THROMBOSPONDIN BINDS AND ACTIVATES THE SMALL AND LARGE FORMS OF LATENT TRANSFORMING GROWTH-FACTOR-BETA IN A CHEMICALLY-DEFINED SYSTEM

Transforming growth factor-beta (TGF-beta) is a potent growth regulato ry protein normally secreted by cells in a latent form. Primary regula tion of TGF-beta activity occurs through factors which control the pro cessing of the latent to the biologically active molecule. Thrombospon din (TSP), a platelet alpha-granule and extracellular matrix protein, forms specific complexes with active TGF-beta in platelet releasate an d activates endogenous latent TGF-beta secreted by endothelial cells v ia a cell- and protease-independent mechanism. In order to better unde rstand TSP-mediated activation of cell-secreted latent TGF-beta, we ex amined the consequences of interactions of the large (platelet-derived ) and small (recombinant) forms of latent TGF-beta with TSP in a chemi cally defined system. Data from these studies show that interactions b etween TSP and both forms of latent TGF-beta result in the generation of biologically active TGF-beta as assayed by the ability of NRK-49F c ells to form colonies in soft agar, by the ability to compete for bind ing to TGF-beta receptors on endothelial cells, and by an enzyme linke d immunosorbent assay selective for the active form of TGF-beta. Activ ation of latent TGF-beta by TSP stripped of associated TGF-beta activi ty (sTSP) is time- and concentration-dependent, but temperature-indepe ndent. The mechanism whereby sTSP activates latent TGF-beta appears to involve the direct binding of sTSP to the latent molecule as shown by gel permeation chromatography. In addition, a polyclonal antibody spe cific for the amino-terminal region of the latency-associated peptide (amino acids 81-94) inhibits sTSP-mediated activation of latent TGF-be ta in both the chemically defined system and in endothelial cell condi tioned medium. These data and the observation that similar concentrati ons of sTSP activate latent TGF-beta in both the chemically defined sy stem and in the endothelial cell system indicate that there is a commo n mechanism by which TSP activates the small, large, and endothelial c ell-derived latent TGF-beta complexes. The ability of TSP to convert l atent TGF-beta to biologically active TGF-beta suggests that TSP is a major regulatory factor in the control of TGF-beta activity.