Antithrombin Regulates Matriptase Activity Involved in Plasmin Generation, Syndecan Shedding, and HGF Activation in Keratinocytes

Ya-Wen Chen,Michael D Johnson,Hiroaki Kataoka,Chen-Yong Lin,Richard Swanson,Zhenghong Xu,Steve T Olson,Jehng-Kang Wang,Adrienne N H Baksh,Chiu-Yuan Chen,Wanjin Hong

doi:10.1371/journal.pone.0062826

Abstract

Matriptase, a membrane-associated serine protease, plays an essential role in epidermal barrier function through activation of the glycosylphosphatidylinositol (GPI)-anchored serine protease prostasin. The matriptase-prostasin proteolytic cascade is tightly regulated by hepatocyte growth factor activator inhibitor (HAI)-1 such that matriptase autoactivation and prostasin activation occur simultaneously and are followed immediately by the inhibition of both enzymes by HAI-1. However, the mechanisms whereby matriptase acts on extracellular substrates remain elusive. Here we report that some active matriptase can escape HAI-1 inhibition by being rapidly shed from the cell surface. In the pericellular environment, shed active matriptase is able to activate hepatocyte growth factor (HGF), accelerate plasminogen activation, and shed syndecan 1. The amount of active matriptase shed is inversely correlated with the amount of antithrombin (AT) bound to the surface of the keratinocytes. Binding of AT to the surface of keratinocytes is dependent on a functional heparin binding site, Lys-125, and that the N-glycosylation site Asn-135 be unglycosylated. This suggests that β-AT, and not α-AT, is responsible for regulation of pericellular matriptase activity in keratinocytes. Keratinocytes appear to rely on AT to regulate the level of pericellular active matriptase much more than breast and prostate epithelial cells in which AT regulation of matriptase activity occurs at much lower levels than keratinocytes. These results suggest that keratinocytes employ two distinct serine protease inhibitors to control the activation and processing of two different sets of matriptase substrates leading to different biological events: 1) HAI-1 for prostasin activation/inhibition, and 2) AT for the pericellular proteolysis involved in HGF activation, accelerating plasminogen activation, and shedding of syndecans.

Highlights

Epidermal differentiation is a carefully controlled process that generates a functional epidermal layer providing the critical barrier function of the skin [1,2]
These results suggest that keratinocytes employ two distinct serine protease inhibitors to control the activation and processing of two different sets of matriptase substrates leading to different biological events: 1) HAI-1 for prostasin activation/inhibition, and 2) AT for the pericellular proteolysis involved in HGF activation, accelerating plasminogen activation, and shedding of syndecans
Our results indicate that the two distinct matriptase inhibitors are apparently involved in the regulation of two different types of substrate processing by matriptase: HAI-1 for prostasin activation and AT for the activation of HGF, acceleration of plasminogen activation, and shedding of syndecan-1

Summary

Introduction

Epidermal differentiation is a carefully controlled process that generates a functional epidermal layer providing the critical barrier function of the skin [1,2]. Among the many proteases and protease inhibitors that are involved in skin functions, matriptase, prostasin, and HAI-1 have been shown to be functionally linked and form a tightly controlled protease/inhibitor network. Increased matriptase zymogen activation has been previously shown to be associated with various human skin diseases and may result from the oxidative environment associated with the inflammation, or acidification of the extracellular milieu associated with many pathologic states, since matriptase activation is induced in cells exposed to H2O2 or a mildly acidic environment [7,8]. A GPI-anchored serine protease appears to be the sole downstream substrate responsible for the epidermal defects associated with matriptase ablation in mice [9]. A remarkable feature of regulation of this serine protease cascade is that both proteases are under extremely tight control by HAI-1 [6]. The unusually tight linkage of the three key players of the protease network is consistent with the similar epidermal defects observed in their respective knockout mice [12,13,14]

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Conclusion