Rhodocetin-αβ–induced Neuropilin-1–cMet Association Triggers Restructuring of Matrix Contacts in Endothelial Cells

Abstract

Objective-The snake venom component rhodocetin-αβ (RCαβ) stimulates endothelial cell motility in an α2β1 integrinindependent manner. We aimed to elucidate its cellular and molecular mechanisms. Methods and Results-We identified neuropilin-1 (Nrp1) as a novel target of RCαβ by protein-chemical methods. RCαβ and vascular endothelial growth factor (VEGF)-A avidly bind to Nrp1. Instead of acting as VEGF receptor 2 coreceptor, Nrp1 associates upon RCαβ treatment with cMet. Furthermore, cell-based ELISAs and kinase inhibitor studies showed that RCαβ induces phosphorylation of tyrosines 1234/1235 and thus activation of cMet. Consequently, paxillin is phosphorylated at Y31, which is redistributed from streak-like focal adhesions to spot-like focal contacts at the cell perimeter, along with α2β1 integrin, thereby regulating cell-matrix interactions. Cortactin is abundant in the cell perimeter, where it is involved in the branching of the cortical actin network of lamellipodia, whereas tensile force-bearing actin stress fibers radiating from focal adhesions disappear together with zyxin, a focal adhesion marker, on RCαβ treatment. Conclusion-Our data demonstrate that (1) Nrp1 is a novel target for venom components, such as RCαβ; (2) Nrp1 coupled to cMet regulates the type of cell-matrix interactions in a manner involving paxillin phosphorylation; and (3) altered cell-matrix interactions determine endothelial cell migration and cellular force management.(Arterioscler Thromb Vasc Biol. 2013;33:544-554.)

Hepatocyte Growth Factor Modulates MET Receptor Tyrosine Kinase and β-Catenin Functional Interactions to Enhance Synapse Formation

“…How MET receptor activation mediates these discrete cellular outcomes is only beginning to be addressed, with most focus on the diversity of downstream signaling pathways initiated following the activation of MET (Finsterwald and Martin, 2011; Eagleson et al, 2016). Evidence from cell lines, however, indicates that the repertoire of MET protein-interacting partners expressed by a cell also can modulate MET signaling to influence biological outcomes (Smyth and Brady, 2005; Wang et al, 2005; Zeng et al, 2006; Reshetnikova et al, 2007; DeAngelis et al, 2010; Bozkaya et al, 2012; Burghy et al, 2012; Lu et al, 2012; Niland et al, 2013). A recent coimmunoprecipitation (Co-IP)/mass spectrometry (MS) study identified the MET interactome with 72 proteins, including β-catenin, in isolated murine neocortical synaptosomes during the peak of synaptogenesis (Xie et al, 2016).…”

Signals of the Neuropilin-1–MET Axis and Cues of Mechanical Force Exertion Converge to Elicit Inflammatory Activation in Coherent Endothelial Cells

“…This stiffening explains the SIRS-like symptoms caused by some snake venoms (44) at the inter-and intracellular level, and it provides insight into the underlying signaling through the NRP1-MET axis and the resulting increased mechanotransduction within a confluent monolayer of tightly interlinked ECs. Within an endothelial monolayer, the molecular mode of action of RCab is based on its binding to NRP1 and, thus, the formation of a ternary complex with MET as signaling receptor tyrosine kinase (29). Because of increased phosphorylation of paxillin, this causes disassembly of focal adhesions, which are the major forcetransmitting adhesion contacts of ECs with their ECM (29).…”

“…In a previous work, we identified rhodocetin-ab (RCab) as the first nonenzymatic component of a snake venom that recognizes neuropilin-1 (NRP1) on ECs (29). NRP1 is a transmembrane glycoprotein expressed inter alia in ECs, neurons, and immune cells (30).…”

“…There, NRP1 functions as coreceptor of VEGF receptor family members and of the hepatocyte growth factor (HGF) receptor MET. RCab binds to NRP1 and induces the formation of a ternary complex with MET (29). In vitro, this leads, in ECs without adjacent cells, to a reorganization of the actin cytoskeleton and to restructuring of the cell matrixanchoring complexes from focal adhesions into focal contacts, thereby promoting cell migration (29).…”

“…RCab binds to NRP1 and induces the formation of a ternary complex with MET (29). In vitro, this leads, in ECs without adjacent cells, to a reorganization of the actin cytoskeleton and to restructuring of the cell matrixanchoring complexes from focal adhesions into focal contacts, thereby promoting cell migration (29). In addition, in an in vivo tumor model, RCab selectively disrupted blood vessels of the tumor but not of healthy tissues (31).…”