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Sample records for 2-macroglobulin receptor recognition

  1. Ligand interaction of human alpha 2-macroglobulin-alpha 2-macroglobulin receptor studied by partitioning in aqueous two-phase systems.

    PubMed

    Birkenmeier, G; Kunath, M

    1996-05-17

    Alpha 2-macroglobulin (alpha 2-M) is a major proteinase inhibitor in human blood and tissue. Besides its antiproteolytic potential, alpha 2-M was found to modulate antigen- and mitogen-driven immune responses and cell growth by binding and transporting distinct cytokines, growth factors and hormones. The inhibitor is cleared from circulation by binding to a multifunctional cellular receptor present on different cell types. Alpha 2-M, as well as its receptor, are capable of binding a variety of ligands. In the present study we have applied aqueous two-phase systems to analyze the interaction of IL-1 beta and alpha 2-M receptor to different forms of alpha 2-M. The partition of IL-1 beta was changed by addition of transformed alpha 2-M to the two-phase systems rather than by the native inhibitor. The interaction between IL-1 beta and alpha 2-M was enhanced by divalent cations. In addition, the complex formation between 125I-labelled receptor and alpha 2-M could clearly be demonstrated by partitioning. In the presence of divalent cations, transformed alpha 2-M, in contrast to the native inhibitor, effectively changed the partition of the receptor. However, the observed alteration of the partition coefficient was found to be less compared with the values obtained by partitioning of the receptor in the presence of whole plasma containing the inhibitor in equivalent concentrations. The results indicate that other components of the plasma exist which competitively bind to the receptor but independent of Ca2+-ions.

  2. The human alpha 2-macroglobulin receptor: identification of a 420-kD cell surface glycoprotein specific for the activated conformation of alpha 2-macroglobulin

    PubMed Central

    1990-01-01

    Ligand affinity chromatography was used to purify a cell surface alpha 2-macroglobulin (alpha 2M) receptor. Detergent extracts of human placenta were applied to an affinity matrix consisting of alpha 2M, previously reacted with methylamine, coupled to Sepharose. Elution with EDTA specifically released polypeptides with apparent molecular masses of 420 and 39 kD. In some preparations, small amounts of a 90-kD polypeptide were observed. The 420- and 39-kD polypeptides appear specific for the forms of alpha 2M activated by reaction with proteinases or methylamine and do not bind to an affinity matrix consisting of native alpha 2M coupled to Sepharose. Separation of these two polypeptides was accomplished by anion exchange chromatography, and binding activity was exclusively associated with the 420-kD polypeptide. The purified 420-kD protein binds to the conformationally altered forms of alpha 2M that are known to specifically interact with alpha 2M receptors and does not bind to native alpha 2M. Binding of the 420-kD polypeptide to immobilized wheat germ agglutinin indicates that this polypeptide is a glycoprotein. The cell surface localization of the 420-kD glycoprotein was confirmed by affinity chromatography of extracts from surface radioiodinated fibroblasts. These properties suggest that the 420-kD polypeptide is a cell surface receptor for the activated forms of alpha 2M. PMID:1691187

  3. Receptor-recognized alpha 2-macroglobulin-methylamine elevates intracellular calcium, inositol phosphates and cyclic AMP in murine peritoneal macrophages.

    PubMed Central

    Misra, U K; Chu, C T; Rubenstein, D S; Gawdi, G; Pizzo, S V

    1993-01-01

    Human plasma alpha 2-macroglobulin (alpha 2M) is a tetrameric proteinase inhibitor, which undergoes a conformational change upon reaction with either a proteinase or methylamine. As a result, a receptor recognition site is exposed on each subunit of the molecule enabling it to bind to its receptors on macrophages. We have used Fura-2-loaded murine peritoneal macrophages and digital video fluorescence microscopy to examine the effects of receptor binding on second messenger levels. alpha 2M-methylamine caused a rapid 2-4-fold increase in intracellular Ca2+ concentration ([Ca2+]i) within 5 s of binding to receptors. The agonists induced a focal increase in [Ca2+]i that spread out to other areas of the cell. The increase in [Ca2+]i was dependent on the alpha 2M-methylamine concentration and on the extracellular [Ca2+]. Both sinusoidal and transitory oscillations were observed, which varied from cell to cell. Neither alpha 2M nor boiled alpha 2M-methylamine, forms that are not recognized by the receptor, affected [Ca2+]i in peritoneal macrophages under identical conditions of incubation. The alpha 2M-methylamine-induced rise in [Ca2+]i was accompanied by a rapid and transient increase in macrophage inositol phosphates, including inositol tris- and tetrakis-phosphates. Native alpha 2M did not stimulate a rise in inositol phosphates. Finally, binding of alpha 2M-methylamine to macrophages increased cyclic AMP transiently. Thus receptor-recognized alpha-macroglobulins behave as agonists whose receptor binding causes stimulation of signal transduction pathways. Images Figure 2 PMID:7681282

