Imaging Cellular Signaling one Molecule at a Time

Imaging Cellular Signaling one Molecule at a Time

Thomas Schmidt
Universiteit Leiden

The interaction of G-protein-coupled receptors with G proteins is a key event in transmembrane signal transduction that leads to vital decision-making by the cell. Here, we applied single-molecule epifluorescence microscopy to study the mobility of both the Gß? and the Ga2 subunits of the G protein heterotrimer in comparison with the cAMP receptor responsible for chemotactic signaling in Dictyostelium discoideum. Our experimental results suggest that ~30% of the G protein heterotrimers exist in receptor-precoupled complexes. Upon stimulation in a chemotactic gradient, this complex dissociates, subsequently leading to a linear diffusion and collision amplification of the external signal. We further found that Gß? was partially immobilized and confined in an agonist-, F-actin- and Ga2-dependent fashion. This led to the hypothesis that functional nanometric domains exist in the plasma membrane, which locally restrict the activation signal, and in turn, lead to faithful and efficient chemotactic signaling.