DEFINING THE DOMAINS OF TYPE-I COLLAGEN INVOLVED IN HEPARIN-BINDING AND ENDOTHELIAL TUBE FORMATION

Cell surface heparan sulfate proteoglycan (HSPG) interactions with typ e I collagen may be a ubiquitous cell adhesion mechanism. However, the HSPG binding sites on type I collagen are unknown. Previously we mapp ed heparin binding to the vicinity of the type I collagen N terminus b y electron microscopy, The present study has identified type I collage n sequences used for heparin binding and endothelial cell-collagen int eractions. Using affinity coelectrophoresis, we found heparin to bind as follows: to type I collagen with high affinity (K-d approximate to 150 nM); triple-helical peptides (THPs) including the basic N-terminal sequence alpha 1(I)87-92, KGH-RGF, with intermediate affinities (K-d approximate to 2 mu M); and THPs including other collagenous sequences , or single-stranded sequences, negligibly (K-d >> 10 mu M). Thus, hep arin-type I collagen binding likely relies on an N-terminal basic trip le-helical domain represented once within each monomer, and at multipl e sites within fibrils. We next defined the features of type I collage n necessary for angiogenesis in a system in which type I collagen and heparin rapidly induce endothelial tube formation in vitro. When pepti des, denatured or monomeric type I collagen, or type V collagen was su bstituted for type I collagen, no tubes formed, However, when peptides and type I collagen were tested together, only the most heparin-avid THPs inhibited tube formation, likely by influencing cell interactions with collagen-heparin complexes. Thus, induction of endothelial tube morphogenesis by type I collagen mag depend upon its triple-helical an d fibrillar conformations and on the N terminal heparin-binding site i dentified here.