Cytoskeletal Regulation Couples LFA-1 Conformational Changes to Receptor Lateral Mobility and Clustering

Abstract

The alpha(L)beta(2) integrin (leukocyte function-associated antigen-1 [LFA-1]) is regulated to engage and maintain T cell adhesion. Conformational changes in the receptor are associated with changes in receptor-ligand affinity and are necessary for firm adhesion. Less well understood is the relationship between receptor conformation and the regulation of its lateral mobility. We have used fluorescence photobleaching recovery and single-particle tracking to measure the lateral mobility of specific conformations of LFA-1. These measurements show that different receptor conformations have distinct diffusion profiles and that these profiles vary according to the activation state of the cell. Notably, a high-affinity conformation of LFA-1 is mobile on resting cells but immobile on phorbol-12-myristate-13-acetate-activated cells. This activation-induced immobilization is prevented by a calpain inhibitor and by an allosteric LFA-1 inhibitor. Our results suggest that current models of LFA-1 regulation are incomplete and that LFA-1 confinement by cytoskeletal attachment regulates cell adhesion both negatively and positively.

Requirement of open headpiece conformation for activation of leukocyte integrin αXβ2

“…Our data suggest that the intermediate affinity state observed in the literature for the related β 2 integrin LFA-1 (16)(17)(18)(19)(20) does not have to come from an individual conformational state, but could instead result from a mixture of conformational states. Previous studies showed that LFA-1 binding to ICAM-3, compared with ICAM-1, required more activation as defined by the number of activation agents needed (19)(20)(21).…”

“…At the leading edge of migrating T lymphocytes, a distinct population of LFA-1 was defined as in an intermediate affinity state by the exposure of different monoclonal antibody epitopes (16). Lateral mobility measurements of LFA-1 on T cell surfaces suggested an intermediate conformational state with distinct diffusion profile (17). In vitro flow chamber experiments showed that an intermediate affinity of LFA-1 on lymphocytes could be induced by immobilized chemokine (18).…”

LFA-1 Regulates CD8+ T Cell Activation via T Cell Receptor-mediated and LFA-1-mediated Erk1/2 Signal Pathways

“…It is controversial whether the affinity or avidity state of LFA-1 is involved in the insideout and outside-in signals. A recent publication demonstrated that cytoskeletal regulation couples LFA-1 conformational changes to receptor lateral mobility and clustering; thus, both affinity and avidity states of LFA-1 provide regulation of receptor function on lymphocytes (42). Previous studies reported that LFA-1 ligation can co-stimulate T cells for IL-2 production, cell proliferation, and Th1 polarization (27,(43)(44).…”

RhoA Is Involved in LFA-1 Extension Triggered by CXCL12 but Not in a Novel Outside-In LFA-1 Activation Facilitated by CXCL9

“…In the present study, we demonstrate that the RhoA 23/40 domain is critical for the earliest CCL21-triggered LFA-1 activation events measured in vivo as well as for a CXCL12-induced LFA-1 extension step critical for integrin adhesiveness to ICAM-1 under shear flow, but is not involved in spontaneous or CXCL9-induced LFA-1 adhesiveness to ICAM-1 in human T lymphocytes. We also find that CCL21 and CXCL12, on one hand, and CXCL9 on the other hand, use different modalities to activate LFA-1 adhesiveness, whereas CCL21 and CXCL12 trigger LFA-1 conformational changes before ligand binding and thereby increases the pool of extended LFA-1 readily available for interacting with surface-bound ICAM-1 (10), CXCL9 triggers a unique LFA-1 activation process that takes place without conformational changes before ligand binding and therefore may rely on spontaneously extended subsets of LFA-1 that preexist on T cells and preferentially engage with ICAM-1 (4,39). We also show that an alternative chemokine-independent, PKC-induced pathway of LFA-1 activation also does not trigger integrin conformational extension or integrin clustering before ICAM-1 binding and takes place independently of RhoA.…”