A virus-free cell fusion assay relying on the transient transfection of Epstein-Barr virus (EBV) glycoproteins into cells provides an efficient and quantitative assay for characterizing the viral requirements necessary for fusion of the viral envelope with the B cell membrane. Extensive cellular fusion occurred when Daudi cells were layered onto Chinese hamster ovary K1 cells transiently expressing EBV glycoproteins gp42, gH, gL, and gB. This is the first direct evidence that gB is involved in the process of EBV entry. Moreover, mutational analysis of gB indicates that the cytoplasmic tail contains two distinct domains that function differentially in the process of fusion. The region from amino acids 802 to 816 is necessary for productive membrane fusion, while amino acids 817 to 841 comprise a domain that negatively regulates membrane fusion.
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