WDFY1 is a key adaptor protein in Axl receptor tyrosine kinase signaling 2310082

Abstract Introduction The WD repeat and FYVE-domain containing protein 1 (WDFY1) encodes a phosphatidylinositol 3-phosphate (PI3P) binding protein localized to early endosomes. WDFY1 involves in endosomal trafficking, cell growth signaling, and autophagy and serves as a key adaptor protein in immune signaling. Axl receptor tyrosine kinase is critical in eliminating apoptotic debris and regulating cell proliferation/survival. We found a close association between Axl and WDFY1 expression. We wondered if WDFY1 serves as an adaptor protein for Axl signal transduction. Methods Mesangial cells were cultured and treated with siRNAs targeting WDFY1 and Axl expression. Cells were then stimulated with Axl agonists. Protein-protein interactions were investigated with co-immunoprecipitation and verified with immunofluorescence co-localization and proximity ligation assay (PLA). Axl signaling cascades were studied by Western blotting with anti-phospho antibodies. WDFY1 C347S point mutation were constructed and transfected into mesangial cells to study the importance of the WDFY1 FYVE domain in the process. Results We found that Axl activation leads to its internalization and merge with early endosomes, where PI3P is generated. WDFY1 binds to PI3P upon Axl activation and then recruits Akt to the site transducing downstream signal cascades. WDFY1 depletion and C347S mutation resulted in reduced Akt phosphorylation and disrupted PI3P-WDFY1 interaction. Conclusion Our data suggest a key role of WDFY1 in bridging the PI3P-Akt interaction, which is necessary for Axl signaling. Funding Source n/a Topic Categories Immune Response Regulation: Molecular Mechanisms (IRM)