Objective:Select C-C and C-X-C chemokines can suppress HIV infection. This is because their receptors are the gateways for HIV-1 entry, determinants of viral tropism and sensitivity. C-C chemokines are most effective against macrophage-tropic viruses, and C-X-C chemokines are most effective against T-tropic viruses. The epitopes on the chemokine molecule responsible for virus inhibition and for chemokines' specificities are not known. The objective of this study was to map the functional domains of prototypic antiviral chemokine, namely, RANTES (regulated-on-activation normal T-expressed and secreted).Study Design:Optimal folding of the chemokine molecule is thought to be important for its biologic activity. Anticipating that it will provide a native milieu for folding, we expressed recombinant RANTES molecules in an HIV-2-derived lentivirus mammalian expression system. We focused on the structural landmarks of RANTES to determine their role in its life and function.Results:We found that the flexible amino-terminal region of RANTES was not important for its structural integrity or antiviral activity, either positively or negatively. It was also not important for binding to the CCR5 receptor. Modification of all other domains was detrimental, implying a functional role. However, a more careful analysis revealed that these domains were crucial for controlling stability, transport, and secretion of the molecule. Although all recombinant clones contained signal sequence and were transcriptionally active, they presented three different phenotypes: normal synthesis and secretion, normal synthesis but blocked secretion, and presumed normal synthesis but rapid degradation. Structural considerations and preliminary experiments showing a lack of effect of proteasome inhibitors suggested that the signal recognition particle pathway of translocation and proteasomal pathway of destruction may not be the major determinant of the life of the chemokine.Conclusions:The amino-terminal domain of RANTES was not essential for its antiviral activity or for its binding to the CCR5 receptor. Although the 1-domain of the core and carboxyterminal domain may contribute to the antiviral activity of RANTES, they were more important for its intracellular life.Journal of Human Virology 1999;2:270-282 © Lippincott Williams & Wilkins, Inc.