共查询到20条相似文献,搜索用时 15 毫秒
1.
Identification of different binding sites in the dendritic cell-specific receptor DC-SIGN for intercellular adhesion molecule 3 and HIV-1. 总被引:15,自引:0,他引:15
Teunis B H Geijtenbeek Gerard C F van Duijnhoven Sandra J van Vliet Elmar Krieger Gert Vriend Carl G Figdor Yvette van Kooyk 《The Journal of biological chemistry》2002,277(13):11314-11320
The novel dendritic cell (DC)-specific human immunodeficiency virus type 1 (HIV-1) receptor DC-SIGN plays a key role in the dissemination of HIV-1 by DC. DC-SIGN is thought to capture HIV-1 at mucosal sites of entry, facilitating transport to lymphoid tissues, where DC-SIGN efficiently transmits HIV-1 to T cells. DC-SIGN is also important in the initiation of immune responses by regulating DC-T cell interactions through intercellular adhesion molecule 3 (ICAM-3). We have characterized the mechanism of ligand binding by DC-SIGN and identified the crucial amino acids involved in this process. Strikingly, the HIV-1 gp120 binding site in DC-SIGN is different from that of ICAM-3, consistent with the observation that glycosylation of gp120, in contrast to ICAM-3, is not crucial to the interaction with DC-SIGN. A specific mutation in DC-SIGN abrogated ICAM-3 binding, whereas the HIV-1 gp120 interaction was unaffected. This DC-SIGN mutant captured HIV-1 and infected T cells in trans as efficiently as wild-type DC-SIGN, demonstrating that ICAM-3 binding is not necessary for HIV-1 transmission. This study provides a basis for the design of drugs that inhibit or alter interactions of DC-SIGN with gp120 but not with ICAM-3 or vice versa and that have a therapeutic value in immunological diseases and/or HIV-1 infections. 相似文献
2.
Force spectroscopy of the leukocyte function-associated antigen-1/intercellular adhesion molecule-1 interaction 总被引:7,自引:0,他引:7
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Interactions between leukocyte function-associated antigen-1 (LFA-1) with its cognate ligand, intercellular adhesion molecule-1 (ICAM-1) play a crucial role in leukocyte adhesion. Because the cell and its adhesive components are subject to external perturbation from the surrounding flow of blood, it is important to understand the binding properties of the LFA-1/ICAM-1 interaction in both steady state and in the presence of an external pulling force. Here we report on atomic force microscopy (AFM) measurements of the unbinding of LFA-1 from ICAM-1. The single molecule measurements revealed the energy landscape corresponding to the dissociation of the LFA-1/ICAM-1 complex and provided the basis for defining the energetic determinants of the complex at equilibrium and under the influence of an external force. The AFM force measurements were performed in an experimental system consisting of an LFA-1-expressing T cell hybridoma, 3A9, attached to the end of the AFM cantilever and an apposing surface expressing ICAM-1. In measurements covering three orders of magnitude change in force loading rate, the LFA-1/ICAM-1 force spectrum (i.e., unbinding force versus loading rate) revealed a fast and a slow loading regime that characterized a steep inner activation barrier and a wide outer activation barrier, respectively. The addition of Mg(2+), a cofactor that stabilizes the LFA-1/ICAM-1 interaction, elevated the unbinding force of the complex in the slow loading regime. In contrast, the presence of EDTA suppressed the inner barrier of the LFA-1/ICAM-1 complex. These results suggest that the equilibrium dissociation constant of the LFA-1/ICAM-1 interaction is regulated by the energetics of the outer activation barrier of the complex, while the ability of the complex to resist a pulling force is determined by the divalent cation-dependent inner activation barrier. 相似文献
3.
