Identification of fibronectin binding sites in dermatopontin and their biological function.

BACKGROUND:Dermatopontin (DP) is a 22kDa acidic extracellular matrix (ECM) protein that plays a critical role in both ECM structure and wound healing. Previously, we demonstrated that DP interacts with fibronectin (Fn) and promotes formation of insoluble Fn fibrils that are called activated Fn (Kato et al., J Biol Chem 2011;286:14861-9). OBJECTIVE:Details of the interaction between DP and Fn are investigated to further examine the biological functions of DP. METHODS:Interactive sites between Fn and DP were examined by a solid-phase assay using Fn, DP, and their respective recombinant proteins and synthetic peptides. The effect of the DP peptides on insoluble Fn fibril formation was examined by both electrophoresis and electron microscopy. RESULTS:A binding site in DP for Fn was identified as the DP-4 (PHGQVVVAVRS) peptide, and the major binding site in Fn for DP was the 14th type III repeat (III14) domain. Further, the major DP binding site in the III14 domain was located around the B- and C-strands and their connecting loop region. A synthetic cyclic peptide mimicking the Fn loop structure enhanced DP binding activity. The DP-4 peptide induced Fn polymerization but the morphology was different from that of Fn fibrils formed by full length DP. The Fn fibrils with DP-4 enhanced integrin α5β1-mediated cell adhesion and spreading. CONCLUSION:Interactive sites between Fn and DP were identified. The DP-4 peptide activated Fn and enhanced cell adhesion activity. DP-4 has the potential to be used for therapeutic applications, such as a wound treatment.

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