Abstract
P-selectin glycoprotein ligand-1 (PSGL-1), a sialomucin expressed on leukocytes, is a major ligand for P-selectin and mediates leukocyte rolling on the endothelium. Here we show that human PSGL-1 interacts with CCL27 (CTACK/ILC/ESkine), a skin-associated chemokine that attracts skin-homing T lymphocytes. A recombinant soluble form of PSGL-1 (rPSGL-Ig) preferentially bound CCL27 among several chemokines tested. This interaction was abrogated by arylsulfatase treatment of rPSGL-Ig, suggesting that sulfated tyrosines play a critical role. In contrast, removal of either N-glycans or O-glycans by glycosidase treatment of rPSGL-Ig did not affect the interaction. The binding of CCL27 to a recombinant PSGL-1 synthesized in the presence of a sulfation inhibitor was lower than that produced in normal medium. Moreover, mutation of the tyrosines at the amino terminus of PSGL-1 to phenylalanine abolished the binding, further supporting the role of sulfated tyrosines in the CCL27-PSGL-1 interaction. Functionally, rPSGL-Ig reduced the chemotaxis of L1.2 cells expressing CCR10, the receptor for CCL27. In addition, the expression of human PSGL-1 on CCR10-expressing L1.2 cells resulted in reduced chemotaxis to CCL27. These findings suggest a role for PSGL-1 in regulating chemokine-mediated responses, in addition to its role as a selectin ligand.
Highlights
Leukocyte migration from the blood to tissues is initiated by transient and reversible interactions that capture leukocytes from flowing blood and allow them to roll on the surface of endothelial cells under blood flow
P-selectin glycoprotein ligand-1 (PSGL-1) Binds CCL27—To investigate the interaction between chemokines and PSGL-1, we first examined the binding of rPSGL-Ig, a recombinant soluble form of PSGL-1, to chemokines immobilized on a nitrocellulose membrane. rPSGL-Ig consists of the 47 amino-terminal amino acids of mature human PSGL-1, fused to a mutated hinge region of human IgG1 that has reduced complement binding and Fc receptor binding [19]. rPSGL-Ig is produced in Chinese hamster ovary cells that have been engineered to express FucT-VII and C2GlcNAcT-I, so it carries the glycans required for binding to P-selectin
All three tyrosines positioned at the amino terminus of PSGL-1 are required for the CCL27 binding
Summary
Leukocyte migration from the blood to tissues is initiated by transient and reversible interactions that capture leukocytes from flowing blood and allow them to roll on the surface of endothelial cells under blood flow. The expression of human PSGL-1 on CCR10-expressing L1.2 cells resulted in reduced chemotaxis to CCL27. CCR5 is modified by O-glycosylation in the amino-terminal region, which, together with tyrosine sulfation, contributes to its high affinity binding to chemokines [31].
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