Paxillin Binding to the α4 Integrin Subunit Stimulates LFA-1 (Integrin αLβ2)-Dependent T Cell Migration by Augmenting the Activation of Focal Adhesion Kinase/Proline-Rich Tyrosine Kinase-2

Abstract

Engagement of very late Ag-4 (integrin α4β1) by ligands such as VCAM-1 markedly stimulates leukocyte migration mediated by LFA-1 (integrin αLβ2). This form of integrin trans-regulation in T cells requires the binding of paxillin to the α4 integrin cytoplasmic domain. This conclusion is based on the abolition of trans-regulation in Jurkat T cells by an α4 mutation (α4(Y991A)) that disrupts paxillin binding. Furthermore, cellular expression of an α4-binding fragment of paxillin that blocks the α4-paxillin interaction, selectively blocked VCAM-1 stimulation of αLβ2-dependent cell migration. The α4-paxillin association mediates trans-regulation by enhancing the activation of tyrosine kinases, focal adhesion kinase (FAK) and/or proline-rich tyrosine kinase-2 (Pyk2), based on two lines of evidence. First, disruption of the paxillin-binding site in the α4 tail resulted in much less α4β1-mediated phosphorylation of Pyk2 and FAK. Second, transfection with cDNAs encoding C-terminal fragments of Pyk2 and FAK, which block the function of the intact kinases, blocked α4β1 stimulation of αLβ2-dependent migration. These results define a proximal protein-protein interaction of an integrin cytoplasmic domain required for trans-regulation between integrins, and establish that augmented activation of Pyk2 and/or FAK is an immediate signaling event required for the trans-regulation of integrin αLβ2 by α4β1.