R P Lauener R S Geha D Vercelli 《Journal of immunology (Baltimore, Md. : 1950)》1990,145(5):1390-1394
Murine anti-CD14 mAb which recognize different CD14 epitopes induced marked homotypic adhesion of normal human monocytes. Induction of aggregation by anti-CD14 mAb required Mg2+, occurred at an optimal temperature of 37 degrees C, but not at 4 degrees C, and exhibited a kinetics which differed from adhesion triggered by IFN-gamma and anti-CD43 mAb. Monocyte adhesion induced by anti-CD14 mAb required neither Fcy gamma R engagement nor cross-linking of CD14, because adhesion was induced by F(ab)'2 fragments, as well as by monovalent F(ab) fragments of anti-CD14 mAb. mAb to CD11a, CD18, and intercellular adhesion molecule-1 (ICAM-1), but not antibodies to CD11b and CD11c, inhibited monocyte adhesion induced by CD14 engagement. These results indicate that CD14-dependent adhesion is mediated by lymphocyte function-associated Ag-1/ICAM-1 interactions. This was confirmed by the absence of aggregation in anti-CD14-stimulated cells from a patient with leukocyte adhesion deficiency. Monocyte adhesion upon CD14 engagement was blocked by an inhibitor of protein kinases, sphingosine. This suggests that protein kinases play a role in the intracellular signaling pathway(s) which couple CD14 to lymphocyte function-associated Ag-1/ICAM-1. 相似文献
4.
Keating SM Clark KR Stefanich LD Arellano F Edwards CP Bodary SC Spencer SA Gadek TR Marsters JC Beresini MH 《Protein science : a publication of the Protein Society》2006,15(2):290-303
The lymphocyte function-associated antigen-1 (LFA-1) binding of a unique class of small-molecule antagonists as represented by compound 3 was analyzed in comparison to that of soluble intercellular adhesion molecule-1 (sICAM-1) and A-286982, which respectively define direct and allosteric competitive binding sites within LFA-1's inserted (I) domain. All three molecules antagonized LFA-1 binding to ICAM-1-Immunoglobulin G fusion (ICAM-1-Ig) in a competition ELISA, but only compound 3 and sICAM-1 inhibited the binding of a fluorescein-labeled analog of compound 3 to LFA-1. Compound 3 and sICAM-1 displayed classical direct competitive binding behavior with ICAM-1. Their antagonism of LFA-1 was surmountable by both ICAM-1-Ig and a fluorescein-labeled compound 3 analog. The competition of both sICAM-1 and compound 3 with ICAM-1-Ig for LFA-1 resulted in equivalent and linear Schild plots with slopes of 1.24 and 1.26, respectively. Cross-linking studies with a photoactivated analog of compound 3 localized the high-affinity small-molecule binding site to the N-terminal 507 amino acid segment of the alpha chain of LFA-1, a region that includes the I domain. In addition, cells transfected with a variant of LFA-1 lacking this I domain showed no significant binding of a fluorescein-labeled analog of compound 3 or ICAM-1-Ig. These results demonstrate that compound 3 inhibits the LFA-1/ICAM-1 binding interaction in a directly competitive manner by binding to a high-affinity site on LFA-1. This binding site overlaps with the ICAM-1 binding site on the alpha subunit of LFA-1, which has previously been localized to the I domain. 相似文献
5.
Purified intercellular adhesion molecule-1 (ICAM-1) is a ligand for lymphocyte function-associated antigen 1 (LFA-1) 总被引:178,自引:0,他引:178
Lymphocyte function-associated antigen 1 (LFA-1) is a leukocyte cell surface glycoprotein that promotes intercellular adhesion in immunological and inflammatory reactions. It is an alpha beta complex that is structurally related to receptors for extracellular matrix components, and thus belongs to the integrin family. ICAM-1 (intercellular adhesion molecule-1) is a distinct cell surface glycoprotein. Its broad distribution, regulated expression in inflammation, and involvement in LFA-1-dependent cell-cell adhesion have suggested that ICAM-1 may be a ligand for LFA-1. We have purified ICAM-1 and incorporated it into artificial supported lipid membranes. LFA-1+ but not LFA-1- cells bound to ICAM-1 in the artificial membranes, and the binding could be specifically inhibited by anti-ICAM-1 treatment of the membranes or by anti-LFA-1 treatment of the cells. The cell binding to ICAM-1 required metabolic energy production, an intact cytoskeleton, and the presence of Mg2+ and was temperature dependent, characteristics of LFA-1- and ICAM-1-dependent cell-cell adhesion. 相似文献
6.
