Mechanism of human keratinocyte migration on fibronectin: Unique roles of RGD site and integrins

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Abstract
The migration of human keratinocytes over the wound bed plays an important role in the re‐epithelialization of cutaneous wounds. Fibronectin, a large glycoprotein matrix component that is abundant within cutaneous wound beds, promotes keratinocyte migration. However, the mechanisms by which keratinocytes migrate over fibronectin are unknown. In this study, we sought to identify specific sites within the fibronectin molecule that induce keratinocyte locomotion and to characterize the cell surface receptors involved. The data show that the domain within the fibronectin molecule that induces human keratinocyte migration is the 120 kD cell‐binding domain close to the carboxyl terminus. The 40 kD heparin‐binding domain near the carboxyl terminus and the 45 kD gelatin‐binding domain near the amino terminus did not promote keratinocyte migration. In addition, keratinocyte migration on both fibronectin and the 120 kD cell‐binding domain was completely inhibited by the presence of GRGDSP peptide, suggesting that keratinocyte migration on fibronectin is mediated by recognizing the RGD sequence located within the cell‐binding domain of fibronectin. Furthermore, keratinocytes were able to migrate directly on immobilized RGD substratum. Cell migration on fibronectin is mediated by the α5β1 integrin since antibodies blocking the α5 and the β1 subunits completely inhibited keratinocyte migration on fibronectin. In addition, we demonstrate that human keratinocytes express α5β1 integrin in culture by flow cytometry.