Our proposed studies will use the measurement of lateral diffusion of specific membrane proteins to study the lateral organization and dynamics of plasma membrane components. A major objective will be to further characterize the cytoplasmic and extracellular matrix factors which restrict the lateral mobility of components of animal cell membranes. Using various biological and physical means, we will perturb the interactions of membrane proteins with cellular structures and measure any resultant changes in lateral mobility using the fluorescence recovery after photobleaching (FRAP) technique. The results of these studies will allow us to identify those moieties of both the membrane proteins themselves and the structures peripheral to the membrane which are crucial to restraining lateral mobility. Using biophysical analysis, we will attempt to identify the mechanisms by which proteins are restricted in their lateral motions. Using recently developed digitized fluorescence microscopy (DFM) techniques in combination with the photobleaching method, we will ascertain whether the lateral mobility of membrane proteins consists of simple, isotropic dif- fusion or is a more complex process involving membrane flow and/or anisotropic diffusion. The feasibility of using DFM technology to study the kinetics of antigen aggregation reactions (patching) will be explored. It is anticipated that information generated in this project will ultimately lead to a deepened understanding of plasma membrane organization in normal and diseased states.
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