Organized spatial patterns of activated β₂ integrins in arresting neutrophils

Abstract

The transition from leukocyte rolling to firm adhesion is called arrest. β₂ integrins are required for neutrophil arrest¹. Chemokines can trigger neutrophil arrest in vivo² and in vitro³. Resting integrins⁴ exist in a “bent-closed” conformation, i.e., not extended (E⁻) and not high affinity (H⁻), unable to bind ligand. Electron microscopic images of isolated β₂ integrins in “open” and “closed” conformations⁵ inspired the switchblade model of integrin activation from E⁻H⁻ to E⁺H⁻ to E⁺H⁺⁶⁷. Recently⁸, we discovered an alternative pathway of integrin activation from E⁻H⁻ to E⁻H⁺ to E⁺H⁺. Spatial patterning of activated integrins is thought to be required for effective arrest, but so far only diffraction-limited localization maps of activated integrins exist⁸. Here, we combine superresolution microscopy with molecular modeling to identify the molecular patterns of H⁺E⁻, H⁻E⁺, and H⁺E⁺ activated integrins on primary human neutrophils. At the time of neutrophil arrest, E⁺H⁺ integrins form oriented (non-random) nanoclusters that contain a total of 4,625±369 E⁺H⁺ β₂ integrin molecules.