Receptor-mediated endocytosis of CC-chemokines.
Chemokines are chemotactic proteins which play a central role in immune and inflammatory responses. Chemokine receptors are members of the seven transmembrane G-protein coupled family and have recently been shown to be involved in the entry of human immunodeficiency virus (HIV) into target cells. To study chemokine endocytosis in detail we have used novel site-specific chemistry to make a fluorescently labeled CC-chemokine agonist (rhodamine-MIP-1alpha) and antagonist (NBD-RANTES). We have also generated a CHO cell line stably expressing a hemagglutinin-tagged version of the CC-chemokine receptor 1 (CCR1), and using these reagents we have examined the receptor-mediated endocytosis of CC-chemokines by confocal microscopy. Our studies reveal that the agonist was internalized and accumulated in transferrin receptor-positive endosomes whereas the antagonist failed to internalize. However, receptor-bound antagonist could be induced to internalize by co-administration of agonist. Analysis of receptor redistribution following chemokine addition confirmed that sequestration was induced by agonists but not by antagonists.
Solari
R
R
Cell Biology Unit, Glaxo Wellcome Medicines Research Centre, Gunnels Wood Road, Stevenage, Hertfordshire SG1 2NY, United Kingdom. RCES4402@ggr.co.uk
Offord
R E
RE
Remy
S
S
Aubry
J P
JP
Wells
T N
TN
Whitehorn
E
E
Oung
T
T
Proudfoot
A E
AE
D009327:4-Chloro-7-nitrobenzofurazan; D054407:Chemokine CCL4; D018946:Chemokine CCL5 / Q000378:metabolism; D018925:Chemokines / Q000378:metabolism*; D004705:Endocytosis; D005456:Fluorescent Dyes; D006678:HIV; D006801:Humans; D019402:Macrophage Inflammatory Proteins / Q000378:metabolism; D054401:Receptors, CCR8; D019707:Receptors, Chemokine; D018121:Receptors, Cytokine / Q000378:metabolism*; D011990:Receptors, Transferrin / Q000032:analysis; D012235:Rhodamines
C103330:CCR8 protein, human; D054407:Chemokine CCL4; D018946:Chemokine CCL5; D018925:Chemokines; D005456:Fluorescent Dyes; D019402:Macrophage Inflammatory Proteins; D054401:Receptors, CCR8; D019707:Receptors, Chemokine; D018121:Receptors, Cytokine; D011990:Receptors, Transferrin; D012235:Rhodamines; D009327:4-Chloro-7-nitrobenzofurazan