Membrane capacitance recordings resolve dynamics and complexity of receptor-mediated endocytosis in Wnt signalling

Abstract

Receptor-mediated endocytosis is an essential process in signaling pathways for an activation of intracellular signaling cascades. One example is the Wnt signaling pathway, which seems to depend on endocytosis of the ligand-receptor complex for initiation of Wnt signal transduction. So far, the role of different endocytic pathways in Wnt signaling, the molecular players and the kinetics of this process are unclear. Here, we monitor endocytosis in Wnt3a and Wnt5a mediated signaling by membrane capacitance recordings of HEK293 cells. Our measurements revealed a fast and substantial increase in the number of endocytic vesicles. This endocytotic activity is specifically elicited by extracellular Wnt ligands; it starts immediately upon ligand binding and ceases over a period of ten minutes. By using specific inhibitors, we can dissect Wnt induced endocytosis into two independent pathways, where canonical Wnt3a is taken up mainly by clathrin-independent endocytosis and Wnt5a exclusively by clathrin-mediated endocytosis.