Human Immunodeficiency Virus (HIV) is a sexually transmitted disease whose spread has become increasingly devastating worldwide. In order for productive HIV transmission to occur between infected males and healthy females, virus in human semen must transverse the mucosal epithelial barrier designed in part to protect the underlying tissues of the genital tract from pathogens. In these studies, we identify a host protein, gp340, which is expressed in cell lines and tissue-derived from the human female genital epithelium. In contrast to other splice variants of the protein expressed elsewhere in the body, gp340 expression induces binding and enhancement of transmission of various strains of HIV-1. Through inhibition of the interaction of gp340 with the envelope protein of HIV-1 (Env or gp120), we show that the enhancement observed in gp340 expressing cells is dependent on direct interaction of gp340 with virions. This protein-protein interaction can influence the ability of cells that express gp340 to bind, sequester and transmit infectious virus to healthy targets of infection in coculture. The previously described ability of certain genital epithelial cells to directly transcytose HIV, led to interest in determining the potential contribution of gp340 to this process. Gp340 was identified as a significant contributor to the transyctosis in both cell lines and mucosal tissue. These observations when taken together present gp340 as a potentially significant mediator of one of the earliest steps in HIV transmission.