α₄β₁-Integrin regulates directionally persistent cell migration in response to shear flow stimulation

α₄β₁-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 α₄- integrin/paxillin binding by phosphorylation at the α₄-integrin cytoplasmic tail. Here, we demonstrate that α₄β ₁-integrin regulates directionally persistent cell migration via a more complex mechanism in which α₄-integrin phosphorylation and paxillin binding act via both cooperative and independent pathways. We show that, in response to shear flow, α₄β₁-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 α ₄β₁-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 α₄-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 α₄-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.