Nck enables directional cell migration through the coordination of polarized membrane protrusion with adhesion dynamics

Abstract

SummaryDirectional migration requires the coordination of cytoskeletal changes essential for cell polarization and adhesion turnover. Extracellular signals that alter tyrosine phosphorylation drive directional migration by inducing reorganization of the actin cytoskeleton. It is recognized that Nck is an important link between tyrosine phosphorylation and actin dynamics; however, the role of Nck in cytoskeletal remodeling during directional migration and the underlying molecular mechanisms remain largely undetermined. In this study, a combination of molecular genetics and quantitative live cell microscopy was used to show that Nck is essential in the establishment of front-back polarity and directional migration of endothelial cells. Time-lapse differential interference contrast and total internal reflection fluorescence microscopy showed that Nck couples the formation of polarized membrane protrusions with their stabilization through the assembly and maturation of cell-substratum adhesions. Measurements by atomic force microscopy showed that Nck also modulates integrin a5b1-fibronectin adhesion force and cell stiffness. Fluorescence resonance energy transfer imaging revealed that Nck depletion results in delocalized and increased activity of Cdc42 and Rac. By contrast, the activity of RhoA and myosin II phosphorylation were reduced by Nck knockdown. Thus, this study identifies Nck as a key coordinator of cytoskeletal changes that enable cell polarization and directional migration, which are crucial processes in development and disease.

Requisite Role for Nck Adaptors in Cardiovascular Development, Endothelial-to-Mesenchymal Transition, and Directed Cell Migration

“…These phenotypes are accompanied by profound defects in stress fiber formation, cell polarity, focal adhesion turnover, and regulation of FAK, RhoA, and Rac1. Our findings complement those of Rivera and colleagues, who employed short hairpin RNAmediated knockdown of Nck in human umbilical vein endothelial cells and live-cell microscopy to elegantly establish a mechanism by which Nck guides cell polarization and the stabilization of cellsubstrate adhesions to facilitate directional cell migration (54). We now extend this model in vivo to propose that in the context of angiogenic sprouting, Nck further modulates the function of endothelial tip cells and other ECs, allowing them to physically extend from the leading edge of nascent vessels and guide their movement in response to chemotactic gradients of growth factors (55).…”

Nephrin Tyrosine Phosphorylation Is Required to Stabilize and Restore Podocyte Foot Process Architecture

“…35,36 Of note, Nck can complex with PINCH-ILK, 37 and it plays a direct role in cell adhesion and spreading. 38,39 In the nephrin-Y3F mutant, which cannot bind Nck, integration of the slit diaphragm with the mechanotransduction machinery seems to be compromised but incompletely severed. As such, tethering of the slit diaphragm to the actin cytoskeleton via mechanisms other than phosphonephrin-Nck1/2 interactions could explain why congenital nephrosis does not manifest in these mice (as it does in Nck-deficient animals 9 ).…”

SDF1α‐induced interaction of the adapter proteins Nck and HS1 facilitates actin polymerization and migration in T cells

“…Recently, also Nck was implicated in the directional migration of endothelial cells, where it coordinates cytoskeletal changes associated with cell polarity [43,44]. Moreover, the HS1 homologue cortactin was shown to recruit Nck to invadopodia in breast carcinoma cells.…”

“…Interestingly, we observed a clear reduction of SDF1α-induced actin polymerization upon overexpression of both Y 378 F and Y 397 F HS1 variants indicating that both phosphorylation sites govern the regulation of the actin cytoskeleton upon CXCR4 ligation. Since the Nck SH2 domain selectively binds to phosphorylated Y 378, this implies that a multipartite molecular complex may be formed by HS1 to facilitate actin polymerization in response to CXCR4 ligation with its exact composition yet to be defined.Recently, also Nck was implicated in the directional migration of endothelial cells, where it coordinates cytoskeletal changes associated with cell polarity [43,44]. Moreover, the HS1 homologue cortactin was shown to recruit Nck to invadopodia in breast carcinoma cells.…”