Development of fluorescence-based methods to study the effect of cytoplasmic proteins, cholesterol, and extracellular ligand on integrin diffusion and clustering in single cells

Components of the cell membrane play a critical role in overall cell health and are often the target of various drug treatments. Cell membrane organization can provide insight into how cells respond to their environment and ultimately provide the knowledge needed to understand disease regulation. Integrins are one of the main transmembrane proteins on animal cells that integrate the extracellular matrix with the intracellular cytoskeleton and are involved in many cellular functions including signaling, growth, and differentiation. It is the goal of this work to measure integrin clustering and diffusion properties in the presence of altered extracellular, membrane, and cellular compositions in order to elucidate the molecular mechanisms of integrins. FRET, fluorescence resonance energy transfer, is a technique that measures interactions on the 0 to 10 nanometer scale. FRAP, fluorescence recovery after photobleaching, monitors the two-dimensional diffusion of fluorescent molecules into a previously photobleached area