Latent TGF-beta binding protein LTBP-1 contains three potential extracellular matrix interacting domains

Latent TGF-beta binding proteins (LTBPs) are components of the extracellula r matrix (ECM). They belong to the fibrillin/LTBP-superfamily, and are high molecular weight glycoproteins characterized by EGF-like repeats and 8-Cys repeats. Most LTBPs associate with the small latent forms of TGF-beta. The ir roles include to facilitate the secretion of latent TGF-beta and to targ et it to the ECM. In order to identify new matrix-binding domains of LTBP-1 and to characterize their association with the extracellular matrix, we ha ve produced (in a mammalian expression system) partly overlapping recombina nt fragments of its shorter form, LTBP-1S, and analyzed the binding of the purified fusion proteins to extracellular matrices of cultured human dermal and lung fibroblasts. Recombinant fragments from three different regions o f the N- and C-termini showed affinity to the matrix, These interacting reg ions contain either the first (hybrid), second or fourth 8-Cys domains of t he LTBP-1S molecule. They bound independently to the matrix, Each of them h ad an ability to inhibit the association of native exogenous LTBP-1 with fi broblast extracellular matrix. The interactions of the LTBP-1 fragments wit h the extracellular matrix resisted treatment with sodium deosycholate, sug gesting strong, possibly covalent binding. The binding occurred in a time- and dose-dependent fashion. The N-terminal fragments bound more readily to the matrices. With all fragments the binding took place both,vith intact fi broblast matrices and with matrices isolated by sodium deoxycholate. When u sing CHO cell layers, which form sparse matrices, only the N-terminal fragm ent of LTBP-1 was efficiently incorporated. The association of the binding fragments with isolated matrices was enhanced by soluble, cell-derived fact ors. The current data suggest that LTBP-1 contains three different domains with an ability to associate with the extracellular matrix.