Sarantos MR Raychaudhuri S Lum AF Staunton DE Simon SI 《The Journal of biological chemistry》2005,280(31):28290-28298
Neutrophil rolling and transition to arrest on inflamed endothelium are dynamically regulated by the affinity of the beta(2) integrin CD11a/CD18 (leukocyte function associated antigen 1 (LFA-1)) for binding intercellular adhesion molecule (ICAM)-1. Conformational shifts are thought to regulate molecular affinity and adhesion stability. Also critical to adhesion efficiency is membrane redistribution of active LFA-1 into dense submicron clusters where multimeric interactions occur. We examined the influences of affinity and dimerization of LFA-1 on LFA-1/ICAM-1 binding by engineering a cell-free model in which two recombinant LFA-1 heterodimers are bound to respective Fab domains of an antibody attached to latex microspheres. Binding of monomeric and dimeric ICAM-1 to dimeric LFA-1 was measured in real time by fluorescence flow cytometry. ICAM-1 dissociation kinetics were measured while LFA-1 affinity was dynamically shifted by the addition of allosteric small molecules. High affinity LFA-1 dissociated 10-fold faster when bound to monomeric compared with dimeric ICAM-1, corresponding to bond lifetimes of 25 and 330 s, respectively. Downshifting LFA-1 into an intermediate affinity state with the small molecule I domain allosteric inhibitor IC487475 decreased the difference in dissociation rates between monomeric and dimeric ICAM-1 to 4-fold. When LFA-1 was shifted into the low affinity state by lovastatin, both monomeric and dimeric ICAM-1 dissociated in less than 1 s, and the dissociation rates were within 50% of each other. These data reveal the respective importance of LFA-1 affinity and proximity in tuning bond lifetime with ICAM-1 and demonstrate a nonlinear increase in the bond lifetime of the dimer versus the monomer at higher affinity. 相似文献
7.
Krieglstein CF Salter JW Cerwinka WH Russell JM Schuermann G Bruewer M Laroux FS Grisham MB Granger DN 《Biochemical and biophysical research communications》2001,282(2):635-642
Adhesion molecules have been implicated in the pathogenesis of inflammatory bowel diseases. We investigated their expression and contribution to leukocyte recruitment in experimental intestinal inflammation. Ileitis was induced in Sprague-Dawley rats by two injections of indomethacin (7.5 mg/kg), given 24 h apart. Endothelial intercellular adhesion molecule-1 (ICAM-1) expression was quantified using the dual radiolabeled monoclonal antibody technique and Mac-1 (CD11b/CD18) expression on leukocytes by flow cytometry. Leukocyte infiltration was monitored by tissue myeloperoxidase (MPO) activity. The first indomethacin injection induced a time- and site-dependent increase of ICAM-1 expression in ileal mucosa and muscularis. The second injection resulted in a reduction of ICAM-1 expression below constitutive levels whereas Mac-1 was upregulated. MPO changes paralleled lesion development over 48 h. ICAM-1 and MPO values were correlated for the first 24 h. Immunoneutralization of either ICAM-1 or Mac-1 attenuated mucosal injury. We conclude that (i) indomethacin-induced ileitis is associated with a temporally disassociated upregulation of ICAM-1 and (ii) despite a reduction in ICAM-1 after 24 h, ICAM-1, in concert with Mac-1, contributes to mucosal injury and leukocyte infiltration elicited by indomethacin. 相似文献
8.
