title: Interaction of platelet-derived growth factor with thrombospondin 1.
abstract: 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 PDGFBB bound tightly and specifically to TSP1, that this interaction was markedly dependent on the disulphide bond arrangement in TSP1, and that binding of PDGFBB to TSP1 did not preclude PDGFBB from binding to its receptor on rat aortic vascular smooth-muscle cells. At physiologic ionic strength and pH, PDGFBB 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 PDGFBB competed for binding of iodinated PDGFBB to immobilized TSP1, whereas other platelet alpha-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 PDGFBB. The interaction of PDGFBB 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 PDGFBB to PDGF receptor on rat aortic vascular smooth-muscle cells by 37 +/- 2%, whereas platelet TSP1 non-competitively inhibited binding of iodinated PDGFBB. Uncomplexed PDGFBB bound to PDGF receptor with an affinity 5 +/- 2 times that of PDGFBB-TSP1 complexes. These results suggest that TSP1 might assist in the targeting of PDGF to its receptor on vascular smooth-muscle cells.
authors.affiliation: Centre for Thrombosis and Vascular Research, School of Pathology, University of New South Wales, Sydney, NSW 2052, Australia.
authors.forename: P J
authors.initials: PJ
authors.lastname: Hogg
authors.forename: K A
authors.initials: KA
authors.lastname: Hotchkiss
authors.forename: B M
authors.initials: BM
authors.lastname: Jiménez
authors.forename: P
authors.initials: P
authors.lastname: Stathakis
authors.forename: C N
authors.initials: CN
authors.lastname: Chesterman
mesh_terms: D000818:Animals; D006801:Humans; D008562:Membrane Glycoproteins / Q000378:metabolism*; D010982:Platelet-Derived Growth Factor / Q000378:metabolism*; D011485:Protein Binding; D011869:Radioligand Assay; D051381:Rats; D017479:Receptors, Platelet-Derived Growth Factor / Q000378:metabolism; D019699:Thrombospondins
chemical_list: D008562:Membrane Glycoproteins; D010982:Platelet-Derived Growth Factor; D019699:Thrombospondins; D017479:Receptors, Platelet-Derived Growth Factor