Journal Article

Molecular dissection of Wnt3a-Frizzled8 interaction reveals essential and modulatory determinants of Wnt signaling activity

Žigman, Mihaela, Trageser, Benjamin, Patel, Trushar R, Rahm, Karolin, Gross, Julia Christina, Gradl, Dietmar, Holstein, Thomas, Steinbeisser, Herbert, Stetefeld, Jörg, Özbek, Suat, Kumar, Sumit, Boutros, Michael

2014

Overview

Background Wnt proteins are a family of secreted signaling molecules that regulate key developmental processes in metazoans. The molecular basis of Wnt binding to Frizzled and LRP5/6 co-receptors has long been unknown due to the lack of structural data on Wnt ligands. Only recently, the crystal structure of the Wnt8-Frizzled8-cysteine-rich-domain (CRD) complex was solved, but the significance of interaction sites that influence Wnt signaling has not been assessed. Results Here, we present an extensive structure-function analysis of mouse Wnt3a in vitro and in vivo . We provide evidence for the essential role of serine 209, glycine 210 (site 1) and tryptophan 333 (site 2) in Fz binding. Importantly, we discovered that valine 337 in the site 2 binding loop is critical for signaling without contributing to binding. Mutations in the presumptive second CRD binding site (site 3) partly abolished Wnt binding. Intriguingly, most site 3 mutations increased Wnt signaling, probably by inhibiting Wnt-CRD oligomerization. In accordance, increasing amounts of soluble Frizzled8-CRD protein modulated Wnt3a signaling in a biphasic manner. Conclusions We propose a concentration-dependent switch in Wnt-CRD complex formation from an inactive aggregation state to an activated high mobility state as a possible modulatory mechanism in Wnt signaling gradients.

• Publisher: BioMed Central,BioMed Central Ltd,Springer Nature B.V
• Subject: Acids / Amino Acid Sequence / Analysis / Animals / Binding sites / Biology / Biomedical and Life Sciences / Biotechnology industry / Colleges & universities / Embryo, Nonmammalian - metabolism / Genetic aspects / HEK293 Cells / Humans / Life Sciences / Ligands / Lipids / Medical research / Mice / Models, Biological / Models, Molecular / Molecular Sequence Data / Mutant Proteins - metabolism / Mutation / Physiological aspects / Point Mutation - genetics / Protein Binding / Protein Structure, Tertiary / Proteins / Receptors, G-Protein-Coupled - chemistry / Receptors, G-Protein-Coupled - metabolism / Research Article / Solubility / Structure-Activity Relationship / Wnt Signaling Pathway / Wnt3A Protein - chemistry / Wnt3A Protein - metabolism / Zebrafish - embryology