Migration‐stimulating factor displays HEXXH‐dependent catalytic activity important for promoting tumor cell migration

Abstract

Like most extracellular matrix (ECM) components, fibronectin (Fn) is proteolyzed generating specific activities. Fibronectin proteinase (Fn-proteinase) represents such a cryptic activity located in the gelatin-binding domain (GBD) of Fn and displays a zinc metalloproteinase activity. The migration-stimulating factor (MSF) is a truncated Fn isoform generated by alternative mRNA splicing and corresponds to the N-terminal part of Fn that comprises the GBD. We show that several human mammary epithelial cells express MSF and constitutively produce Fn-proteinase activity. Furthermore, recombinant MSF produced by HEK-293 and MCF-7 cells possesses a constitutive Fn-proteinase activity. Mutating the putative zinc-binding motif, HEXXH, of the protein abolishes its activity thereby demonstrating its specificity. Using PCR, we showed that MSF is barely expressed in normal breast tissues, whereas its expression is significantly increased in tumors. Furthermore, an association between MSF expression and invasive capacity is observed in various breast adenocarcinoma cell lines. Indeed, when stably transfected in non-invasive MCF-7 cells, MSF promotes cell migration in a mechanism mostly dependent on its Fn-proteinase activity. In summary, our study shows that: (i) MSF displays constitutive Fn-proteinase activity; (ii) MSF expression is induced in human breast cancer; and (iii) MSF confers pro-migratory activity that depends mostly on its Fn-proteinase activity. These results suggest that MSF may be involved in tumor progression. ' 2005 Wiley-Liss, Inc.Key words: fibronectin; alternative splicing; Fn-proteinase; migration-stimulating factor; cancer Fibronectin (Fn) is an adhesive glycoprotein, secreted as a dimer, present in body fluids and within extracellular matrices. Each monomer ( 250 kDa) is organized into proteolysis-resistant functional domains. These domains can bind fibrin, collagens, integrins and proteoglycans, accounting for the wide range of Fn functions (Fig. 1a). 1 In addition to these well-characterized properties of ''fulllength'' Fn, proteolytically generated Fn fragments show additional biological activities. For instance, the gelatin-binding domain (GBD) influences cell proliferation and migration, 2 and induces cartilage catabolism. 3 In addition, the GBD isolated after Fn proteolysis or produced in recombinant cells, referred to fibronectin proteinase (Fn-proteinase), 4 displays a zinc-dependent collagenase/gelatinase activity. [5][6][7] Fn-proteinase activity is inhibited by a specific phosphinic pseudo peptide. 5 This activity is involved in the degradation of cartilage induced by the GBD. 8 In addition, we showed recently that Fn-proteinase is activated by plasmin and therefore may play a role in tissue remodeling. 4 Zhao et al. 9 recently cloned and characterized a C-terminus truncated Fn isoform generated by alternative splicing in zebrafish. This isoform, called Fn2, is a 120-kDa monomeric protein with distinct properties compared to ''full-length '' Fn. 9 This raised the possibility that Fn fragm...

Mice lacking the matrix metalloproteinase-9 gene reduce renal interstitial fibrosis in obstructive nephropathy

“…There is growing evidence suggesting that the TBM is not only a static physical barrier that separates tubular epithelial and interstitial compartments but also a regulator of tubular epithelial cell phenotype and physiology in the kidney (28). For example, a report shows that alterations in TBM composition regulate tubular epithelial cell transdifferentiation in vitro (7,13,18). It is also reported that interstitial collagen type I tends to promote EMT, whereas the collagen type IV is likely to inhibit this phenotypic transition.…”

“…The TBM consists predominantly of collagen IV and laminin (7,29). It provides a structural foundation for the proper function of tubular epithelial cells.…”

Tumor-Conditioned Macrophages Secrete Migration-Stimulating Factor: A New Marker for M2-Polarization, Influencing Tumor Cell Motility

“…It is also expressed during wound healing and strongly overexpressed in a wide range of common human cancers (62,102). rMSF displays a number of potent bioactivities relevant to cancer development processes, including stimulation of cell migration, hyaluronan synthesis (103,104), proteolytic activity (105), and angiogenesis (62,102).…”

Angiogenesis and tumour progression: migration-stimulating factor as a novel target for clinical intervention

“…These cells show a bell-shaped dose response with half-maximal activity manifesting at 0.1-1.0 pg/ml, that is, femtomolar concentrations. MSF also stimulates the migration of non-epithelial tumour cells and displays other bioactivities relevant to tumour progression, such as the upregulation of hyaluronan (HA) synthesis, proteolysis, and induction of angiogenesis 20,21,[27][28][29] (unpublished observations). The angiogenic activity of MSF has been demonstrated in both in vitro and in vivo models.…”

“…It is important to note that none of these bioactivities are manifested by any full-length fibronectin isoform, apparently as a consequence of steric hindrance. 31,32 Houard et al 29 have identified a second motif (HEEGH) in MSF module I-8 that seems to mediate the motogenic response of a breast cancer cell line (MCF7) and is additionally required for manifestation of its proteinase activity. Our recent data indicate that the motogenic response of certain target cells may be mediated by both IGD and HEEGH, whereas other cells only respond to the IGD motif.…”