Force Engages Vinculin and Promotes Tumor Progression by Enhancing PI3K Activation of Phosphatidylinositol (3,4,5)-Triphosphate

Abstract

Extracellular matrix stiffness induces focal adhesion assembly to drive malignant transformation and tumor metastasis. Nevertheless, how force alters focal adhesions to promote tumor progression remains unclear. Here, we explored the role of the focal adhesion protein vinculin, a force-activated mechano-transducer, in mammary epithelial tissue transformation and invasion. We found that extracellular matrix stiffness stabilizes the assembly of a vinculin-talin-actin scaffolding complex that facilitates PI3-kinase mediated phosphatidylinositol (3,4,5)-triphosphate phosphorylation. Using defined two and three dimensional matrices, a mouse model of mammary tumorigenesis with vinculin mutants and a novel super resolution imaging approach, we established that ECM stiffness, per se, promotes the malignant progression of a mammary epithelium by activating and stabilizing vinculin and enhancing Akt signaling at focal adhesions. Our studies also revealed that vinculin strongly co-localizes with activated Akt at the invasive border of human breast tumors, where the ECM is stiffest and we detected elevated mechano-signaling. Thus, extracellular matrix stiffness could induce tumor progression by promoting the assembly of signaling scaffolds; a conclusion underscored by the significant association we observed between highly expressed focal adhesion plaque proteins and malignant transformation across multiple types of solid cancer.

Selective Role of Vinculin in Contractile Mechanisms of Endothelial Permeability

“…In contrast, tensile forces mediated by actomyosin are required for conformational changes and activation of native vinculin in living cells (32), which is a critical step in force-induced enhancement of cell adhesion to the substrate. The next experiments tested if vinculin linkage to both the FA complex and actin cytoskeleton is required for full activation of signaling, and cytoskeletal and permeability responses by pulmonary ECs to thrombin and IL-6.…”

Low-intensity pulsed ultrasound promotes cell motility through vinculin-controlled Rac1 GTPase activity

“…The conversion of extracellular forces to intracellular signalling events is termed mechanotransduction; focal adhesions (FAs), which link the intracellular actin cytoskeleton to the ECM via integrins, are implicated as mechanosensitive cellular organelles (Puklin-Faucher and Sheetz, 2009;Schwarz and Gardel, 2012), capable of sensing forces and generating signalling events in response (Goldmann, 2012). The FA protein vinculin has been shown to be particularly important for mechanotransduction (Atherton et al, 2016); the link between vinculin and actin is required for cell polarization and Rac1 activation in response to cyclic stretching (Carisey et al, 2013), and vinculin is involved in generating cellular responses to different ECM rigidities (Holle et al, 2013;Rubashkin et al, 2014;Yamashita et al, 2014).…”

Applied stretch initiates directional invasion via the action of Rap1 GTPase as a tension sensor

“…The formation of vinculin-containing focal adhesions in 3D matrices is important for tumor cell growth and invasion (Rubashkin et al, 2014). Because Rap-dependent focal adhesion formation in 3D collagen persisted for at least 5 days (Fig.…”

“…To rule out the possibility that the changes in integrin dynamics within pre-existing adhesions were simply due to activation of the integrin, we expressed a mutant form of the β1 integrin subunit (G429N) that constitutively forces the integrin heterodimer into an extended active form (Rubashkin et al, 2014). Within pre-existing adhesions, this mutant β1 integrin subunit exhibited the same fluorescence recovery after photobleaching as the wild-type integrin (Fig.…”