Study Of The Molecular Mechanism Of Dkk1-Mediated Inhibition Of Canonical Wnt Pathway

Wnt signaling pathway is a conservative signaling pathway, playing very important roles in diverse physiological processes. Dkk is one of the important antagonists in Wnt signaling pathway and was found to specifically inhibit canonical Wnt pathway. Two models have been proposed to explain molecular mechanism of Dkk's inhibition role in canonical Wnt pathway: in the first model, Dkk1 competes for binding of Wnt co-receptor LRP6; in the second model, Dkk1 can form a ternary complex with LRP6 and Kremen at the presence of Kremen, and triggers the endocytosis of this ternary complex quickly. The controversies between these two models exist during these years. In this project, we studied these two models respectively. Our results suggest that both these two models function in Dkk's inhibition role and there is no conflict between them. Using semi-quantity immuno-staining assay, we firstly detected that Dkk1 can inhibit the formation of Wnt-Frizzled-LRP6 ternary complex. In the study of the molecular mechanism of endocytosis proposed in the long-term effect, we found that the intracellular domain of Kremen is required for the endocytosis of Dkk1-LRP6-Kremen ternary complex, and it also plays important roles in Dkk's inhibition role in Wnt canonical pathway. Our results also suggested that this endocytosis is Clathrin-dependent and AP-2 might be the adaptor protein that recognizes the ternary complex. Moreover, we found that Kremen can interact with LRP6 independent of Dkk, and full-length Kremen can even promote the membrane localization of LRP6, indicating that Kremen might play other roles besides assisting Dkk to inhibit canonical Wnt pathway.