title: Selectivity in the Use of Gi/o Proteins Is Determined by the DRF Motif in CXCR6 and Is Cell-Type Specific.
abstract: CXCR6, the receptor for CXCL16, is expressed on multiple cell types and can be a coreceptor for human immunodeficiency virus 1. Except for CXCR6, all human chemokine receptors contain the D(3.49)R(3.50)Y(3.51) sequence, and all but two contain A(3.53) at the cytoplasmic terminus of the third transmembrane helix (H3C), a region within class A G protein-coupled receptors that contacts G proteins. In CXCR6, H3C contains D(3.49)R(3.50)F(3.51)I(3.52)V(3.53) at positions 126-130. We investigated the importance and interdependence of the canonical D126 and the noncanonical F128 and V130 in CXCR6 by mutating D126 to Y, F128 to Y, and V130 to A singly and in combination. For comparison, we mutated the analogous positions D142, Y144, and A146 to Y, F, and V, respectively, in CCR6, a related receptor containing the canonical sequences. Mutants were analyzed in both human embryonic kidney 293T and Jurkat E6-1 cells. Our data show that for CXCR6 and/or CCR6, mutations in H3C can affect both receptor signaling and chemokine binding; noncanonical H3C sequences are functionally linked, with dual changes mitigating the effects of single mutations; mutations in H3C that compromise receptor activity show selective defects in the use of individual Gi/o proteins; and the effects of mutations in H3C on receptor function and selectivity in Gi/o protein use can be cell-type specific. Our findings indicate that the ability of CXCR6 to make promiscuous use of the available Gi/o proteins is exquisitely dependent on sequences within the H3C and suggest that the native sequence allows for preservation of this function across different cellular environments.
authors: Laboratory of Molecular Immunology (S.P.S., J.F.F., H.H.Z., J.L.R., C.S.S., F.L., J.M.F.) and Bioinformatics and Scientific IT Program, Office of Technology Information Systems, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland (D.E.H.); and Howard Hughes Medical Institute, National Institutes of Health Research Scholars Program, Bethesda, Maryland (C.S.S.).
authors: Satya P
authors: SP
authors: Singh
authors: Laboratory of Molecular Immunology (S.P.S., J.F.F., H.H.Z., J.L.R., C.S.S., F.L., J.M.F.) and Bioinformatics and Scientific IT Program, Office of Technology Information Systems, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland (D.E.H.); and Howard Hughes Medical Institute, National Institutes of Health Research Scholars Program, Bethesda, Maryland (C.S.S.).
authors: John F
authors: JF
authors: Foley
authors: Laboratory of Molecular Immunology (S.P.S., J.F.F., H.H.Z., J.L.R., C.S.S., F.L., J.M.F.) and Bioinformatics and Scientific IT Program, Office of Technology Information Systems, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland (D.E.H.); and Howard Hughes Medical Institute, National Institutes of Health Research Scholars Program, Bethesda, Maryland (C.S.S.).
authors: Hongwei H
authors: HH
authors: Zhang
authors: Laboratory of Molecular Immunology (S.P.S., J.F.F., H.H.Z., J.L.R., C.S.S., F.L., J.M.F.) and Bioinformatics and Scientific IT Program, Office of Technology Information Systems, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland (D.E.H.); and Howard Hughes Medical Institute, National Institutes of Health Research Scholars Program, Bethesda, Maryland (C.S.S.).
authors: Darrell E
authors: DE
authors: Hurt
authors: Laboratory of Molecular Immunology (S.P.S., J.F.F., H.H.Z., J.L.R., C.S.S., F.L., J.M.F.) and Bioinformatics and Scientific IT Program, Office of Technology Information Systems, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland (D.E.H.); and Howard Hughes Medical Institute, National Institutes of Health Research Scholars Program, Bethesda, Maryland (C.S.S.).
authors: Jennifer L
authors: JL
authors: Richards
authors: Laboratory of Molecular Immunology (S.P.S., J.F.F., H.H.Z., J.L.R., C.S.S., F.L., J.M.F.) and Bioinformatics and Scientific IT Program, Office of Technology Information Systems, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland (D.E.H.); and Howard Hughes Medical Institute, National Institutes of Health Research Scholars Program, Bethesda, Maryland (C.S.S.).
authors: Craig S
authors: CS
authors: Smith
authors: Laboratory of Molecular Immunology (S.P.S., J.F.F., H.H.Z., J.L.R., C.S.S., F.L., J.M.F.) and Bioinformatics and Scientific IT Program, Office of Technology Information Systems, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland (D.E.H.); and Howard Hughes Medical Institute, National Institutes of Health Research Scholars Program, Bethesda, Maryland (C.S.S.).
authors: Fang
authors: F
authors: Liao
authors: Laboratory of Molecular Immunology (S.P.S., J.F.F., H.H.Z., J.L.R., C.S.S., F.L., J.M.F.) and Bioinformatics and Scientific IT Program, Office of Technology Information Systems, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland (D.E.H.); and Howard Hughes Medical Institute, National Institutes of Health Research Scholars Program, Bethesda, Maryland (C.S.S.) jfarber@niaid.nih.gov.
authors: Joshua M
authors: JM
authors: Farber
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D002478:Cells, Cultured; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000737:chemistry / Q000502:physiology*; D057809:HEK293 Cells; D006801:Humans; D019169:Jurkat Cells; D008958:Models, Molecular; D016296:Mutagenesis; D000075744:Receptors, CXCR6; D019707:Receptors, Chemokine / Q000737:chemistry* / Q000502:physiology; D011991:Receptors, Virus / Q000737:chemistry* / Q000502:physiology; D013329:Structure-Activity Relationship
chemical_list: C106330:CXCR6 protein, human; D000075744:Receptors, CXCR6; D019707:Receptors, Chemokine; D011991:Receptors, Virus; D019206:GTP-Binding Protein alpha Subunits, Gi-Go