Biological Activation of pro-HGF (Hepatocyte Growth Factor) by Urokinase Is Controlled by a Stoichiometric Reaction

doi:10.1074/jbc.270.2.603

Biological Activation of pro-HGF (Hepatocyte Growth Factor) by Urokinase Is Controlled by a Stoichiometric Reaction (*)

From the Department of Biomedical Sciences and Oncology, University of Torino Medical School, corso Massimo D'Azeglio 52, 10126, Torino, Italy

^§ To whom correspondence should be addressed:
Dept. of Biomedical Sciences and Oncology, University of Torino Medical School, corso Massimo D'Azeglio 52, 10126, Torino, Italy.
Tel.: 39-11-6707739; Fax: 39-11-6509105.

Abstract

Hepatocyte growth factor (HGF) is a paracrine inducer of morphogenesis and invasive growth in epithelial and endothelial cells. HGF is secreted by mesenchymal cells as an inactive precursor (pro-HGF). The crucial step for HGF activation is the extracellular hydrolysis of the Arg-Val bond, which converts pro-HGF into αβ-HGF, the high-affinity ligand for the Met receptor. We previously reported that the urokinase-type plasminogen activator (uPA) activates pro-HGF in vitro. We now show that this is a stoichiometric reaction, and provide evidence for its occurrence in tissue culture.

Activation involves the formation of a stable complex between pro-HGF and uPA. This complex was isolated from the in vitro reaction of pure uPA with recombinant pro-HGF, as well as from the membrane of target cells, after sequential addition of uPA and pro-HGF. On the cell membrane, the uPA•HGF complex was bound to the Met receptor.

Monocytic cell lines, and primary monocytes after adhesion, activated efficiently pro-HGF both on their surface and in the culture medium. This activation was inhibited by anti-catalytic anti-uPA antibodies, and occurred by a stoichiometric reaction.

The stoichiometry of the activation reaction suggests that the biological effects of HGF can be titrated in vivo by the level of uPA activity. Adequate amounts of uPA can be locally provided by the macrophages, which would condition the tissue microenvironment by rendering HGF bioavailable to its target cells.

Footnotes

↵^¶ Supported by the Ministero della Sanita' (Progetto AIDS).
↵* This work was supported in part by grants from the Associazione Italiana Ricerche sul Cancro and the Consiglio Nazionale delle Ricerche (special project ACRO, Grant 92.02169.PF39). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

HGF

hepatocyte growth factor

uPA

urokinase plasminogen activator

PN-1

protease nexin-1

PAGE

polyacrylamide gel electrophoresis

CHAPS

3-[(3-cholamidopropyl-dimethylammonio]-1-propanesulfonic acid

DST

disuccinimidyl tartrate

PBS

phosphate-buffered saline.
↵²F. Galimi, M. F. Brizzi, E. Cottone, S. Giordano, L. Naldini, E. Vigna, C. Boccaccio, L. Pegoraro, and P. M. Comoglio, manuscript in preparation.
- Received August 19, 1994.
- Revision received October 20, 1994.