Endothelial cells proactively form microvilli-like membrane projections upon intercellular adhesion molecule 1 engagement of leukocyte LFA-1 总被引:12,自引:0,他引:12
Carman CV Jun CD Salas A Springer TA 《Journal of immunology (Baltimore, Md. : 1950)》2003,171(11):6135-6144
Specific leukocyte/endothelial interactions are critical for immunity and inflammation, yet the molecular details of this interaction interface remain poorly understood. Thus, we investigated, with confocal microscopy, the distribution dynamics of the central adhesion molecules ICAM-1 and LFA-1 in this context. Monolayers of activated HUVECs stained with fluorescent anti-ICAM-1 Fabs or Chinese hamster ovary-K1 cells expressing ICAM-1-green fluorescent protein were allowed to bind LFA-1-bearing monocytes, neutrophils, or K562 LFA-1 transfectants. ICAM-1 was rapidly relocalized to newly formed microvilli-like membrane projections in response to binding LFA-1 on leukocytes. These ICAM-1-enriched projections encircled the leukocytes extending up their sides and clustered LFA-1 underneath into linear tracks. Projections formed independently of VCAM-1/very late Ag 4 interactions, shear, and proactive contributions from the LFA-1-bearing cells. In the ICAM-1-bearing endothelial cells, projections were enriched in actin but not microtubules, required intracellular calcium, and intact microfilament and microtubule cytoskeletons and were independent of Rho/Rho kinase signaling. Disruption of these projections with cytochalasin D, colchicine, or BAPTA-AM had no affect on firm adhesion. These data show that in response to LFA-1 engagement the endothelium proactively forms an ICAM-1-enriched cup-like structure that surrounds adherent leukocytes but is not important for firm adhesion. This finding leaves open a possible role in leukocyte transendothelial migration, which would be consistent with the geometry and kinetics of formation of the cup-like structure. 相似文献
9.
Carcinoembryonic antigen, a human tumor marker, functions as an intercellular adhesion molecule 总被引:56,自引:0,他引:56
Carcinoembryonic antigen (CEA) is a member of a family of cell surface glycoproteins that are produced in excess in essentially all human colon carcinomas and in a high proportion of carcinomas at many other sites. The function of this widely used tumor marker and its relevance to malignant transformation is therefore of considerable interest. We demonstrate here that CEA mediates Ca2+-independent, homotypic aggregation of cultured human colon adenocarcinoma cells (LS-180) and rodent cells transfected with functional CEA cDNA. Furthermore, CEA can effect the homotypic sorting of cells in heterogeneous populations of aggregating cells. CEA can thus be considered a new addition to the family of intercellular adhesion molecules. We also show that, whereas CEA is localized mainly to epithelial cell membranes facing the lumen in normal adult intestine, it is found on adjacent cell membranes in both embryonic intestine and colonic tumors. A model for the role of CEA in the tissue architecture of adult, embryonic, and aberrant tumor intestinal epithelium is presented. 相似文献
10.
Song G Lazar GA Kortemme T Shimaoka M Desjarlais JR Baker D Springer TA 《The Journal of biological chemistry》2006,281(8):5042-5049
The interaction between integrin lymphocyte function-associated antigen-1 (LFA-1) and its ligand intercellular adhesion molecule-1 (ICAM-1) is critical in immunological and inflammatory reactions but, like other adhesive interactions, is of low affinity. Here, multiple rational design methods were used to engineer ICAM-1 mutants with enhanced affinity for LFA-1. Five amino acid substitutions 1) enhance the hydrophobicity and packing of residues surrounding Glu-34 of ICAM-1, which coordinates to a Mg2+ in the LFA-1 I domain, and 2) alter associations at the edges of the binding interface. The affinity of the most improved ICAM-1 mutant for intermediate- and high-affinity LFA-1 I domains was increased by 19-fold and 22-fold, respectively, relative to wild type. Moreover, potency was similarly enhanced for inhibition of LFA-1-dependent ligand binding and cell adhesion. Thus, rational design can be used to engineer novel adhesion molecules with high monomeric affinity; furthermore, the ICAM-1 mutant holds promise for targeting LFA-1-ICAM-1 interaction for biological studies and therapeutic purposes. 相似文献
11.
12.
Evidence that measles virus hemagglutinin initiates modulation of leukocyte function-associated antigen 1 expression.
