9723532 Satir Regulative secretion in eukaryotic cells involves a large number of components that interact to control membrane fusion and exocytosis via a signal transduction cascade. In many systems, this cascade is initiated by an increase in cytosolic Ca2+. The targets of this Ca2+ increase are not completely understood. They probably include a phosphoglycoprotein, parafusin (PFUS), that is widely found from protozoa to mammalian cells. In Paramecium where PFUS has been carefully studied, a Ca2+-dependent dephosphoglucosylation of the molecule is clearly correlated with exocytosis. The possibility that PFUS is a critical component of the secretory apparatus has now been enhanced by the finding that PFUS localizes to the surface "coat" of the dense core secretory vesicles in Paramecium and to the cell membrane prior to exocytosis and that it changes its localization when exocytosis is induced. Coat proteins are important in many membranes to determine localization within the cell. In this proposal we seek to explore the function of PFUS in membrane fusion and exocytosis, including a possible relationship of PFUS to specific components of the 20S docking complex. This function may have fundamental consequences for the signal transduction cascade leading to exocytosis and secretory product release in tissues of multicellular organisms. The secretion of proteins from cells is an essential physiological process in cells as diverse as unicellular organisms to organs of animals. This process involves the intracellular packaging of secretory proteins into membrane-bounded vesicles that fuse with the cell's limiting membrane, or plasma membrane, to be secreted into extracellular space. A conserved protein, termed parafusin appears to be involved in the fusion of the secretory vesicles to the plasma membrane. The PI has found that the reversible attachment of a phosphoglucose to the parafusin protein may regulate the fusion of the secretory vesicle to the plasma membrane. This type of modification of a protein is apparently unique to this protein. The regulation of the attachment and removal of the phosphoglucose is proposed as a critical step to regulate the secretion of proteins from cells. This award will fund basic research in the functional role and regulation of parafusin phosphorylation in protein secretion.
