Many animal viruses enter their respective host cells by the process of adsorptive endocytosis. This route of internalization is also used by many hormones and ligands which bind to specific receptors on the cell plasma membrane. After binding to the cell surface, viruses can follow different pathways dependent upon their particular structure and organization. For example, vesicular stomatitis virus (an enveloped, RNA virus) enters an acidic endosomal vesicle and, consequent to exposure to the acidic pH, is believed to fuse with the membrane of the vesicle. Fusion results in the release of viral nucleic acid into the cell cytoplasm. In contrast to VSV, SV40 (a non-enveloped, DNA virus) appears to translocate quickly from the endosomal vesicle to the nucleus where it is uncoated and its DNA released. The mechanisms of this specific and rapid translocation is not known. Our laboratory is studying the mechanisms by which two viruses (VSV and SV40) are """"""""targeted"""""""" to different and specific sites in the host cell. Specifically, we have asked: (1) are these specific binding sites for viruses at the cell surface; (2) what are the cellular and viral components which participte in binding; (3) do the binding sites contribute to the final """"""""destination"""""""" of the virus; (4) how do viruses cross cell membrane barriers; and (5) do viruses contain signals to direct their intracellular translocation? Multiiple approaches will be used in these studies. Radiolabelled virus will be used to permit the detection and characterization of specific viral binding as well as the fate of internalized virus. Cell fractionation, electron microscopy, and immunofluorescence microscopy will also be used to follow the pathway of viral infection. Fusion of virus with cell membranes will be monitored by fluorescence energy transfer and EM of in vitro fusion models. Attempts will be made to delineate specific, biologically-active domains of viral spike and capsid proteins by use of both synthetic peptides and their corresponding antibodies.
