Macrophage Stimulating Protein (MSP) Binds to Its Receptor via the MSP β Chain

Abstract

Macrophage stimulating protein (MSP)1 was originally purified from human plasma, based on its activity for murine resident peritoneal macrophages (1). It is a 78-kDa heterodimeric protein composed of a disulfide-linked 53-kDa ␣ chain and a 25-kDa ␤ chain (calculated from amino acid composition). The ␣ chain contains a N-terminal hairpin loop followed by four kringle domains. The ␤ chain has a serine protease-like domain but is devoid of enzymatic activity due to amino acid substitutions in the catalytic triad. MSP belongs to the kringle protein family that includes plasminogen (2) and hepatocyte growth factor/scatter factor (HGF/SF) (3, 4). MSP is synthesized mainly by liver cells (5, 6), circulates in blood as a biologically inactive single chain precursor (7), and is cleaved by members of the kallikrein family (8, 9) or by trypsin-like enzymes located on macrophage surfaces (7). Recent functional studies have revealed that in addition to induction of macrophage shape change, chemotactic migration (10), and phagocytosis of C3bi-coated erythrocytes (1), MSP has other activities. These include inhibition of expression of inducible nitric oxide synthase mRNA in endotoxin or cytokine-stimulated macrophages (11), induction of interleukin-6 production and differentiation of megakaryocytes (12), suppression of colony formation of human bone marrow cells induced by Steel factor plus granulocyte macrophage-stimulating factor (13), increase in beat frequency of nasal epithelium cilia (14), and stimulation in vitro of proliferation of certain epithelial cell lines (15-17).The receptor for MSP was recently identified as the human RON gene product (18), a transmembrane receptor proteintyrosine kinase cloned from a human keratinocyte cDNA library (19). The murine STK gene cloned from hematopoietic stem cells of bone marrow is the homologue of human RON (20,21). The RON gene encodes a 190-kDa heterodimeric protein composed of a 40-kDa extracellular ␣ chain and 150-kDa transmembrane ␤ chain with intrinsic tyrosine kinase activity (21). This property places the product of the RON/STK gene into a subfamily of receptor tyrosine kinases that includes proto-oncogene MET and SEA (22,23). These receptors share many unique structural properties including a putative proteolytic cleavage site, similar location of cysteine residues in their extracellular domain, and two conserved tyrosines in the Cterminal tail (19,20,22,23). Studies of the signaling pathways of RON have shown that tyrosine-phosphorylated RON associates in vivo with intracellular signal transducers, including Grb-2-Sos and phosphatidylinositol 3-kinase (17, 24).In this work, we initiated a structure-activity study of MSP to identify functionally important domains that interact with the RON receptor. Five purified recombinant proteins were used, including pro-MSP, MSP, MSP ␣ and ␤ chains, and the MSP N terminus (including the first two kringles) fused to human IgG Fc. We report the binding capacity of MSP and its subunits to RON receptor in intact cells. We also analyzed ...

Coordinate expression and functional profiling identify an extracellular proteolytic signaling pathway

“…Macrophages respond to MSP-1 via RON activation, leading to changes in cell shape and increased migration, as well as an inhibition of nitric oxide production (23)(24)(25). We found that primary mouse peritoneal macrophages, but not primary mouse bone marrow macrophages, underwent a characteristic shape change upon activation by MSP-1 ( Fig.…”

“…We found that primary mouse peritoneal macrophages, but not primary mouse bone marrow macrophages, underwent a characteristic shape change upon activation by MSP-1 ( Fig. 3) (25). The effects of the endogenous MT-SP1 inhibitor, HAI-1, and a highly specific MT-SP1 antibody inhibitor (Fig.…”

Cross-talk between the proto-oncogenes Met and Ron

“…The response to HGF and MSP through the activation of Met and Ron is mediated by the same pathways, mainly by Ras and by PI-3 Kinase (Wang et al, 1996;Bardelli et al, 1999). Met can regulate growth and transformation by activating the MAP kinase cascade through Grb2/Sos/Ras complex whereas the Ron pathway is directed more towards cellular movement and matrix invasion (Santoro et al, 1996;Wang et al, 1997). When Met and Ron receptors are co-expressed, the formation of heterodimers may balance the activation of Ras and PI-3 kinase pathways.…”

Structural Basis for the Binding Specificity of Human Recepteur d'Origine Nantais (RON) Receptor Tyrosine Kinase to Macrophage-stimulating Protein

“…1B), and both ligands belong to the plasminogen-like growth factor family (31). Circulating MSP is synthesized in liver cells as a single-chain precursor (pro-MSP) that does not bind to RON (32,33). Under cellular stress, pro-MSP undergoes proteolytic maturation to become a disulfide-linked MSP␣␤, which binds and activates RON (33).…”

“…Circulating MSP is synthesized in liver cells as a single-chain precursor (pro-MSP) that does not bind to RON (32,33). Under cellular stress, pro-MSP undergoes proteolytic maturation to become a disulfide-linked MSP␣␤, which binds and activates RON (33). Several serine proteases (kallikreins, matriptase, hepsin, and human airway trypsin-like protease) recognize the cleavage site (Arg 483 -Val 484 ) between the K 4 and SP domains (33).…”