The purpose of this training grant is twofold: 1. to enhance the research experience of individuals with the Ph.D. degree in the area of cellular and molecular physiology in order to prepare them for academic careers, 2. to provide clinically trained fellows with adequate research training and experience to enable them to carry out independent research in an academic setting. The training objective is to educate four trainees annually in the study of membrane function, more particularly of membrane channels, transporters, pumps and receptors in a wide range of cell types. The research expertise will be acquired through a program lasting one to three years. Research areas cover the molecular and structural biology of membrane proteins, the biophysics of their function, the basic mechanisms whereby ions, water and organic molecules are transferred across biological membranes, the regulation of membrane proteins, the involvement of membranes in signal transduction, the biosynthesis, assembly, intracellular sorting, targeting and insertion of membrane proteins, protein-protein interactions, the signaling pathways in cell regulation and morphogenesis, and the integrated function of cells in epithelia, in whole kidney and transgenic animals. The biological systems being studied include isolated membrane proteins, crystal structures of channels, cDNA clones that encode these proteins, chimeric membrane proteins, mutant transport systems, cytoskeletal elements associated with membrane proteins, specific monoclonal antibodies, membrane fractions and membrane vesicles, liposomes, bilayer lipid membranes, organelles within cells, isolated separated single cells, cellular protoplasts, frog oocytes, symmetrical and polarized cells in culture, epithelial cell sheets in culture, isolated perfused renal tubules, renal tubule segments in situ, isolated epithelia, and transgenic animals. The methodologies which can be acquired though this program cover molecular biological techniques, expression cloning, genetic recombination, in vitro mutagenesis, transfection, expression of membrane proteins, protein isolation and chemistry, microarray assays, reconstitution of membrane components, x-ray crystallography, cryo-electron microscopy, electron- confocal- and fluorescence microscopy, digital video microscopy, immunocytochemistry, atomic force microscopy, molecular probing of membrane components, biosensors and imaging of intracellular composition, cell culturing, patch-clamping, intracellular and transepithelial voltage and conductance measurements, intracellular ion activity measurements, ion fluxes, microdissection, in vitro and in situ microperfusion, as well as kinetic modeling.
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