Analysis of GPI-AP Diffusion on Plasma Membrane of Endothelial Cell Using Photochromic Fluorescent Protein

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The viscosity of the endothelial cell membrane changes during cell migration or in response to shear stress exposure. Interestingly, the degree of viscosity change differs among the various regions of the endothelial cell. These local variations in membrane viscosity may contribute to spatially specific signal transductions. It is unclear, however, whether local membrane viscosity affects to membrane protein dynamics. To address this issue, we examined membrane protein diffusion at different sites in endothelial cells. We fused the photocromic fluorescent protein Dronpa to glycosil phosphatidylinositol-anchored protein (DGGPI) which diffuses on the extracellular surface of the endothelial cell membrane. The DGGPI diffusion coefficient D was 0.165 ± 0.013 μm2/s (mean ± SE) at 37°C. Examining different sites in the endothelial cells at 28°C (room temperature) revealed that the DGGPI diffusion coefficient at the cell edge was 27% lower than the value measured at the of cell body center.