Proteolysis of CCN1 by Plasmin: Functional Implications

Abstract

Plasmin is shown to play a crucial role in many pathophysiologic processes primarily through its ability to degrade extracellular matrix (ECM) and/or mobilizing growth factors that are sequestered in the ECM. Cysteine-rich 61 (CCN1) is a matricellular protein of which expression is up-regulated in cancer and various vascular diseases. The present study was undertaken to investigate whether plasmin liberates CCN1 from the ECM and whether the released growth factor modulates endothelial cell migration. Treatment of breast carcinoma cells (MDA-MB-231) with plasmin released a truncated form of CCN1 (28 kDa) into the overlying medium. Experiments with recombinant CCN1 confirmed that plasmin effectively cleaves CCN1. Thrombin and other clotting/fibrinolytic proteases are ineffective in cleaving CCN1. Further studies revealed that the conditioned medium of plasmintreated carcinoma cells supports endothelial cell migration and that antibodies specific to CCN1 blocked this enhancing effect. These data were the first to show that plasmin can liberate a pluripotent matrix signaling protein, CCN1, from the ECM. Because both CCN1 and the components of the plasmin generation system are present in tumor cells and a variety of other cells, the proteolysis of CCN1 by plasmin may play a role in many pathophysiologic processes, including tumor cellmediated angiogenesis. (Cancer Res 2005; 65(21): 9705-11)

CCN1 secretion and cleavage regulate the lung epithelial cell functions after cigarette smoke

“…The majority of flCCN1 or cCCN1 was detected in exosome or the soluble component, respectively (Fig. 1H), suggesting a differential route of secretion for flCCN1 or cCCN1 after CSE stimulation despite that CCN1 cleavage has been reported in breast cancer tissue previously (14,37). CS-induced CCN1 cleavage in lung tissue and its functional roles in lung diseases remain unexplored.…”

Matricellular Signaling Molecule CCN1 Attenuates Experimental Autoimmune Myocarditis by Acting as a Novel Immune Cell Migration Modulator

“…For this purpose, 2 cDNAs expressing native full-length mCCN1or a variant thereof (mCCN1-HA) with a carboxy-terminal 9 -amino acid HA tag (the hemagglutinin epitope YPYDVPDYA from influenza virus A) were cloned into the hepatotropic adenovirus mutant RR5 to yield the vectors AdV-mCCN1 and AdV-mCCN1-HA, respectively ( Figure 1A). When AdV-mCCN1 or AdV-mCCN1-HA is expressed in EA.hy926 cells, a major fraction of CCN1 remains cell-bound, but there is free full-length CCN1 protein and an expected 21-kDa plasmin degradation product 28 in the cell supernatants ( Figure 1B). By intracellular flow cytometry, recombinant CCN1 protein could be detected both intracellularly and on the surface of EA.hy926 cells ( Figure 1C).…”

“…8 Because adenoviral gene transfer is highly liver-selective, with Ͼ99% of the vector located and expressed in this organ, the detection of HA-tagged protein on macrophages proves secretion of recombinant CCN1 protein and binding to immune cells. Furthermore, CCN1 protein could be detected by Western blot analysis in sera with 50-to 42-kDa bands corresponding to glycosylated full-length CCN1, an unglycosylated form, and a 21-kDa plasmin degradation fragment, 28 respectively ( Figure 1F). ELISA detected high CCN1-HA levels in heparin plasma from AdV-CCN1-HA-treated mice compared with RR5-treated or nontreated mice ( Figure 1E).…”

Intrinsic biological activity of the thrombospondin structural homology repeat in connective tissue growth factor

“…1A) that may function both independently and interdependently (Bork 1993, Brigstock 1999. Isoforms of CCN2 comprising essentially module 4 alone are bioactive and contain binding sites for the integrins v 3 and 5 1, which account for the ability of CCN2 to interact with stellate cells in the liver and pancreas, respectively (Gao & Brigstock 2004, 2005. Stellate cells are normally quiescent, but following injury they become activated and myofibroblastic, expressing high levels of -smooth muscle actin ( SMA).…”