Dataset Information

Ligand- and cation-induced structural alterations of the leukocyte integrin LFA-1.

ABSTRACT: In ?I integrins, including leukocyte function-associated antigen 1 (LFA-1), ligand-binding function is delegated to the ?I domain, requiring extra steps in the relay of signals that activate ligand binding and coordinate it with cytoplasmic signals. Crystal structures reveal great variation in orientation between the ?I domain and the remainder of the integrin head. Here, we investigated the mechanisms involved in signal relay to the ?I domain, including whether binding of the ligand intercellular adhesion molecule-1 (ICAM-1) to the ?I domain is linked to headpiece opening and engenders a preferred ?I domain orientation. Using small-angle X-ray scattering and negative-stain EM, we define structures of ICAM-1, LFA-1, and their complex, and the effect of activation by Mn2+ Headpiece opening was substantially stabilized by substitution of Mg2+ with Mn2+ and became complete upon ICAM-1 addition. These agents stabilized ?I-headpiece orientation, resulting in a well-defined orientation of ICAM-1 such that its tandem Ig-like domains pointed in the opposite direction from the ?-subunit leg of LFA-1. Mutations in the integrin ?I domain ?1/?1' helix stabilizing either the open or the closed ?I-domain conformation indicated that ?1/?1' helix movements are linked to ICAM-1 binding by the ?I domain and to the extended-open conformation of the ectodomain. The LFA-1-ICAM-1 orientation described here with ICAM-1 pointing anti-parallel to the LFA-1 ?-subunit leg is the same orientation that would be stabilized by tensile force transmitted between the ligand and the actin cytoskeleton and is consistent with the cytoskeletal force model of integrin activation.

SUBMITTER: Sen M

PROVIDER: S-EPMC5925802 | biostudies-literature | 2018 Apr

REPOSITORIES: biostudies-literature

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Ligand- and cation-induced structural alterations of the leukocyte integrin LFA-1.

Sen Mehmet M Koksal Adem C AC Yuki Koichi K Wang Jianchuan J Springer Timothy A TA

The Journal of biological chemistry 20180305 17

In αI integrins, including leukocyte function-associated antigen 1 (LFA-1), ligand-binding function is delegated to the αI domain, requiring extra steps in the relay of signals that activate ligand binding and coordinate it with cytoplasmic signals. Crystal structures reveal great variation in orientation between the αI domain and the remainder of the integrin head. Here, we investigated the mechanisms involved in signal relay to the αI domain, including whether binding of the ligand intercellul ...[more]

PMID: 29507098

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Project description:Using single-molecule force spectroscopy to probe ICAM-1 interactions with recombinant alphaLbeta2 immobilized on microspheres and beta2 integrin on neutrophils, we quantified an impressive hierarchy of long-lived, high-strength states of the integrin bond, which start from basal levels with activation in solutions of divalent cations and shift dramatically upward to hyperactivated states with cell signaling. Taking advantage of very rare events, we used repeated measurements of bond lifetimes under steady ramps of force to achieve a direct assay for the off-rates of ICAM-1 from beta2 integrin throughout the course of each experiment. In our companion article I, we demonstrate the assay using results from tests of a monovalent ICAM-1 probe against recombinant alphaLbeta2 on microspheres in millimolar solutions of divalent cations (Ca2+, Mg2+, Mn2+). In this article, we examine the impact of inside-out and outside-in signaling in neutrophils on the lifetimes and mechanical strengths of ICAM-1 bonds to beta2 integrin on the cell surface. Even though ICAM-1 bonds to recombinant alphaLbeta2 on microspheres in Mg2+ or Mn2+ can live for long periods of time under slow pulling, here we show that stimulation of neutrophils in Mg2+ plus the chemokine IL-8 (i.e., inside-out signaling) induces several-hundred-fold longer lifetimes for ICAM-1 attachments to LFA-1, creating strong bonds at very slow pulling speeds where none are perceived in Mg2+ or Mn2+ alone. Similar changes are observed with outside-in signaling, i.e., long lifetimes and increased bond strength also occur when neutrophils are bound with the activating (anti-CD18) monoclonal 240Q. Limiting our investigation to rare events using very dilute ICAM-1 probes, we show that although the prolonged lifetimes of cell surface attachments for both inside-out and outside-in signaling exhibit single-bond-like statistics for dissociation under force, they are consistent with a tightly coupled dimeric ICAM-1 interaction with a pair of LFA-1 heterodimers.

Dataset Information

Ligand- and cation-induced structural alterations of the leukocyte integrin LFA-1.

Publications

Ligand- and cation-induced structural alterations of the leukocyte integrin LFA-1.

Similar Datasets

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