The N terminus of Kaposi's sarcoma-associated herpesvirus G protein-coupled receptor is necessary for high affinity chemokine binding but not for constitutive activity.
Kaposi's sarcoma-associated herpesvirus (KSHV) contains a gene encoding a G protein-coupled receptor (KSHV-GPCR) that is homologous to mammalian chemokine receptors. KSHV-GPCR signals constitutively (in an agonist-independent manner) via the phosphoinositide-inositol 1,4,5-trisphosphate pathway. Because it has been proposed that the N terminus (N-TERM) of other GPCRs may act as tethered agonists, we determined whether the N-TERM of KSHV-GPCR is necessary for constitutive signaling activity or ligand binding, or both. We show that replacement of the entire N-TERM of KSHV-GPCR with those of two other GPCRs, deletion of residues within the N-TERM, and disruption of a putative disulfide bond that may hold the N-TERM in close proximity to extracellular loop 3 do not affect constitutive signaling activity but decrease chemokine binding. There were differences in the effects of mutation of the N-TERM on binding of the chemokines growth-related oncogene alpha, which is an agonist, and interferon-gamma-inducible protein-10, which is an inverse agonist. The effects on chemokine binding were accompanied by changes in chemokine regulation of KSHV-GPCR signaling. We conclude that the N-TERM is not necessary for constitutive KSHV-GPCR signaling, i.e. the N-TERM is not a tethered agonist, but plays a crucial role in binding of chemokine ligands and of chemokine regulation of KSHV-GPCR signaling.
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Program in Molecular Medicine, Weill Graduate School of Medical Sciences, Division of Molecular Medicine, Department of Medicine, Weill Medical College of Cornell University, New York, New York 10021, USA.
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Gershengorn
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D000818:Animals; D019556:COS Cells; D054360:Chemokine CXCL1; D054357:Chemokine CXCL10; D018925:Chemokines / Q000378:metabolism*; D019743:Chemokines, CXC / Q000378:metabolism; D002630:Chemotactic Factors / Q000378:metabolism; D006133:Growth Substances / Q000378:metabolism; D019288:Herpesvirus 8, Human; D007295:Inositol Phosphates / Q000378:metabolism; D036341:Intercellular Signaling Peptides and Proteins; D008024:Ligands; D054367:Receptors, CXCR3; D019707:Receptors, Chemokine / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D017384:Sequence Deletion; D015398:Signal Transduction
C475829:CXCL1 protein, human; C515187:CXCR3 protein, human; D054360:Chemokine CXCL1; D054357:Chemokine CXCL10; D018925:Chemokines; D019743:Chemokines, CXC; D002630:Chemotactic Factors; C115448:G protein-coupled receptor, Human herpesvirus 8; D006133:Growth Substances; D007295:Inositol Phosphates; D036341:Intercellular Signaling Peptides and Proteins; D008024:Ligands; D054367:Receptors, CXCR3; D019707:Receptors, Chemokine; D011994:Recombinant Proteins