Structural and functional analysis of Wntless in Wnt signaling

Abstract

Wnt signal transduction plays important roles in numerous phases of organismal development and tissue homeostasis. When aberrant, Wnt signaling can drive cell transformation that results in cancer and various developmental diseases. Over the years, it has become clear that Wnt activity is strictly regulated, not only in signal-receiving cells, but also in signal-producing cells at the level of Wnt production, transport and secretion. Post-translational modifications of Wnts are critical for Wnt signal regulation at the level of intracellular trafficking, extracellular dissemination, and ligand-receptor interaction. Modified Wnts are escorted through the secretory pathway for extracellular release by Wntless (Wls), the dedicated Wnt chaperone protein. Wls directly interacts with lipid-modified Wg ( Drosophila Wnt) within the ER of the signal-producing cells in order to assist its navigation through the secretory pathway for cell surface deployment, an indispensible step for downstream signal activation. In this study, I have shown that DWls ( Drosophila Wls) forms homo-oligomeric structures required for Wg interaction, in which both its first transmembrane domain (aa14 to aa35) and amino acids between 137 and 157 play a role. While the presence of the Wg binding region was not a requirement for DWls oligomers to form, DWls oligomerization was requisite for the Wg-DWls interaction; thus, likely necessary for proper Wg intracellular transport. Additionally, mutational studies in our laboratory revealed that Cysteines C50 and C72 are important for intermolecular disulfide bridges and oligomer formation. These studies also revealed that DWls is efficiently N-glycosylated at Asparagine 58. While glycosylation of DWls was not required for Wg binding or DWls oligomerization, it was required for extracellular release of DWls. The exact mechanism of DWls oligomer formation and its effect on Wg binding and release remain to be further investigated.