MUTATIONAL ANALYSIS OF HIV-1 GP160-MEDIATED RECEPTOR INTERFERENCE - INTRACELLULAR COMPLEX-FORMATION

Formation of CD4-gp160 intracellular complexes represents an important mechanism leading to the induction of receptor interference. Previous studies have demonstrated that cells coexpressing gp160 and CD4 forme d complexes of CD4 and gp160 which became blocked within the endoplasm ic reticulum (ER), preventing CD4 from reaching the cell surface. In t his report we have investigated the domains and residues of CD4 and gp 160 involved in intracellular interaction. Accordingly, we have introd uced mutations in both CD4 and gp160 at sites previously shown to disr upt CD4-gp120 interactions at the cell surface. Using a T7-vaccinia Vi rus transient expression system, we expressed these gp160 and CD4 muta nts in HeLa cells and analyzed their effects on intracellular complex formation and CD4 surface modulation. we observed that a number of gp1 60 mutants which failed to interact with CD4 at the cell surface also failed to bind and trap CD4 within the ER as expected. However, mutati ons at a critical residue, W427, did not abrogate intracellular CD4 bi nding. These gp160 mutants continued to interact with intracellular CD 4 and inhibit CD4 transport to the cell surface, although gp120 produc ed from these mutants did not bind CD4 at the cell surface as expected . A number CD4 mutants also continued to form intracellular complexes with gp160, resulting in the loss of CD4 surface expression. Again, th ese CD4 mutants did not bind to gp120 at the cell surface, consistent with earlier reports. These results demonstrate that intracellular int eractions between gp160 and CD4 in the ER may utilize different contac t sites compared to those used during CD4 and gp120 binding at the cel l surface. The data provide further evidence that the environment in w hich CD4 and the HIV-1 envelope glycoprotein interact can have a signi ficant effect on their interaction. (C) 1996 Academic Press, Inc.