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 chemo kine receptors, KSHV-GPCR signals constitutively (in an agonist-independent manner) via the phosphoinositide-inositol 1,4,5-trisphosphate pathway. Bec ause it has been proposed that the N terminus (N-TERM) of other GPCRs may a ct as tethered agonists, we determined whether the N-TERM of KSHV-GPCR is n ecessary 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 o ther GPCRs, deletion of residues within the N-TERM, and disruption of a put ative disulfide bond that may hold the N-TERM in close proximity to extrace llular loop 3 do not affect constitutive signaling activity but decrease ch emokine binding. There were differences in the effects of mutation of the N -TERM on binding of the chemokines growth-related oncogene a, which is an a gonist, and interferon-gamma-inducible protein-10, which is an inverse agon ist. The effects on chemokine binding were accompanied by changes in chemok ine regulation of KSHV-GPCR signaling. We conclude that the N-TERM is not n ecessary for constitutive KSHV-GPCR signaling, i.e. the N-TERM is not a tet hered agonist, but plays a crucial role in binding of chemokine ligands and of chemokine regulation of KSHV-GPCR signaling.