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- Title
EGFR-dependent endocytosis of Wnt9a and Fzd9b promotes β-catenin signaling during hematopoietic stem cell development in zebrafish.
- Authors
Nguyen, Nicole; Carpenter, Kelsey A.; Ensing, Jessica; Gilliland, Carla; Rudisel, Emma J.; Mu, Emily M.; Thurlow, Kate E.; Triche Jr., Timothy J.; Grainger, Stephanie
- Abstract
Cell-to-cell communication through secreted Wnt ligands that bind to members of the Frizzled (Fzd) family of transmembrane receptors is critical for development and homeostasis. Wnt9a signals through Fzd9b, the co-receptor LRP5 or LRP6 (LRP5/6), and the epidermal growth factor receptor (EGFR) to promote early proliferation of zebrafish and human hematopoietic stem cells during development. Here, we developed fluorescently labeled, biologically active Wnt9a and Fzd9b fusion proteins to demonstrate that EGFR-dependent endocytosis of the ligand-receptor complex was required for signaling. In human cells, the Wnt9a-Fzd9b complex was rapidly endocytosed and trafficked through early and late endosomes, lysosomes, and the endoplasmic reticulum. Using small-molecule inhibitors and genetic and knockdown approaches, we found that Wnt9a-Fzd9b endocytosis required EGFR-mediated phosphorylation of the Fzd9b tail, caveolin, and the scaffolding protein EGFR protein substrate 15 (EPS15). LRP5/6 and the downstream signaling component AXIN were required for Wnt9a-Fzd9b signaling but not for endocytosis. Knockdown or loss of EPS15 impaired hematopoietic stem cell development in zebrafish. Other Wnt ligands do not require endocytosis for signaling activity, implying that specific modes of endocytosis and trafficking may represent a method by which Wnt-Fzd specificity is established. Editor's summary: Wnt9a-mediated activation of the receptor Fzd9b is required for hematopoietic stem cell development in zebrafish and for specification of human hematopoietic stem cells in a manner dependent on epidermal growth factor receptor (EGFR). Nguyen et al. found that EGFR-mediated phosphorylation of the tail of Fzd9b induced endocytosis of the Wnt9a-Fzd9b complex that was necessary for signaling. The adaptor protein EPS15, which promotes the endocytosis of EGFR and other receptors, was also required for Wnt9a-Fzd9b endocytosis in cells and for the development of hematopoietic stem cells in zebrafish. Thus, Wnt9a-Fzd9b complexes stimulate downstream signaling only after they are endocytosed in an EGFR- and EPS15-dependent manner. —Annalisa M. VanHook
- Subjects
HEMATOPOIETIC stem cells; WNT signal transduction; EPIDERMAL growth factor receptors; ENDOCYTOSIS; HUMAN stem cells; BRACHYDANIO
- Publication
Science Signaling, 2024, Vol 17, Issue 832, p1
- ISSN
1945-0877
- Publication type
Article
- DOI
10.1126/scisignal.adf4299