Genetic perturbation of the putative cytoplasmic membrane-proximal salt bridge aberrantly activates α₄ integrins

α ₄ integrins play a pivotal role in leukocyte migration and tissue-specific homing. The ability of integrins to bind ligand is dynamically regulated by activation-dependent conformational changes triggered in the cytoplasmic domain. An NMR solution structure defined a putative membrane-proximal salt bridge between the α _llbβ ₃ integrin cytoplasmic tails, which restrains integrins in their low-affinity state. However, the physiological importance of this salt bridge in α ₄ integrin regulation remains to be elucidated. To address this question, we disrupted the salt bridge in murine germ line by mutating the conserved cytoplasmic arginine R ^GFFKR in α ₄ integrins. In lymphocytes from knock-in mice (α ₄-R/A ^GFFKR), α ₄β ₁ and α ₄β ₇ integrins exhibited constitu-tively up-regulated ligand binding. However, transmigration of these cells across VCAM-1 and MAdCAM-1 substrates, or across endothelial monolayers, was reduced. Perturbed detachment of the tail appeared to cause the reduced cell migration of α ₄-R/A ^GFFKR lymphocytes. In vivo, α ₄-R/A ^GGGKR cells exhibited increased firm adhesion to Peyer patch venules but reduced homing to the gut. Our results demonstrate that the membrane-proximal salt bridge plays a critical role in supporting proper α ₄ integrin adhesive dynamics. Loss of this interaction destabilizes the nonadhesive conformation, and thereby perturbs the properly balanced cycles of adhesion and deadhesion required for efficient cell migration.