Interaction of platelet-derived growth factor with thrombospondin 1

Key factors that mediate vascular smooth muscle cell proliferation and migration are platelet-derived growth factor (PDGF) and thrombospondin 1 (TSP1). We now report that PDGF_BB bound tightly and specifically to TSP1, that this interaction was markedly dependent on the disulphide bond arrangement in TSP1, and that binding of PDGF_BB to TSP1 did not preclude PDGF_BB from binding to its receptor on rat aortic vascular smooth-muscle cells. At physiological ionic strength and pH, PDGF_BB bound to Ca²⁺-depleted TSP1 with a dissociation constant of 11±2 nM and to Ca²⁺-replete TSP1 with a dissociation constant of 32±5 nM. Binding was specific, as both soluble TSP1 and unlabelled PDGF_BB competed for binding of iodinated PDGF_BB to immobilized TSP1, whereas other platelet α-granule proteins did not compete. The tertiary structure of TSP1 is regulated by intramolecular disulphide interchange; we found that catalysis of disulphide interchange in TSP1 by protein disulphide isomerase ablated the binding of PDGF_BB. The interaction of PDGF_BB with TSP1 was weakened by increasing salt concentration and essentially ablated at 0.65 ionic strength; it was inhibited by heparin with a half-maximal effect at 20 i.u./ml, implying that the binding was mediated largely by ionic interactions. An anti TSP1 monoclonal antibody decreased the binding of iodinated PDGF_BB to PDGF receptor on rat aortic vascular smooth-muscle cells by 37±2%, whereas platelet TSP1 non-competitively inhibited binding of iodinated PDGF_BB. Uncomplexed PDGF_BB bound to PDGF receptor with an affinity 5±2 times that of PDGF_BB–TSP1 complexes. These results suggest that TSP1 might assist in the targeting of PDGF to its receptor on vascular smooth-muscle cells.