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Measles virus (MV), human immunodeficiency virus, Epstein-Barr virus, and other leukotropic viruses can modulate the expression of leukocyte function antigen 1 (LFA-1) on the surface of infected and nearby leukocytes. This ability to induce changes in LFA-1 expression may play an important role in the pathogenesis of these viruses. However, the mechanism(s) involved in virus-mediated regulation of LFA-1 is unknown. Evidence is presented in this report that it is the MV hemagglutinin (H) protein that initiates up-regulation of LFA-1 expression in leukocyte cultures infected with this virus. Indeed, comparison of the abilities of different MV strains to modulate LFA-1 expression, examination of published nucleotide sequences for the H proteins of different vaccine strains, and competitive inhibition assays using oligopeptides homologous or heterologous to a region of the H protein gene encompassing amino acid 116 (from the amino terminus) all suggest that it is this portion of the H protein that is responsible for MV-induced alteration of LFA-1. These comparisons also support the hypothesis that there is a relationship between the abilities of different MV strains to alter LFA-1 expression and their pathogenic potentials. 相似文献
13.
14.
Cellular activation of leukocyte function-associated antigen-1 and its affinity are regulated at the I domain allosteric site 总被引:9,自引:0,他引:9
Lupher ML Harris EA Beals CR Sui LM Liddington RC Staunton DE 《Journal of immunology (Baltimore, Md. : 1950)》2001,167(3):1431-1439
The I domain of the integrin LFA-1 possesses a ligand binding interface that includes the metal ion-dependent adhesion site. Binding of the LFA-1 ligand, ICAM-1 to the metal ion-dependent adhesion site is regulated by the I domain allosteric site (IDAS). We demonstrate here that intracellular signaling leading to activation of LFA-1 binding to ICAM-1 is regulated at the IDAS. Inhibitory mutations in or proximal to the IDAS are dominant to cytoplasmic signals that activate binding to ICAM-1. In addition, mutational activation at the IDAS greatly increases the binding of lymphocyte-expressed LFA-1 to ICAM-1 in response to PMA, but does not result in constitutive binding. Binding of a novel CD18 activation epitope mAb to LFA-1 in response to soluble ICAM-1 binding was also blocked by inhibitory and was enhanced by activating IDAS mutations. Surface plasmon resonance using soluble wild-type LFA-1 and an IDAS mutant of LFA-1 indicate that the IDAS can regulate a 6-fold change in the K(d) of ICAM-1 binding. The K(d) of wild-type LFA-1 (1.2 x 10(-1) s(-1)) differed with that of the activating IDAS mutant (1.9 x 10(-2) s(-1)), but their K(a) values were identical (2.2 x 10(5) M(-1)s(-1)). We propose that IDAS regulates the binding of LFA-1 to ICAM-1 activated by intracellular signals. IDAS can control the affinity state of LFA-1 with concomitant I domain and CD18 conformational changes. 相似文献
15.
Taheri M Saragovi U Fuks A Makkerh J Mort J Stanners CP 《The Journal of biological chemistry》2000,275(35):26935-26943
The homophilic binding of extracellular domains of membrane-bound immunoglobulin superfamily (IgSF) molecules is often required for intercellular adhesion and signaling. Carcinoembryonic antigen (CEA), a member of the IgSF, is a widely used tumor marker that functions in vitro as a homotypic intercellular adhesion molecule. CEA has also been shown to contribute to tumorigenicity by inhibiting cellular differentiation, an effect that requires the homophilic binding of its extracellular domains. It was of interest, therefore, to identify small subdomain sequences in CEA that could serve as a focus in the design of peptides that disrupt CEA-mediated intercellular adhesion. Three subdomains in the N-terminal domain of CEA, identified by site-directed deletions and point mutations, were shown to be required for intercellular adhesion. Cyclized peptides representing two of these subdomains, (42)NRQII and (80)QNDTG, were found to be effective in blocking CEA-mediated cellular aggregation when added to CEA-expressing transfectants in suspension. Intermolecular binding involving each of these subdomains is therefore essential for intercellular adhesion and cannot be compensated for by known binding contributions of other regions in the CEA molecule. In further support of this assumption, the binding epitope of an anti-CEA monoclonal antibody (monoclonal antibody A20) known to block CEA-mediated adhesion, was shown to bridge two of the three required subdomains: (42)NRQII and (30)GYSWYK. 相似文献
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17.
