DESCRIPTION: Initial Epstein-Barr virus (EBV) infection occurs via the oropharyngeal mucosa, but mechanisms of EBV entry into epithelial cells are poorly understood. Using polarized oropharyngeal epithelial cells, we showed that EBV enters these cells through two CD21-independent pathways: direct cell-to-cell contact of apical epithelial cell membranes with EBV-infected lymphocytes, and entry of cell-free EBV virions through basolateral membranes. Our data show that interaction between the EBV BMRF-2 glycoprotein and beta1 integrin play a key role in cell-free virion entry. We showed that cell-to-cell spread of progeny versions from infected into uninfected cells occurs across lateral membranes. The amino acid sequence of BMRF-2 reveals several potential basolateral-sorting signals, and we have shown that it is transported to basolateral membranes. We posit that BMRF-2 may be involved in targeting of intracellular progeny virions to the basolateral membrane and may facilitate translocation of virions across lateral intercellular junctions. The goal of this study is to characterize molecular mechanisms of BMRF-2/beta1 integrin interactions and their functional significance in EBV infection and dissemination in single-layer polarized and multi-layer stratified oropharyngeal epithelial cells. The Specific Aims of the study are: 1) To investigate the role of EBV BMRF- 2 in virion attachment and entry in oropharyngeal epithelial cells; 2) To identify the sorting signals of BMRF-2 that govern its transport to the basolateral membranes of polarized oropharyngeal epithelial cells; and 3) To study the functions of the BMRF-2 protein in EBV cell-to-cell spread in oropharyngeal epithelium. These data will greatly advance knowledge of the mechanisms of EBV infection of oropharyngeal epithelial cells and may be of value to design new approaches to prevention of initial EBV infection.