High-Affinity Bent β(2)-Integrin Molecules in Arresting Neutrophils Face Each Other through Binding to ICAMs In cis

Leukocyte adhesion requires β(2)-integrin activation. Resting integrins exist in a bent-closed conformation—i.e., not extended (E(−)) and not high affinity (H(−))—unable to bind ligand. Fully activated E(+)H(+) integrin binds intercellular adhesion molecules (ICAMs) expressed on the opposing cell in trans. E(−)H(−) transitions to E(+)H(+) through E(+)H(−) or through EH(+), which binds to ICAMs on the same cell in cis. Spatial patterning of activated integrins is thought to be required for effective arrest, but no high-resolution cell surface localization maps of activated integrins exist. Here, we developed Super-STORM by combining super-resolution microscopy with molecular modeling to precisely localize activated integrin molecules and identify the molecular patterns of activated integrins on primary human neutrophils. At the time of neutrophil arrest, E(−)H(+) integrins face each other to form oriented (non-random) nanoclusters. To address the mechanism causing this pattern, we blocked integrin binding to ICAMs in cis, which significantly relieved the face-to-face orientation.