Single-molecule imaging of Wnt3A protein diffusion on living cell membranes

Abstract

Wnt proteins are secreted, hydrophobic, lipidated proteins found in all animals that play essential roles in development and disease. Lipid modification is thought to facilitate the interaction of the protein with its receptor, Frizzled, but may also regulate the transport of Wnt protein and its localisation at the cell membrane. Here, by employing single-molecule fluorescence techniques, we show that Wnt proteins associate with and diffuse on the plasma membranes of living cells in the absence of any receptor binding. We find that labelled Wnt3A transiently and dynamically associates with the membranes of Drosophila S2 cells, diffuses with Brownian kinetics on flattened membranes and on cellular protrusions, and does not transfer between cells in close contact. In S2R+ cells, which express Frizzled receptors, membrane diffusion rate is reduced and membrane residency time is increased. These results provide direct evidence of Wnt3A interaction with living cell membranes and represent a new system for investigating the dynamics of Wnt transport.

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Identifiers

ORCID for Agnieszka Janeczek: orcid.org/0000-0002-9745-8346

ORCID for Nicholas Evans: orcid.org/0000-0002-3255-4388

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