The Lymphocyte Function–associated Antigen 1 I Domain Is a Transient Binding Module for Intercellular Adhesion Molecule (ICAM)-1 and ICAM-3 in Hydrodynamic Flow

Abstract

The I domain of lymphocyte function–associated antigen (LFA)-1 contains an intercellular adhesion molecule (ICAM)-1 and ICAM-3 binding site, but the relationship of this site to regulated adhesion is unknown. To study the adhesive properties of the LFA-1 I domain, we stably expressed a GPI-anchored form of this I domain (I-GPI) on the surface of baby hamster kidney cells. I-GPI cells bound soluble ICAM-1 (sICAM-1) with a low avidity and affinity. Flow cell experiments demonstrated a specific rolling interaction of I-GPI cells on bilayers containing purified full length ICAM-1 or ICAM-3. The LFA-1 activating antibody MEM-83, or its Fab fragment, decreased the rolling velocity of I-GPI cells on ICAM-1–containing membranes. In contrast, the interaction of I-GPI cells with ICAM-3 was blocked by MEM-83. Rolling of I-GPI cells was dependent on the presence of Mg2+. Mn2+ only partially substituted for Mg2+, giving rise to a small fraction of rolling cells and increased rolling velocity. This suggests that the I domain acts as a transient, Mg2+-dependent binding module that cooperates with another Mn2+-stimulated site in LFA-1 to give rise to the stable interaction of intact LFA-1 with ICAM-1.

Structural Basis of Integrin Regulation and Signaling

“…In in vitro shear flow assays and in vivo, the antagonists enhance rolling of leukocytes and inhibit firm adhesion (57). These results on ICAM-1 substrates suggest that the postulated αL Glu-310-β2 MIDAS interaction is not required for rolling adhesion, in agreement with the ability of isolated, wild-type αL I domains to support rolling adhesion (58,59), but is required for firm adhesion. LFA-1 containing an αL Glu-310 → Ala mutation shows lowered expression of activation epitopes in the β2 I domain and leg, demonstrating cooperativity between the postulated αL Glu-310-β2 MIDAS interaction and conformational rearrangements elsewhere in the LFA-1 molecule.…”

Expression of a structural domain of the β2 subunit essential for αMβ2 ligand recognition

“…Conceivably, the inability of the cells expressing either the ␣ M , ␤ 2 , or ␤ 2 (LB) chain alone to adhere to iC3b may be due to their lower functional affinity of these individual subunits for iC3b compared to complete ␣ M ␤ 2 heterodimer. This hypothesis is supported by the recent demonstration in a flow cell system that a GPI-linked I domain of ␣ L supports binding but not stable adhesion to immobilized ICAM-1 [56]. Other site(s) within ␣ L ␤ 2 are proposed to cooperate with the I domain, resulting in high-affinity stable adhesion between ␣ L ␤ 2 and ICAM-1.…”

An isolated, surface-expressed I domain of the integrin αLβ2 is sufficient for strong adhesive function when locked in the open conformation with a disulfide bond

“…Although the I domain is implicated as an important site for ligand binding in those integrins that contain it, we found that the isolated wild-type ␣L I domain did not support K562 transfectant binding to immobilized ICAM-1. Previous work using a glycosylphosphatidylinositol-anchored, isolated wildtype ␣L I domain has shown that ligand binding by the isolated I domain is much weaker than that by the intact integrin, and is only detected when the I domain is expressed at very high levels (33). It was suggested that the I domain alone might not be sufficient to mediate strong and stable interaction with ligand.…”