RESUMO
Several seven-transmembrane chemokine receptors are known to function as entry coreceptors for human immunodeficiency virus type 1. CCR5 and CXCR4 are the major coreceptors for non-syncytium-inducing (NSI) and syncytium-inducing (SI) viruses, respectively. During the natural course of infection, the emergence of variants with a phenotypic transition from NSI to SI and rapid disease progression is associated with expanded coreceptor usage to CXCR4. Characteristic amino acids at several positions in the hypervariable region 3 (V3) of gp120 have been linked to CXCR4 utilization. Previously, we reported that a highly conserved arginine residue of V3 played an important role in CCR5 utilization. In this study, the possible involvement of the same arginine residue in CXCR4 utilization was investigated. Amino acid substitutions introduced to this arginine on R5X4 viruses were found to have a significant effect on their utilization of CXCR4. These results, taken together with those reported previously, suggest that this highly conserved arginine may contribute to the functional convergence of chemokine coreceptor utilization by human immunodeficiency viruses and may represent a unique target for future antiviral design.
Assuntos
Proteína gp120 do Envelope de HIV/química , Proteína gp120 do Envelope de HIV/metabolismo , HIV-1/fisiologia , Fragmentos de Peptídeos/metabolismo , Receptores CXCR4/metabolismo , Sequência de Aminoácidos , Substituição de Aminoácidos , Arginina , Linhagem Celular , Regiões Determinantes de Complementaridade/química , Regiões Determinantes de Complementaridade/genética , Proteína gp120 do Envelope de HIV/genética , HIV-1/patogenicidade , Humanos , Dados de Sequência Molecular , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/genéticaRESUMO
Crystallographic characterization of a ternary complex containing a monomeric gp120 core, parts of CD4, and a mAb, revealed a region that bridges the inner and outer domains of gp120. In a related genetic study, several residues conserved among primate lentiviruses were found to play important roles in CC-chemokine receptor 5 (CCR5) coreceptor utilization, and all but one were mapped to the bridging domain. To reconcile this finding with previous reports that the hypervariable region 3 (V3) of gp120 plays an important role in chemokine coreceptor utilization, elucidating the roles of various V3 residues in this critical part of the HIV type 1 (HIV-1) life cycle is essential. Alanine-scanning mutagenesis was carried out to identify V3 residues critical for CCR5 utilization. Our findings demonstrated that several residues in V3 were critical to CCR5 utilization. Furthermore, these residues included not only those conserved across HIV-1 subtypes, but also those that varied among HIV-1 subtypes. Although the highly conserved V3 residues may represent unique targets for antiviral designs, the involvement of variable residues raises the possibility that antigenic variation in the coreceptor binding domain could further complicate HIV-1 vaccine design.
Assuntos
Proteína gp120 do Envelope de HIV/química , Proteína gp120 do Envelope de HIV/metabolismo , HIV-1/fisiologia , Receptores CCR5/fisiologia , Alanina , Sequência de Aminoácidos , Substituição de Aminoácidos , Sítios de Ligação , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/virologia , Linhagem Celular , Sequência Consenso , Variação Genética , Proteína gp120 do Envelope de HIV/genética , HIV-1/genética , Humanos , Fragmentos Fab das Imunoglobulinas/química , Modelos Moleculares , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Conformação Proteica , Receptores CCR5/química , Alinhamento de Sequência , Homologia de Sequência de AminoácidosRESUMO
The seven-transmembrane CCR5 was recently found to double as a coreceptor for a genetically diverse family of human and nonhuman primate lentiviruses. Paradoxically, the main region of the envelope protein believed to be involved in CCR5 utilization was mapped to hypervariable region 3, or V3, of the envelope glycoprotein gp120. In this study, we addressed the question of whether functional convergence in CCR5 utilization is mediated by certain V3 residues that are highly conserved among HIV type 1 (HIV-1), HIV type 2, and simian immunodeficiency virus. Site-directed mutagenesis carried out on three such V3 residues revealed that the Arg-298 of HIV-1 gp120 has an important role in CCR5 utilization. In contrast, no effect was observed for the other residues we tested. The inability of Arg-298 mutants to use CCR5 was not attributed to global alteration of gp120 conformation. Neither the expression, processing, and incorporation of mutant envelope proteins into virions, nor CD4 binding were significantly affected by the mutations. This interpretation is further supported by the finding that alanine substitutions of five residues immediately adjacent to the arginine residue had no effect on CCR5 utilization. Taken together, our data strongly suggests that the highly conserved Arg-298 residue identified in the V3 of HIV-1 has a significant role in CCR5 utilization, and may represent an unusually conserved target for future anti-viral designs.
