Ligation of Integrin α3β1 by Laminin 5 at the Wound Edge Activates Rho-dependent Adhesion of Leading Keratinocytes on Collagen

Beth P. Nguyen,Xiang-Dong Ren,William G. Carter,Martin A. Schwartz

doi:10.1074/jbc.m103404200

Abstract

Wounding of the epidermis signals the transition of keratinocytes from quiescent anchorage on endogenous basement membrane laminin 5 to migration on exposed dermal collagen. In this study, we attempt to characterize activation signals that transform quiescent keratinocytes into migratory leading cells at the wound edge. Previously, we reported that adhesion and spreading on collagen via integrin alpha(2)beta(1) by cultured human foreskin keratinocytes (HFKs) requires RhoGTP, a regulator of actin stress fibers. In contrast, adhesion and spreading on laminin 5 requires integrins alpha(3)beta(1) and alpha(6)beta(4) and is dependent on phosphoinositide 3-hydroxykinase (Nguyen, B. P., Gil, S. G., and Carter, W. G. (2000) J. Biol. Chem. 275, 31896-31907). Here, we report that quiescent HFKs do not adhere to collagen but adhere and spread on laminin 5. By using collagen adhesion as one criterion for conversion to a "leading wound cell," we found that activation of collagen adhesion requires elevation of RhoGTP. Adhesion of quiescent HFKs to laminin 5 via integrin alpha(3)beta(1) and alpha(6)beta(4) is sufficient to increase levels of RhoGTP required for adhesion and spreading on collagen. Consistently, adhesion of quiescent HFKs to laminin 5, but not collagen, also promotes expression of the precursor form of laminin 5, a characteristic of leading keratinocytes in the epidermal outgrowth. We suggest that wounding of quiescent epidermis initiates adhesion and spreading of keratinocytes at the wound edge on endogenous basement membrane laminin 5 via alpha(3)beta(1) and alpha(6)beta(4) in a Rho-independent mechanism. Spreading on endogenous laminin 5 via alpha(3)beta(1) is necessary but not sufficient to elevate expression of precursor laminin 5 and RhoGTP, allowing for subsequent collagen adhesion via alpha(2)beta(1), all characteristics of leading keratinocytes in the epidermal outgrowth.

Highlights

Wounding of the epidermis signals the transition of keratinocytes from quiescent anchorage on endogenous basement membrane laminin 5 to migration on exposed dermal collagen
We suggest that adhesion and spreading of keratinocytes at the wound edge via integrins ␣6␤4 and ␣3␤1 on endogenous basement membrane (BM) laminin 5, respectively, are necessary to increase levels of RhoGTP, collagen adhesion, and expression of precursor laminin 5 that define the migratory keratinocytes in the epidermal outgrowth
Quiescent human foreskin keratinocytes (HFKs) were isolated fresh from neonatal foreskin and assayed for cell adhesion and spreading on exogenous laminin 5 or collagen

Summary

Introduction

Wounding of the epidermis signals the transition of keratinocytes from quiescent anchorage on endogenous basement membrane laminin 5 to migration on exposed dermal collagen. Adhesion of quiescent HFKs to laminin 5, but not collagen, promotes expression of the precursor form of laminin 5, a characteristic of leading keratinocytes in the epidermal outgrowth. Spreading on endogenous laminin 5 via ␣3␤1 is necessary but not sufficient to elevate expression of precursor laminin 5 and RhoGTP, allowing for subsequent collagen adhesion via ␣2␤1, all characteristics of leading keratinocytes in the epidermal outgrowth. We attempt to characterize the initial adhesion-dependent activation signals (B) that transform quiescent keratinocytes (A) at the wound margin into migratory leading keratinocytes in the epidermal outgrowth (C) [2, 4, 5]. ␣3␤1 Ligation to Laminin 5 Activates Rho-dependent Adhesion and spread on the processed form of laminin 5 in the BM before they up-regulate expression of precursor laminin 5 as a downstream response

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Conclusion