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Forcing switch from short- to intermediate- and long-lived states of the alphaA domain generates LFA-1/ICAM-1 catch bonds.

ABSTRACT: Binding of lymphocyte function-associated antigen-1 (LFA-1) to intercellular adhesion molecule-1 (ICAM-1) mediates leukocyte adhesion under force. Using a biomembrane force probe capable of measuring single bond interactions, we showed ICAM-1 binding to LFA-1 at different conformations, including the bent conformation with the lowest affinity. We quantify how force and conformations of LFA-1 regulate its kinetics with ICAM-1. At zero-force, on-rates were substantially changed by conditions that differentially favor a bent or extended LFA-1 with a closed or open headpiece; but off-rates were identical. With increasing force, LFA-1/ICAM-1 bond lifetimes (reciprocal off-rates) first increased (catch bonds) and then decreased (slip bonds). Three states with distinct off-rates were identified from lifetime distributions. Force shifted the associated fractions from the short- to intermediate- and long-lived states, producing catch bonds at low forces, but increased their off-rates exponentially, converting catch to slip bonds at high forces. An internal ligand antagonist that blocks pulling of the ?(7)-helix suppressed the intermediate-/long-lived states and eliminated catch bonds, revealing an internal catch bond between the ?A and ?A domains. These results elucidate an allosteric mechanism for the mechanochemistry of LFA-1/ICAM-1 binding.

SUBMITTER: Chen W

PROVIDER: S-EPMC2975219 | biostudies-literature | 2010 Nov

REPOSITORIES: biostudies-literature

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Forcing switch from short- to intermediate- and long-lived states of the alphaA domain generates LFA-1/ICAM-1 catch bonds.

Chen Wei W Lou Jizhong J Zhu Cheng C

The Journal of biological chemistry 20100906 46

Binding of lymphocyte function-associated antigen-1 (LFA-1) to intercellular adhesion molecule-1 (ICAM-1) mediates leukocyte adhesion under force. Using a biomembrane force probe capable of measuring single bond interactions, we showed ICAM-1 binding to LFA-1 at different conformations, including the bent conformation with the lowest affinity. We quantify how force and conformations of LFA-1 regulate its kinetics with ICAM-1. At zero-force, on-rates were substantially changed by conditions that ...[more]

PMID: 20819952

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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.

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 integrin activation in solutions of divalent cations and shift dramatically upward to hyperactivated states with cell signaling in leukocytes. 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 in each experiment. Of fundamental importance, the assay for off-rates does not depend on how the force is applied over time, and remains valid when the rates of dissociation change with different levels of force. In this first article, we present results from tests of a monovalent ICAM-1 probe against immobilized alphaLbeta2 in environments of divalent cations (Ca2+, Mg2+, and Mn2+) and demonstrate in detail the method for assay of off-rates. When extrapolated to zero force, the force-free values for the off-rates are found to be consistent with published solution-based assays of soluble ICAM-1 dissociation from immobilized LFA-1, i.e., approximately 10(-2)/s in Mg2+ or Mn2+ and approximately 1/s in Ca2+. At the same time, as expected for adhesive function, we find that the beta2 integrin bonds activated in Mn2+ or Mg2+ possess significant and persistent mechanical strength (e.g., >20 pN for >1 s) even when subjected to slow force ramps (<10 pN/s). As discussed in our companion article, using the same assay, we find that although the rates of dissociation for diICAM-1fc bonds to LFA-1 on neutrophils in Mn2+ are similar to those for mICAM-1 bonds to recombinant alphaLbeta2 on microspheres, they appear to represent a dimeric attachment to a pair of tightly clustered integrin heterodimers. The mechanical strengths and lifetimes of the dimeric interactions increase dramatically when the neutrophils are stimulated by the chemokine IL-8 or are bound with an allosterically activating (anti-CD18) monoclonal antibody, demonstrating the major impact of cell signaling on LFA-1.

Dataset Information

Forcing switch from short- to intermediate- and long-lived states of the alphaA domain generates LFA-1/ICAM-1 catch bonds.

Publications

Forcing switch from short- to intermediate- and long-lived states of the alphaA domain generates LFA-1/ICAM-1 catch bonds.

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