Tyrosine Kinase Activity of Discoidin Domain Receptor 1 Is Necessary for Smooth Muscle Cell Migration and Matrix Metalloproteinase Expression

Abstract

Smooth muscle cell (SMC) interactions with collagen mediate cell migration during the pathogenesis of atherosclerosis and restenosis. Discoidin domain receptors (DDRs) have been identified as novel collagen receptors. We used aortic SMCs from wild-type and DDR1؊/؊ mice to evaluate the function of the DDR1 in regulating migration. DDR1؊/؊ SMCs exhibited impaired attachment to and migration toward a type I collagen substrate. Matrix metalloproteinase-2 (MMP-2) and MMP-9 activities were concomitantly reduced in these cells. Transfection of a full-length cDNA for DDR1b rescued these deficits, whereas kinase-dead mutants of DDR1 restored attachment but not migration and MMP production. These results suggest that active DDR1 kinase is a central mediator of SMC migration. Interactions between smooth muscle cells (SMCs) and collagen are important in cell migration during the pathogenesis of atherosclerosis and restenosis. The synthesis of several collagens, including fibrillar type I and III and short-chain type VIII, is upregulated after injury. 1,2 These collagens act as chemotactic factors for SMCs, and migration through the extracellular matrix is facilitated when type VIII collagen triggers matrix metalloproteinase (MMP) synthesis. 3,4 The discoidin domain receptors (DDRs) are two novel collagen receptors distinguished by an extracellular domain homologous to discoidin-I in Dictyostelium discoideum and by a catalytic tyrosine kinase domain in the cytoplasmic tail. The activation of DDR2 by collagen results in increased production of MMP-1 5 and MMP-2, 6 suggesting an important role for this receptor in matrix degradation. Targeted deletion of the DDR1 gene in mice resulted in severe defects in placental implantation and mammary gland development, 7 whereas DDR2 Ϫ/Ϫ mice exhibit skeletal malformation and delayed wound healing, 8 suggesting potential roles in migration and matrix degradation for both receptors. DDR1 was expressed on migrating SMCs after balloon catheter injury of the rat carotid artery, and intimal thickening after arterial injury was reduced in the DDR1 Ϫ/Ϫ mouse. 9 In the present study, we have investigated the role of DDR1 in SMC migration by transfecting DDR1 Ϫ/Ϫ SMCs with full-length or kinase-mutated DDR1 constructs to determine whether kinase activity is required for migration and MMP production. Materials and MethodsSMCs were isolated from the aortas of DDR1 Ϫ/Ϫ mice and wild-type littermates (mixture of 129/Sv and ICR strains). Knockout cells were transfected with the full-length cDNA of DDR1b, with a truncated form of DDR1b that lacks the catalytic domain (K529*) or with a kinase-dead form of DDR1a containing a lysine to alanine mutation at position 618 in the catalytic domain (K618A). 10 SMC attachment was measured by toluidine blue staining; chemotaxis chamber migration assays and gelatin zymograms were performed using methods we have described. 3 To localize the gelatinolytic activity in SMC cultures, the cells were overlaid with FITC-labeled DQ gelatin that is intermolecularly quenched ...

Collagens in the progression and complications of atherosclerosis

“…25 Furthermore, Ddr1 -/-SMCs exhibited reduced proliferation, migration, and MMP activity in response to type I or type VIII collagen in vitro, 25 and overexpression of DDR1 rescued these deficits. 30 In agreement with this, overexpression of DDR1 or DDR2 in human SMCs increased MMP-1 expression and decreased procollagen α 1 (I) mRNA expression. 29 Recent studies from our laboratory have demonstrated that SMC responses to collagen mediated by DDR1 may depend upon the disease context and the experimental model used.…”

Expression and mutation analysis of the discoidin domain receptors 1 and 2 in non-small cell lung carcinoma

“…Furthermore we have shown that DDR1 is an independent favourable prognostic marker for early-stage NSCLC patients, and that mutations in DDR1 and DDR2 appear less frequently than previously reported. The collagen-binding RTKs, DDR1 and DDR2, have previously been linked to various human diseases including fibrosis (Alves et al, 1995;Mao et al, 2002;Lee et al, 2004;Avivi-Green et al, 2006), atherosclerosis (Hou et al, 2001;Hou et al, 2002;Ferri et al, 2004), and cancer (Johnson et al, 1993;Alves et al, 1995;Barker et al, 1995;Nemoto et al, 1997;Weiner et al, 2000;Dejmek et al, 2003;Ongusaha et al, 2003;Heinzelmann-Schwarz et al, 2004;Ram et al, 2006;Vogel et al, 2006). The mechanism by which DDRs may contribute to oncogenesis is as yet unknown.…”

Antitumor activity of 7RH, a discoidin domain receptor 1 inhibitor, alone or in combination with dasatinib exhibits antitumor effects in nasopharyngeal carcinoma cells

“…The two categories of DDRs exhibit different tissue-specific expression patterns: DDR1 is predominantly expressed in epithelial cells and DDR2 is predominantly expressed in mesenchymal cells (19,20). Previous studies have demonstrated that DDR1 and DDR2 exhibit vital functions in the regulation of fundamental cellular processes, including proliferation, survival, differentiation, adhesion and matrix remodeling (21,22). In addition, the dysregulation of DDR1 and DDR2 has been associated with a number of human diseases, including fibrotic disorders, atherosclerosis and cancer (5,23).…”