Surface-bound Regulation of Actin Polymerization

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DESCRIPTION (provided by applicant): Changes in cell shape are important for immunological defense against pathogens and for normal tissue development, including cell migration. Aberrant regulation is a key point in some disease processes, including metastases of cancer. Because the actin cortex is responsible for these changes in cell shape, our long-term goal is to understand its regulation and its mechanisms for changing shape. We have pioneered new biophysical approaches and novel nanotechnology for probing the mechanical functions of the actin cortex. Cortical regulation often occurs with membrane-bound factors controlling a patch of cortex containing many barbed ends. By examining ARP2/3 and Ena/VASP-dependent reactions, we will probe the physicochemical constraints of membrane-surface catalysis in two modes of regulating cortical actin: lamellae and filopodia protrusions. Essentially building a model of the leading edges of cells, our longer-term Aim is to reconstitute these representative reactions for direct visualization on nanofabricated surfaces. Even without visualization, surface-adsorption already reveals biochemical interactions unexpected from solution studies. Surface-activation of ARP2/3 reveals a novel mechanical role for capping protein, whereas surface-activation of Ena/VASP suggests that certain proposed biochemical associations are not required to speed protrusion. Constituting our first two Specific Aims, understanding these two effects also requires biophysical measurements to monitor mechanical properties. In addition, understanding these two representative processes will provide a rational basis to guide development of the third, longer-term Aim of visualizing cortical dynamics.