CC and CX3C chemokines differentially interact with the N terminus of the human cytomegalovirus-encoded US28 receptor.
Human cytomegalovirus (HCMV) is the causative agent of life-threatening systemic diseases in immunocompromised patients as well as a risk factor for vascular pathologies, like atherosclerosis, in immunocompetent individuals. HCMV encodes a G-protein-coupled receptor (GPCR), referred to as US28, that displays homology to the human chemokine receptor CCR1 and binds several chemokines of the CC family as well as the CX3C chemokine fractalkine with high affinity. Most importantly, following HCMV infection, US28 activates several intracellular pathways, either constitutively or in a chemokine-dependent manner. In this study, our goal was to understand the molecular interactions between chemokines and the HCMV-encoded US28 receptor. To achieve this goal, a double approach has been used, consisting in the analysis of both receptor and ligand mutants. This approach has led us to identify several amino acids located in the N terminus of US28 that differentially contribute to the high affinity binding of CC versus CX3C chemokines. Additionally, our results highlight the importance of secondary modifications occurring at US28, such as sulfation, for ligand recognition. Finally, the effects of chemokine dimerization and interaction with glycosaminoglycans (GAGs) on chemokine binding and activation of US28 were investigated as well using CCL4 as model ligand. In line with the two-state model describing chemokine/receptor interaction, we show that an aromatic residue in the N-loop region of CCL4 promotes tight binding to US28, whereas receptor activation depends on the presence of the N terminus of CCL4, as shown previously for CCR5.
Casarosa
Paola
P
Leiden/Amsterdam Center for Drug Research, Division of Medicinal Chemistry, Faculty of Chemistry, De Boelelaan 1083, 1081 HV Amsterdam, The Netherlands.
Waldhoer
Maria
M
LiWang
Patricia J
PJ
Vischer
Henry F
HF
Kledal
Thomas
T
Timmerman
Henk
H
Schwartz
Thue W
TW
Smit
Martine J
MJ
Leurs
Rob
R
D000595:Amino Acid Sequence; D024322:Amino Acids, Aromatic / Q000378:metabolism; D000818:Animals; D019556:COS Cells; D054407:Chemokine CCL4; D019742:Chemokines, CC / Q000378:metabolism*; D020523:Chemokines, CX3C / Q000378:metabolism*; D002522:Chlorocebus aethiops; D017124:Conserved Sequence; D015870:Gene Expression; D006801:Humans; D019402:Macrophage Inflammatory Proteins / Q000235:genetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000235:genetics; D019707:Receptors, Chemokine / Q000235:genetics / Q000378:metabolism*; D013455:Sulfur / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
D024322:Amino Acids, Aromatic; C066507:CCL4 protein, human; D054407:Chemokine CCL4; D019742:Chemokines, CC; D020523:Chemokines, CX3C; D019402:Macrophage Inflammatory Proteins; D011506:Proteins; D019707:Receptors, Chemokine; C411191:US28 receptor, Cytomegalovirus; D014764:Viral Proteins; D014443:Tyrosine; D013455:Sulfur