THE CARBOXYL-TERMINAL REGION OF HIV-1 NEF PROTEIN IS A CELL-SURFACE DOMAIN THAT CAN INTERACT WITH CD4(-CELLS() T)

Our previous studies have shown that the HIV-1 Nef Ag is expressed, at least in part, on the surface of infected cells. We demonstrated this by using membrane immunofluorescence and flow cytometry with Nef muri ne mAbs. To identify the domain of Nef exposed on the cell surface, ep itope mapping of these and a new mAb was performed by ELISAs by using several recombinant truncated Nef fusion proteins and synthetic peptid es. The results showed that mAbs F1, E7, E9, and 4H4 recognized Nef ep itopes located at amino acid residues 148-157, 192-206, 158-206, and 1 -33, respectively. The intensity of cell surface Nef staining was stro nger with mAbs E7 and E9 than with F1, and there was no staining by 4H 4, which indicates that the carboxyl-terminal region of Nef is predomi nantly exposed on the surface of HIV-1-infected T cell lines and PBMC. This surface Nef domain displayed high affinity for the surface of un infected CD4(+) T cells; because the binding of a soluble form of reco mbinant Nef protein to the cell surface was specifically blocked by th e E7 and E9 mAbs or by synthetic peptides that contained the carboxyl- terminal region of Nef. In addition, syncytium formation between infec ted and uninfected cells also was specifically reduced by the same mAb s or peptides. Thus, the cell surface domain of Nef seems to play an i mportant role in the interaction between HIV-1-infected and CD4(+) uni nfected T cells.