Lateral Mobility and Nanoscale Spatial Arrangement of Chemokine-activated α4ß1 Integrins on T Cells.

ABSTRACT

Chemokine stimulation of integrin α4ß1-dependent T lymphocyte adhesion is a key step during lymphocyte trafficking. A central question regarding α4ß1 function is how its lateral mobility and organization influence its affinity and avidity following cell stimulation with chemokines and/or ligands. Using single particle tracking and superresolution imaging approaches, we explored the lateral mobility and spatial arrangement of individual α4ß1integrins on T cells exposed to different activating stimuli. We show that CXCL12 stimulation leads to rapid and transient α4ß1activation, measured by induction of the activation epitope recognized by the HUTS-21 anti-ß1antibody and by increased talin-ß1 association. CXCL12-dependent α4ß1 activation directly correlated with restricted lateral diffusion and integrin immobilization. Moreover, co-stimulation by CXCL12 together with soluble VCAM-1 potentiated integrin immobilization with a 5-fold increase in immobile integrins compared with unstimulated conditions. Our data indicate that docking by talin of the chemokine-activated α4ß1 to the actin cytoskeleton favors integrin immobilization, which likely facilitates ligand interaction and increased adhesiveness. Superresolution imaging showed that the nanoscale organization of high-affinity α4ß1 remains unaffected following chemokine and/or ligand addition. Instead, newly activated α4ß1 integrins organize on the cell membrane as independent units without joining pre-established integrin sites to contribute to cluster formation. Altogether, our results provide a rationale to understand how the spatiotemporal organization of activated α4ß1 integrins regulates T lymphocyte adhesion.