title: Multiple residues contribute to the inability of murine CCR-5 to function as a coreceptor for macrophage-tropic human immunodeficiency virus type 1 isolates.
abstract: Infection of CD4-positive cells by human immunodeficiency virus type 1 (HIV-1) requires functional interaction of the viral envelope protein with a coreceptor belonging to the chemokine receptor family of seven-membrane-spanning receptors. For the majority of macrophage-tropic HIV-1 isolates, the physiologically relevant coreceptor is the human CCR-5 (hCCR-5) receptor. Although the murine homolog of CCR-5 (mCCR-5) is unable to mediate HIV-1 infection, chimeric hCCR-5/mCCR-5 molecules containing single extracellular domains derived from hCCR-5 are effective coreceptors for certain macrophage-tropic HIV-1 isolates. Here, we have sought to identify residues in hCCR-5 critical for HIV-1 infection by substitution of mCCR-5-derived residues into the context of functional chimeric hCCR-5/mCCR-5 receptor molecules. Using this strategy, we demonstrate that residues 7, 13, and 15 in the first extracellular domain and residue 180 in the third extracellular domain of CCR-5 are important for HIV-1 envelope-mediated membrane fusion. Of interest, certain substitutions, for example, at residues 184 and 185 in the third extracellular domain, have no phenotype when introduced individually but strongly inhibit hCCR-5 coreceptor function when present together. We hypothesize that these changes, which do not preclude chemokine receptor function, may inhibit a conformational transition in hCCR-5 that contributes to HIV-1 infection. Finally, we report that substitution of glycine for valine at residue 5 in CCR-5 can significantly enhance the level of envelope-dependent cell fusion by expressing cells. The diversity of the mutant phenotypes observed in this mutational analysis, combined with their wide distribution across the extracellular regions of CCR-5, emphasizes the complexity of the interaction between HIV-1 envelope and coreceptor.
authors.affiliation: Department of Genetics, Duke University Medical Center, Durham, North Carolina 27710, USA.
authors.forename: T M
authors.initials: TM
authors.lastname: Ross
authors.forename: P D
authors.initials: PD
authors.lastname: Bieniasz
authors.forename: B R
authors.initials: BR
authors.lastname: Cullen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D002461:Cell Line, Transformed; D002462:Cell Membrane / Q000378:metabolism; D015870:Gene Expression; D015497:HIV-1 / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D008264:Macrophages / Q000821:virology; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019713:Receptors, CCR5 / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D013329:Structure-Activity Relationship
chemical_list: D019713:Receptors, CCR5; D011993:Recombinant Fusion Proteins