Interferon gamma binds to extracellular matrix chondroitin-sulfate proteoglycans, thus enhancing its cellular response.
The amino acid sequence of interferon gamma (IFN-gamma) has basic amino acid clusters similar to the heparin-binding consensus sequences found in other proteins that bind to proteoglycans (PGs). We investigated whether recombinant human IFN-gamma could bind to extracellular matrix (ECM) PGs secreted by human arterial smooth muscle cells (HASMCs) in vitro and whether the interaction affected the cellular response to IFN-gamma. As an in vitro model of ECM we used the basement membrane from HASMCs in culture. The binding of 125I-IFN-gamma to ECM was reduced significantly by pretreatment of ECM with chondroitinase ABC, an enzyme that degrades chondroitin-sulfate glycosaminoglycans. IFN-gamma binding to ECM was reduced by increasing concentrations of chondroitin-6-sulfate. 125I-IFN-gamma (0.05 to 2 ng/mL) binding data indicated an apparent Kd of 2 x 10(-11) mol/L and a maximum binding of 1.6 x 10(6) IFN-gamma molecules bound per square millimeter of ECM. Experiments with synthetic peptides suggested that residues 127 through 135 (AKTGKRKRS) are involved in the binding. The binding to chondroitin-sulfate PGs was confirmed by affinity chromatography of isolated [35S]chondroitin-sulfate PGs from ECM and cell-culture medium on immobilized IFN-gamma. The binding was abolished by treatment with chondroitinase ABC. ECM-bound IFN-gamma was more effective in inducing the expression of class II major histocompatibility antigens such as HLA-DR in HASMCs and human arterial endothelial cells than soluble IFN-gamma. These results suggest a role for chondroitin-sulfate PGs in immobilizing IFN-gamma in the ECM compartment and enhancing the cellular response to IFN-gamma.
Camejo
E H
EH
Wallenberg Laboratory for Cardiovascular Research, University of Gothenburg, Sahlgrenska University Hospital, Sweden.
Rosengren
B
B
Camejo
G
G
Sartipy
P
P
Fager
G
G
Bondjers
G
G
D000595:Amino Acid Sequence; D001665:Binding Sites; D001667:Binding, Competitive; D002478:Cells, Cultured; D011508:Chondroitin Sulfate Proteoglycans / Q000378:metabolism*; D002809:Chondroitin Sulfates / Q000494:pharmacology; D002810:Chondroitinases and Chondroitin Lyases / Q000494:pharmacology; D002846:Chromatography, Affinity; D004730:Endothelium, Vascular / Q000378:metabolism; D005109:Extracellular Matrix / Q000378:metabolism*; D005260:Female; D006025:Glycosaminoglycans / Q000032:analysis / Q000378:metabolism; D006684:HLA-DR Antigens / Q000032:analysis; D019763:Heparin Lyase; D006801:Humans; D007371:Interferon-gamma / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000378:metabolism; D011133:Polysaccharide-Lyases / Q000494:pharmacology; D011994:Recombinant Proteins; D014599:Uterus / Q000098:blood supply
D011508:Chondroitin Sulfate Proteoglycans; D006025:Glycosaminoglycans; D006684:HLA-DR Antigens; D011994:Recombinant Proteins; D007371:Interferon-gamma; D002809:Chondroitin Sulfates; D002810:Chondroitinases and Chondroitin Lyases; D011133:Polysaccharide-Lyases; D019763:Heparin Lyase