Wnt7a interaction with Fzd5 and detection of signaling activation using a split eGFP.
Wnts are secreted glycoproteins that regulate important cellular processes including proliferation, differentiation, and cell fate. In the beta-catenin/canonical pathway, Wnt interacts with Fzd receptors to inhibit degradation of beta-catenin and promote its translocation into the nucleus where it regulates transcription of a number of genes. Dysregulation of this pathway has been attributed to a host of diseases including cancer. As a result, components of the beta-catenin/canonical pathway have been gaining recognition as promising targets for the discovery of novel therapeutic agents. Here, we show, using an ELISA-based protein-protein binding assay that purified Wnt7a binds to the extracellular cysteine-rich domain of Fzd5 in the nanomolar range. We have developed a novel split eGFP complementation assay to visually detect Wnt7a-Fzd5 interactions and subsequent pathway activation in cells. These biological tools could help lead to a better understanding of Wnt-Fzd interactions and the identification of new modulators of Wnt signaling.
Carmon
Kendra S
KS
Department of Integrative Biology and Pharmacology, University of Texas Health Science Center Houston, 6431 Fannin Street, Houston, TX 77030, USA.
Loose
David S
DS
D051157:Frizzled Receptors / Q000737:chemistry* / Q000378:metabolism*; D049452:Green Fluorescent Proteins; D006801:Humans; D025941:Protein Interaction Mapping / Q000379:methods*; D043562:Receptors, G-Protein-Coupled / Q000737:chemistry* / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D051153:Wnt Proteins / Q000737:chemistry* / Q000378:metabolism*
C498589:FZD5 protein, human; D051157:Frizzled Receptors; D043562:Receptors, G-Protein-Coupled; C092778:WNT7A protein, human; D051153:Wnt Proteins; D049452:Green Fluorescent Proteins