Bilateral regulation of EGFR activity and local PI dynamics observed with superresolution microscopy

Anionic lipid molecules, including phosphatidylinositol-4,5-bisphosphate (PI(4,5)P₂), are implicated in the regulation of epidermal growth factor receptor (EGFR). However, the role of the spatiotemporal dynamics of PI(4,5)P₂in the regulation of EGFR activity in living cells is not fully understood, as it is difficult to visualize the local lipid domains around EGFR. In this study, both EGFR and PI(4,5)P₂nanodomains in the plasma membrane were visualized using super-resolution single-molecule microscopy. The EGFR and PI(4,5)P₂nanodomains aggregated before stimulation with epidermal growth factor (EGF) through transient visits of EGFR to the PI(4,5)P₂nanodomains. The degree of coaggregation decreased after EGF stimulation and depended on phospholipase Cγ, the EGFR effector hydrolyzing PI(4,5)P₂. Artificial reduction in the PI(4,5)P₂content of the plasma membrane reduced both the dimerization and autophosphorylation of EGFR after stimulation with EGF. Inhibition of PI(4,5)P₂hydrolysis after EGF stimulation decreased phosphorylation of EGFR-Thr654. Thus, EGFR kinase activity and the density of PI(4,5)P₂around EGFR molecules were found to be mutually regulated.