The Epidermal Growth Factor Receptor (EGFR) is a prototypical receptor tyrosine kinase, which belongs to the ErbB family. EGFR signaling plays an important role in the cellular processes such as proliferation, migration and apoptosis. The function of EGFR is known to be strongly related to its localization and the state of activation in the plasma membrane. However, the spatial distribution of EGFR in the plasma membrane is still debated, mainly due to limited detection methods.
In this project, the organization of EGFR in live cell membranes is explored by the recently developed Imaging Fluorescence Correlation Spectroscopy (FCS) at physiological conditions. Imaging FCS is a multiplexed version of the commonly used confocal FCS, which can quantitatively measure diffusion coefficients simultaneously over hundreds of contiguous points and thereby provides maps of membrane dynamics with high temporal resolution and diffraction-limited spatial resolution. Here we use Imaging Total Internal Reflection FCS (ITIR-FCS), which uses total internal reflection as mode of excitation. This allows in particular observing membrane dynamics close to a cover slide surface with low background. Imaging FCS exploits its spatio-temporal information to calculate the so-called FCS diffusion law which can probe the nanoscopic membrane organization with microscopic measurements. The FCS diffusion law can determine the presence of domains and meshworks on the plasma membrane below the diffraction limit. In the current study, dynamics and organization of EGFR on plasma membranes is explored under different drug treatments with mβCD and latrunculin A. Preliminary results indicate that EGFR is localized in cholesterol sensitive domains on the plasma membrane while it couples at the same time to the cytoskeleton.