RhoB regulates cell migration through altered focal adhesion dynamics

Vega, Francisco; Colomba, Audrey; Reymond, Nicolas; Thomas, Mairian; Ridley, Anne J.

doi:10.1098/rsob.120076

Cited by 52 publications

(59 citation statements)

References 29 publications

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“…B30.2 domains have been reported to function as scaffold modules by spatiotemporally sequestering signaling proteins to distinct subcellular locations (Perfetto et al, 2013). This interaction favors an enhanced submembranous cytoskeleton arrangement, which leads to the observed immobilization of BTN3A1 at the cell surface, similar to its reported role in modulating membrane structures such as focal adhesions (Allal et al, 2002, Vega et al, 2012). Moreover, our data suggest a sequential, second cascade of events in the transmembrane compartment, which is necessary for recognition by a Vγ9Vδ2 TCR+ T cell: RhoB dissociates from immobilized BTN3A1 most likely after phosphoantigen binding to B30.2 and the conformational change in the extracellular domain of BTN3A1, which is then independent of RhoB GTPase activity.…”

Section: Discussionsupporting

confidence: 74%

RhoB Mediates Phosphoantigen Recognition by Vγ9Vδ2 T Cell Receptor

et al. 2016

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Summary Human Vγ9Vδ2 T cells respond to tumour cells by sensing elevated levels of phosphorylated intermediates of the dysregulated mevalonate pathway, which is translated into activating signals by the ubiquitously expressed butyrophilin A1 (BTN3A1) through yet unknown mechanisms. Here, we developed an unbiased, genome-wide screening method that identified RhoB as a critical mediator of Vγ9Vδ2 TCR activation in tumour cells. Our results show that Vγ9Vδ2 TCR activation is modulated by the GTPase activity of RhoB and its redistribution to BTN3A1. This is associated with cytoskeletal changes that directly stabilize BTN3A1 in the membrane, and the subsequent dissociation of RhoB from BTN3A1. Furthermore, phosphoantigen accumulation induces a conformational change in BTN3A1, rendering its extracellular domains recognizable by Vγ9Vδ2TCRs. These complementary events provide further evidence for inside-out signaling as an essential step in the recognition of tumor cells by a Vγ9Vδ2TCR.

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Section: Discussionsupporting

confidence: 74%

RhoB Mediates Phosphoantigen Recognition by Vγ9Vδ2 T Cell Receptor

et al. 2016

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“…The aberrant migration and focal adhesion turnover of PKP2 KD cells could therefore be due to changes in integrin expression (Vega et al, 2012). The expression of the β1 and β4 integrins was analyzed by immunofluorescence and immunoblot analyses, and an increased amount of both integrins was observed in PKP2 KD cells compared with the control keratinocytes (Figure 5a and b).…”

Section: Resultsmentioning

confidence: 99%

Plakophilin 2 Affects Cell Migration by Modulating Focal Adhesion Dynamics and Integrin Protein Expression

Koetsier

Amargo

Todorović

et al. 2014

Journal of Investigative Dermatology

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Plakophilin 2 (PKP2), a desmosome component, modulates the activity and localization of the small GTPase RhoA at sites of cell–cell contact. PKP2 regulates cortical actin rearrangement during junction formation, and its loss is accompanied by an increase in actin stress fibers. We hypothesized that PKP2 may regulate focal adhesion dynamics and cell migration. Here we show that PKP2-deficient cells bind efficiently to the extracellular matrix, but upon spreading display total cell areas ~30% smaller than control cells. Focal adhesions in PKP2-deficient cells are ~2× larger and more stable than in control cells, and vinculin displays an increased time for fluorescence recovery after photobleaching. Furthermore, β4 and β1 integrin protein and mRNA expression is elevated in PKP2-silenced cells. Normal focal adhesion phenotypes can be restored in PKP2-null cells by dampening the RhoA pathway or silencing β1 integrin. However, integrin expression levels are not restored by RhoA signaling inhibition. These data uncover a potential role for PKP2 upstream of β1 integrin and RhoA in integrating cell–cell and cell–substrate contact signaling in basal keratinocytes necessary for the morphogenesis, homeostasis, and reepithelialization of the stratified epidermis.

