Activation Of Leukocyte Function-associated Antigen-1 Research Articles

Extracellular Ca2+ modulates leukocyte function-associated antigen-1 cell surface distribution on T lymphocytes and consequently affects cell adhesion.

Transition of leukocyte function-associated antigen-1 (LFA-1), from an inactive into an activate state depends on the presence of extracellular Mg2+ and/or Ca2+ ions. Although Mg2+ is directly involved in ligand binding, the role of Ca2+ in LFA-1 mediated adhesion remained obscure. We now demonstrate that binding of Ca2+, but not Mg2+, directly correlates with clustering of LFA-1 molecules at the cell surface of T cells, thereby facilitating LFA-1-ligand interaction. Using a reporter antibody (NKI-L16) that recognizes a Ca(2+)-dependent epitope on LFA-1, we found that Ca2+ can be bound by LFA-1 with different strength. We noticed that weak binding of Ca2+ is associated with a dispersed LFA-1 surface distribution on T cells and with non-responsiveness of these cells to stimuli known to activate LFA-1. In contrast, stable binding of Ca2+ by LFA-1 correlates with a patch-like surface distribution and vivid ligand binding after activation of LFA-1. Mg(2+)-dependent ligand binding does not affect binding of Ca2+ by LFA-1 as measured by NKI-L16 expression, suggesting that Mg2+ binds to a distinct site, and that both cations are important to mediate adhesion. Only Sr2+ ions can replace Ca2+ to express the L16 epitope, and to induce clustering of LFA-1 at the cell surface. We conclude that Ca2+ is involved in avidity regulation of LFA-1 by clustering of LFA-1 molecules at the cell surface, whereas Mg2+ is important in regulation of the affinity of LFA-1 for its ligands.

Plasma membrane lipid packing and leukocyte function-associated antigen-1-dependent aggregation of lymphocytes.

A simple model system for study of adhesion mediated by leukocyte function-associated antigen-1 (LFA-1) is aggregation of lymphocytes stimulated in vitro. Although aggregation is blocked by monoclonal antibodies to LFA-1, not all lymphocytes expressing LFA-1 aggregate, indicating that LFA-1 is necessary but not sufficient for aggregation. To investigate whether the lipid bilayer plays a role in the functional activation of LFA-1, human peripheral blood lymphocytes and murine splenic lymphocytes were stimulated in culture, and measurements made of aggregation vs. packing of plasma membrane lipids. Progression of cells into aggregates was paralleled by a decrease in lipid packing of the population as a whole, as monitored by increased staining with the fluorescent probe merocyanine 540. Cells from aggregates stained more intensely than nonaggregated cells from the same population, indicating that aggregates are preferentially formed from cells in the population with the loosest packed membrane. In contrast, aggregated cells were found to express equivalent or even lower amounts of LFA-1 than nonaggregated cells. Looser lipid packing is therefore associated with the development of LFA-1-dependent aggregation, and might be involved in the functional activation of this cell adhesion molecule.