications on stem cell research and regenerative medicine.

Our studies demonstrate that Lgl, PAR-1 and atypical PKC, as well as some other polarity proteins appear to be required for cell polarity and asymmetric cell division during primary neurogenesis in Xenopus ectoderm, and its localization may be controlled by Wnt signaling (Dollar et al., 2005; Ossipova et al., 2009; Lake and Sokol, 2009). As neural stem cells may undergo similar asymmetric divisions, we hypothesize that polarity proteins regulate neuronal differentiation. We obtained experimental support for this hypothesis in several models (Ossipova et al., 2009; Lake and Sokol, 2009). Since the Wnt signaling pathway is known to be involved in the regulation of cell polarity and mitotic spindle orientation in C. elegans embryos, we use frog embryos and mouse progenitor cells to investigate a role for Wnt signaling in asymmetric cell divisions during vertebrate neurogenesis.