Characterization of a neutrophil cell surface glycosaminoglycan that mediates binding of platelet factor 4.
Platelet factor 4 (PF-4) is a platelet-derived alpha-chemokine that binds to and activates human neutrophils to undergo specific functions like exocytosis or adhesion. PF-4 binding has been shown to be independent of interleukin-8 receptors and could be inhibited by soluble chondroitin sulfate type glycosaminoglycans or by pretreatment of cells with chondroitinase ABC. Here we present evidence that surface-expressed neutrophil glycosaminoglycans are of chondroitin sulfate type and that this species binds to the tetrameric form of PF-4. The glycosaminoglycans consist of a single type of chain with an average molecular mass of approximately 23 kDa and are composed of approximately 85-90% chondroitin 4-sulfate disaccharide units type CSA (-->4GlcAbeta1-->3GalNAc(4-O-sulfate)beta1-->) and of approximately 10-15% di-O-sulfated disaccharide units. A major part of these di-O-sulfated disaccharide units are CSE units (-->4GlcAbeta1-->3GalNAc(4,6-O-sulfate)beta1-->). Binding studies revealed that the interaction of chondroitin sulfate with PF-4 required at least 20 monosaccharide units for significant binding. The di-O-sulfated disaccharide units in neutrophil glycosaminoglycans clearly promoted the affinity to PF-4, which showed a Kd approximately 0.8 microM, as the affinities of bovine cartilage chondroitin sulfate A, porcine skin dermatan sulfate, or bovine cartilage chondroitin sulfate C, all consisting exclusively of monosulfated disaccharide units, were found to be 3-5-fold lower. Taken together, our data indicate that chondroitin sulfate chains function as physiologically relevant binding sites for PF-4 on neutrophils and that the affinity of these chains for PF-4 is controlled by their degree of sulfation.
Petersen
F
F
Department of Medical Biochemistry and Microbiology, Uppsala University, Biomedical Center, Box 575, S-75123 Uppsala, Sweden.
Brandt
E
E
Lindahl
U
U
Spillmann
D
D
D000818:Animals; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D002852:Chromatography, Ion Exchange; D002854:Chromatography, Paper; D006025:Glycosaminoglycans / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009504:Neutrophils / Q000378:metabolism*; D010978:Platelet Factor 4 / Q000378:metabolism*; D011485:Protein Binding
D006025:Glycosaminoglycans; D010978:Platelet Factor 4