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“…Unbound cells were washed from the plates with PBS, and attached cells were fixed with 4% paraformaldehyde for 15 min at room temperature. The surface area of cells was determined in at least 50 cells per view in triplicate manually outlined using ImageJ software as described previously (25)(26)(27). For immunofluorescence assays, fixed cells were stained for actin and vinculin using the Actin Cytoskeleton/Focal Adhesion Staining kit (Millipore; FAK100).…”

Section: Methodsmentioning

confidence: 99%

Regulation of Receptor for Advanced Glycation End Products (RAGE) Ectodomain Shedding and Its Role in Cell Function

Braley

Kwak

Jules

et al. 2016

Journal of Biological Chemistry

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The receptor for advanced glycation end products (RAGE) is a multiligand transmembrane receptor that can undergo proteolysis at the cell surface to release a soluble ectodomain. Here we observed that ectodomain shedding of RAGE is critical for its role in regulating signaling and cellular function. Ectodomain shedding of both human and mouse RAGE was dependent on ADAM10 activity and induced with chemical activators of shedding (ionomycin, phorbol 12-myristate 13-acetate, and 4-aminophenylmercuric acetate) and endogenous stimuli (serum and RAGE ligands). Ectopic expression of the splice variant of RAGE (RAGE splice variant 4), which is resistant to ectodomain shedding, inhibited RAGE ligand dependent cell signaling, actin cytoskeleton reorganization, cell spreading, and cell migration. We found that blockade of RAGE ligand signaling with soluble RAGE or inhibitors of MAPK or PI3K blocked RAGE-dependent cell migration but did not affect RAGE splice variant 4 cell migration. We finally demonstrated that RAGE function is dependent on secretase activity as ADAM10 and ␥-secretase inhibitors blocked RAGE ligand-mediated cell migration. Together, our data suggest that proteolysis of RAGE is critical to mediate signaling and cell function and may therefore emerge as a novel therapeutic target for RAGE-dependent disease states.Receptor for advanced glycation end products (RAGE) 3 is a transmembrane, multiligand receptor expressed by most cells but is characterized by its up-regulation in a range of inflammatory disease states including diabetes, various cancers, and cardiovascular disease (1). RAGE activation through ligand binding transduces intracellular signaling and in turn induces cell migration, invasion, and adhesion (1). Work from multiple groups has demonstrated using rodent models that blocking RAGE signaling impairs the development of numerous pathologic states and therefore highlights RAGE as an attractive therapeutic target (2-5). The most widely used means for blocking RAGE signaling is with soluble RAGE (sRAGE), the recombinantly produced extracellular domain of RAGE. Soluble RAGE acts as a decoy receptor and therefore neutralizes RAGE ligands (2, 6). Endogenous sRAGE isoforms, which have been identified in human and mouse sera, are generated through two distinct biological mechanisms: the alternative splicing of the transmembrane region of RAGE leading to a secreted isoform (esRAGE/RAGEv1) and the cleavage of extracellular domain (ectodomain or ECD) at the cell surface by ectodomain shedding (1, 7-10). Ectodomain shedding is a tightly regulated process and is mediated by various metalloproteinases (11, 12). Shedding is essential for normal physiological function but can be deregulated in various pathological disorders where altered levels of metalloproteinases are found (11, 12). In fact, many transmembrane proteins are known to undergo ectodomain shedding including cell adhesion molecules, ligands for growth factor receptors, immunoglobulins, and various enzymes (10 -13). Ectodomain shedding of cell sur...

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RhoB regulates cell migration through altered focal adhesion dynamics

Cited by 52 publications

References 29 publications

RhoB Mediates Phosphoantigen Recognition by Vγ9Vδ2 T Cell Receptor

RhoB Mediates Phosphoantigen Recognition by Vγ9Vδ2 T Cell Receptor

Plakophilin 2 Affects Cell Migration by Modulating Focal Adhesion Dynamics and Integrin Protein Expression

Regulation of Receptor for Advanced Glycation End Products (RAGE) Ectodomain Shedding and Its Role in Cell Function

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