Neurons are quintessential polarized cells possessing axonal and dendritic processes with distinct morphologies, signaling properties, cytoskeletal architectures and physiological activities. Neuronal polarity is essential for the assembly and function of the nervous system, yet the cellular and molecular mechanisms that polarize neurons and shape the structures of their axons and dendrites are not well understood. Secreted Wnt proteins establish the anteroposterior (AP) polarity of the neurons ALM and PLM in the nematode Caenorhabditis elegans. Disruption of Wnt signaling leads to a complete inversion of ALM and PLM polarity: the anterior process adopts the length, branching pattern and synaptic properties of the wild- type posterior process and vice versa.
In Aim 1, we plan to identify and characterize the role of other Wnt pathway genes in this process by analyzing mutants and RNAi animals and determining the expression pattern, subcellular localization and site of action of their gene products. We will test whether Wnts act as instructive or permissive signals and investigate whether Wnt signaling influences centrosome position. Precise regulation of Wnt receptors is crucial for the correct temporal and spatial response to different Wnts expressed throughout development. The conserved transmembrane RING finger protein PLR-1 modulates the response to Wnts by reducing the cell surface levels of the Frizzled family of Wnt receptors.
In Aim 2, we will address whether PLR-1 acts by trafficking Frizzleds to endosomes instead of the plasma membrane or by increasing their internalization. We will assay PLR-1 for E3 ubiquitin protein ligase activity and determine whether Frizzleds are targets of PLR-1 ubiquitination. The retromer, which mediates endosome-to-Golgi trafficking, functions in Wnt-producing cells to form an extracellular Wnt gradient.
In Aim 3, we will investigate whether retromer controls trafficking of MOM-3/Wntless/Evi, a multipass transmembrane protein that acts to promote Wnt secretion. A greater understanding of the molecular basis of neuronal polarity is instrumental for the development of methods to treat neuronal tissue damage caused by trauma or neurodegenerative disease. Highly conserved Wnt signaling pathways play a fundamental role in organizing the nervous system of vertebrates and nematodes; as such, our studies in C. elegans can provide insight into Wnt-mediated processes involved in human development and disease. ? ?
