Distinct kinetic and mechanical properties govern selectin-leukocyte interactions

Journal of Cell Science, May 2004

William D. Hanley, Denis Wirtz, Konstantinos Konstantopoulos

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Distinct kinetic and mechanical properties govern selectin-leukocyte interactions

William D. Hanley 1 Denis Wirtz ) 0 1 Konstantinos Konstantopoulos 1 0 Department of Materials Science and Engineering, The Johns Hopkins University , 3400 N. Charles Street, Baltimore, MD 21218 , USA 1 Department of Chemical and Biomolecular Engineering, The Johns Hopkins University , 3400 N. Charles Street, Baltimore, MD 21218 , USA - Leukocytes are recruited from the bloodstream to sites of inflammation by the selectin family of adhesion receptors. In vivo and in vitro studies reveal distinctive rolling velocities of polymorphonuclear leukocytes over E-, Pand L-selectin substrates. The kinetic and mechanical properties of the selectin-ligand bonds responsible for these differences at the single-molecule level are not well understood. Using single-molecule force spectroscopy, we probe in situ the rupture force, unstressed off-rate and reactive compliance of single selectin receptors to single ligands on whole human polymorphonuclear leukocytes (PMNs) under conditions that preserve the proper orientation and post-translational modifications of the selectin ligands. Single L-selectin bonds to PMNs were more labile than either E- or P-selectin in the presence of an applied force. This outcome, along with a higher Central to all cell-cell communications are the adhesive interactions between biological receptors and their respective ligands. The kinetics regulating receptor-ligand binding impart unique properties that allow specialized cells to interact with one other amidst the challenge of physiological and competitive stresses. This phenomenon is perhaps best represented by the remarkable ability of the selectins (E-, Pand L-selectin) to mediate the initial adhesion events during leukocyte recruitment to sites of inflammation. The distinctiveness of selectin binding is attributed to their fast association/dissociation rates, as well as their ability to form high-strength tethers under rapid loading. These properties allow polymorphonuclear leukocytes (PMNs) first to tether and roll on activated vascular endothelium under hydrodynamic shear, before firm adhesion and extravasation into the tissue space (Konstantopoulos et al., 1998; McEver, 2002; Springer, 1995). In vivo (Jung et al., 1996; Kunkel and Ley, 1996) and in vitro (Alon et al., 1997; Alon et al., 1998; Puri et al., 1997) studies both reveal that E-, P- and L-selectin-mediated interactions display characteristic leukocyte rolling velocities. Structurally similar, the selectins consist of an N-terminal, calcium-dependent lectin domain, an epidermal-growth-factorlike (EGF) domain, a variable number of consensus repeat units, a transmembrane domain (TM) and an intracellular cytoplasmic tail (cyto) (Fig. 1A) (Konstantopoulos et al., 1998; McEver et al., 1995). They also share a relatively high degree unstressed off-rate and a higher reactive compliance, explain the faster L-selectin-mediated rolling. By quantifying binding frequency in the presence of a specific blocking monoclonal antibody or following enzyme treatment, we determined that P-selectin glycoprotein ligand-1 is a high-affinity ligand for E-selectin on PMNs under force. The rupture force spectra and corresponding unstressed off-rate and reactive compliance of selectinligand bonds provide mechanistic insights that might help to explain the variable rolling of leukocytes over different selectin substrates. Supplemental figure available online of homology within each region (McEver et al., 1995). The primary module for adhesion is the lectin domain but other features (including the EGF region and the number of consensus repeat units) might confer unique biophysical and mechanical properties on each selectin when binding to its ligand (McEver et al., 1995). Additionally, selectin ligands recognize a stereochemically precise arrangement of sialic acid and fucose, termed sialyl Lewisx (McEver, 2002; Varki, 1997). Although they share many common features, the selectins possess differences in their expression kinetics, tissue distributions and ability to promote leukocyte attachment, perhaps reflecting their unique roles in diverse immunological processes. E-selectin, expressed on the vascular endothelium, is transcriptionally regulated and induced in response to inflammatory stimuli (Konstantopoulos et al., 1998). P-selectin expression on both the vascular endothelium and on platelets is also inducible, but its storage in the Weibel-Palade bodies of endothelial cells and the a -granules of platelets permits rapid mobilization (within seconds to minutes) to the plasma membrane upon cell activation (Konstantopoulos et al., 1998). L-selectin, by contrast, is constitutively expressed on most leukocyte subpopulations and is responsible for amplifying the inflammatory response through leukocyte-leukocyte interactions or so-called secondary tethers (Alon et al., 1996; Konstantopoulos et al., 1998). P-selectin glycoprotein ligand 1 (PSGL-1) is the bestcharacterized ligand capable of binding to all thr (...truncated)


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William D. Hanley, Denis Wirtz, Konstantinos Konstantopoulos. Distinct kinetic and mechanical properties govern selectin-leukocyte interactions, Journal of Cell Science, 2004, pp. 2503-2511, 117/12, DOI: 10.1242/jcs.01088