The proteins that span lipid bilayers are extremely important in nature. It is their role to selectively bridge the barriers that maintain cellular organization. In particular, those proteins that span the plasma membrane of cells not only mediate the transfer of metabolites into and out of cells, but also the transduction of signals across the cell boundary. Alterations in the cell surface have been noted in many diseased tissues including tumors. It remains largely a mystery how these important proteins are transported from their site of synthesis to the cell surface. Biogenesis of the glycoprotein (G) of vesicular stomatitis virus (VSV) has been widely studied as a model for the biogenesis of plasma membrane glycoproteins that span the lipid bilayer. We have been using immunofluorescence microscopy and cryoultramicrotomy in conjunction with immunoelectron microscopy to study the cellular structures through which this protein passes en route to the plasma membrane. The use of cryoultramicrotomy (ultra-thin frozen section technology) is of tremendous importance when attempting to localize antigens within a tissue or cell. By cutting through the membrane that effectively compartmentalize such structures, one renders the antigen accessible to antibodies during immunostaining. We propose to examine: 1) Intermeidates in the intracellular pathway followed by the VSV G protein. 2) The role of the overall structural organization of the cytoplasm in the intracellular transport of the G protein. 3) The mechanisms involved in the sorting of membrane and secretory proteins. 4) The biogenesis of the interaction between an integral membrane protein (the VSV G protein) and a peripheral membrane protein (the VSV M protein). It is hoped that these studies will contribute to our understanding of the creation and maintainance of the cell surface and subcellular organization.