M Yoshida G Fuse T Matsui S Ouchi 《Biochemical and biophysical research communications》1992,188(2):794-798
It has been believed that Dictyostelium discoideum cell membranes contain no sialic acid. In this study, however, we found that contact site A, the cell adhesion molecule of D. discoideum, is a major glycoprotein containing sialic acids. This suggests that sialic acid in non-reducing terminal plays an important role in the cell adhesion in which contact site A is involved. 相似文献
18.
The effects of several cytokines and phorbol myristate acetate (PMA) on LFA-1 and ICAM-1 expression on a human eosinophilic leukemia cell line, EoL-3, were investigated and compared with those of a human monocytic leukemia cell line, U937. EoL-3 cells expressed large amounts of LFA-1 and small amounts of ICAM-1, and their expression was regulated similarly in EoL-3 cells and U937 cells. Interferon-gamma (IFN-gamma) enhanced ICAM-1 expression but not LFA-1 expression, and PMA augmented both LFA-1 and ICAM-1 expression. IFN-gamma and PMA showed an additive effect on ICAM-1 expression. These results collectively suggest that expression of LFA-1 and ICAM-1 is regulated differently and that IFN-gamma and PMA regulate the expression through different mechanisms. PMA but not IFN-gamma induced homotypic adhesion of EoL-3 and U937 cells, suggesting that PMA but not IFN-gamma activated the adhesive function of these cells. Staurosporin, an inhibitor of protein kinases (PKs), partly suppressed IFN-gamma- and PMA-augmented expression of ICAM-1 on EoL-3 and U937 cells, but did not affect PMA-augmented LFA-1 expression, suggesting that staurosporin-sensitive PKs are involved in IFN-gamma- and PMA-augmented ICAM-1 expression but not in PMA-augmented LFA-1 expression. The role of protein kinase C (PK-C) in these mechanisms was not revealed because a PK-C inhibitor, H-7, did not show any definitive effect on IFN-gamma- and PMA-induced expression of LFA-1 and ICAM-1. Moreover, cyclic AMP (cAMP)- and cGMP-dependent pathways were not shown to be involved in the augmentation of the expression of these molecules. 相似文献
19.
The G-protein-coupled receptor CIRL1/latrophilin-1 (CL1) and the type-1 membrane proteins neurexins represent distinct neuronal cell adhesion molecules that exhibit no similarities except for one common function: both proteins are receptors for α-latrotoxin, a component of black widow spider venom that induces massive neurotransmitter release at synapses. Unexpectedly, we have now identified a direct binding interaction between the extracellular domains of CL1 and neurexins that is regulated by alternative splicing of neurexins at splice site 4 (SS4). Using saturation binding assays, we showed that neurexins lacking an insert at SS4 bind to CL1 with nanomolar affinity, whereas neurexins containing an insert at SS4 are unable to bind. CL1 competed for neurexin binding with neuroligin-1, a well characterized neurexin ligand. The extracellular sequences of CL1 contain five domains (lectin, olfactomedin-like, serine/threonine-rich, hormone-binding, and G-protein-coupled receptor autoproteolysis-inducing (GAIN) domains). Of these domains, the olfactomedin-like domain mediates neurexin binding as shown by deletion mapping. Cell adhesion assays using cells expressing neurexins and CL1 revealed that their interaction produces a stable intercellular adhesion complex, indicating that their interaction can be trans-cellular. Thus, our data suggest that CL1 constitutes a novel ligand for neurexins that may be localized postsynaptically based on its well characterized interaction with intracellular SH3 and multiple ankyrin repeats adaptor proteins (SHANK) and could form a trans-synaptic complex with presynaptic neurexins. 相似文献
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