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Alpha4 beta1-Integrin regulates directionally persistent cell migration in response to shear flow stimulation.

ABSTRACT: alpha(4)beta(1)-Integrin plays a pivotal role in cell migration in vivo. This integrin has been shown to regulate the front-back polarity of migrating cells via localized inhibition of alpha(4)-integrin/paxillin binding by phosphorylation at the alpha(4)-integrin cytoplasmic tail. Here, we demonstrate that alpha(4)beta(1)-integrin regulates directionally persistent cell migration via a more complex mechanism in which alpha(4)-integrin phosphorylation and paxillin binding act via both cooperative and independent pathways. We show that, in response to shear flow, alpha(4)beta(1)-integrin binding to the CS-1 region of fibronectin was necessary and sufficient to promote directionally persistent cell migration when this integrin was ectopically expressed in CHO cells. Under shear flow, the alpha(4)beta(1)-integrin-expressing cells formed a fan shape with broad lamellipodia at the front and retracted trailing edges at the back. This "fanning" activity was enhanced by disrupting paxillin binding alone and inhibited by disrupting phosphorylation alone or together with disrupting paxillin binding. Notably, the phosphorylation-disrupting mutation and the double mutation resulted in the formation of long trailing tails, suggesting that alpha(4)-integrin phosphorylation is required for trailing edge retraction/detachment independent of paxillin binding. Furthermore, the stable polarity and directional persistence of shear flow-stimulated cells were perturbed by the double mutation but not the single mutations alone, indicating that paxillin binding and alpha(4)-integrin phosphorylation can facilitate directionally persistent cell migration in an independent and compensatory manner. These findings provide a new insight into the mechanism by which integrins regulate directionally persistent cell migration.

SUBMITTER: Dikeman DA

PROVIDER: S-EPMC2493558 | biostudies-literature | 2008 Jul

REPOSITORIES: biostudies-literature

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alpha4 beta1-Integrin regulates directionally persistent cell migration in response to shear flow stimulation.

Dikeman Dustin A DA Rivera Rosado Leslie A LA Horn Troy A TA Alves Christina S CS Konstantopoulos Konstantinos K Yang Joy T JT

American journal of physiology. Cell physiology 20080521 1

alpha(4)beta(1)-Integrin plays a pivotal role in cell migration in vivo. This integrin has been shown to regulate the front-back polarity of migrating cells via localized inhibition of alpha(4)-integrin/paxillin binding by phosphorylation at the alpha(4)-integrin cytoplasmic tail. Here, we demonstrate that alpha(4)beta(1)-integrin regulates directionally persistent cell migration via a more complex mechanism in which alpha(4)-integrin phosphorylation and paxillin binding act via both cooperative ...[more]

PMID: 18495811

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Project description:BackgroundIntegrin-mediated cell adhesion and spreading is dramatically enhanced by activation of the small GTPase, R-Ras. Moreover, R-Ras localizes to the leading edge of migrating cells, and regulates membrane protrusion. The exact mechanisms by which R-Ras regulates integrin function are not fully known. Nor is much known about the spatiotemporal relationship between these two molecules, an understanding of which may provide insight into R-Ras regulation of integrins.ResultsGFP-R-Ras localized to the plasma membrane, most specifically in membrane ruffles, in Cos-7 cells. GFP-R-Ras was endocytosed from these ruffles, and trafficked via multiple pathways, one of which involved large, acidic vesicles that were positive for Rab11. Cells transfected with a dominant negative form of GFP-R-Ras did not form ruffles, had decreased cell spreading, and contained numerous, non-trafficking small vesicles. Conversely, cells transfected with the constitutively active form of GFP-R-Ras contained a greater number of ruffles and large vesicles compared to wild-type transfected cells. Ruffle formation was inhibited by knock-down of endogenous R-Ras with siRNA, suggesting that activated R-Ras is not just a component of, but also an architect of ruffle formation. Importantly, beta1-integrin co-localized with endogenous R-Ras in ruffles and endocytosed vesicles. Expression of dominant negative R-Ras or knock down of R-Ras by siRNA prevented integrin accumulation into ruffles, impaired endocytosis of beta1-integrin, and decreased beta1-integrin-mediated adhesion. Knock-down of R-Ras also perturbed the dynamics of another membrane-localized protein, GFP-VSVG, suggesting a more global role for R-Ras on membrane dynamics. However, while R-Ras co-internalized with integrins, it did not traffic with VSVG, which instead moved laterally out of ruffles within the plane of the membrane, suggesting multiple levels of regulation of and by R-Ras.ConclusionsOur results suggest that integrin function involves integrin trafficking via a cycle of membrane protrusion, ruffling, and endocytosis regulated by R-Ras, providing a novel mechanism by which integrins are linked to R-Ras through control of membrane dynamics.

Dataset Information

Alpha4 beta1-Integrin regulates directionally persistent cell migration in response to shear flow stimulation.

Publications

alpha4 beta1-Integrin regulates directionally persistent cell migration in response to shear flow stimulation.

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