  4. Alpha-2-macroglobulin gene, oxidized low-density lipoprotein receptor-1 locus, and sporadic Alzheimer's disease.

    PubMed

    Colacicco, Anna Maria; Solfrizzi, Vincenzo; D'Introno, Alessia; Capurso, Cristiano; Kehoe, Patrick G; Seripa, Davide; Pilotto, Alberto; Santamato, Andrea; Capurso, Antonio; Panza, Francesco

    2009-09-01

    A total sample of 169 AD patients, and 264 age- and sex-matched unrelated caregivers from Apulia, southern Italy, were genotypized for alpha-2-macroglobulin (A2M) Val1000/Ile single-nucleotide polymorphism (SNP) (rs669), apolipoprotein E (APOE), and SNPs (+1073 and +1071) in the oxidized low-density lipoprotein receptor-1 (OLR1) gene on chromosome 12. A2M allele and genotype frequencies were similar between AD patients and controls, also after stratification for late onset (>/=70 years) and early onset (<70 years) or APOE varepsilon4 status. However, there was evidence in support of LD between the OLR1+1071, the OLR1+1073, and the rs669 SNPs, with T-C-A haplotype being associated with significant increased risk of AD in both the whole sample and when we stratified according to early and late onset AD subjects, with the allelic association with AD predominantly from the OLR1+1073 SNP, further supporting the role of OLR1 as a candidate risk gene for sporadic AD.

  5. Gene transfer mediated by alpha2-macroglobulin.

    PubMed Central

    Schneider, H; Huse, K; Birkenmeier, G; Otto, A; Scholz, G H

    1996-01-01

    alpha2-Macroglobulin covalently linked to poly(L)-lysine can be used as a vehicle for receptor-mediated gene transfer. This modified alpha2-macroglobulin maintains its ability to bind to the alpha2-macroglobulin receptor, and was shown to introduce a luciferase reporter gene plasmid into HepG2 human hepatoma cells in vitro. The alpha2-macroglobulin receptor is a very large and multifunctional cell surface receptor, whose rapid and efficient internalization rate makes it attractive for gene therapy, e.g. for hepatic gene targeting via injection into the portal vein. PMID:8871570

  6. Expression of alpha 2-macroglobulin receptor/low density lipoprotein receptor-related protein and scavenger receptor in human atherosclerotic lesions.

    PubMed Central

    Luoma, J; Hiltunen, T; Särkioja, T; Moestrup, S K; Gliemann, J; Kodama, T; Nikkari, T; Ylä-Herttuala, S

    1994-01-01

    Macrophage- and smooth muscle cell (SMC)-derived foam cells are typical constituents of human atherosclerotic lesions. At least three receptor systems have been characterized that could be involved in the development of foam cells: alpha 2-macroglobulin receptor/LDL receptor-related protein (alpha 2 MR/LRP), scavenger receptor, and LDL receptor. We studied the expression of these receptors in human atherosclerotic lesions with in situ hybridization and immunocytochemistry. An abundant expression of alpha 2MR/LRP mRNA and protein was found in SMC and macrophages in both early and advanced lesions in human aortas. alpha 2MR/LRP was also present in SMC in normal aortas. Scavenger receptor mRNA and protein were expressed in lesion macrophages but no expression was found in lesion SMC. LDL receptor was absent from the lesion area but was expressed in some aortas in medial SMC located near the adventitial border. The results demonstrate that (a) alpha 2MR/LRP is, so far, the only lipoprotein receptor expressed in lesions SMC in vivo; (b) scavenger receptors are expressed only in lesion macrophages; and (c) both receptors may play important roles in the development of human atherosclerotic lesions. Images PMID:8182133

  7. Role of the scavenger receptor in the uptake of methylamine-activated alpha 2-macroglobulin by rat liver.

    PubMed Central

    van Dijk, M C; Boers, W; Linthorst, C; van Berkel, T J

    1992-01-01

    Alpha 2-Macroglobulin (alpha 2M) requires activation by small nucleophiles (e.g. methylamine; giving alpha 2M-Me) or proteolytic enzymes (e.g. trypsin; giving alpha 2M-Tr) in order to be rapidly removed from the circulation by the liver. Separation of rat liver cells into parenchymal, endothelial and Kupffer cells at 10 min after injection indicates that liver uptake of alpha 2M-Me is shared between parenchymal and endothelial cells, with relative contributions of 51.3% and 48.3% respectively of total liver-associated radioactivity. In contrast, alpha 2M-Tr is almost exclusively taken up by the parenchymal cells (90.1% of liver-associated radioactivity). A preinjection of 5 mg of poly(inosinic acid) decreased liver uptake of alpha 2M-Me to 39.9% of the control value, while it had no effect on liver uptake of alpha 2M-Tr. It appears that poly(inosinic acid) specifically reduces the uptake of alpha 2M-Me in vivo by endothelial cells, leaving uptake by parenchymal cells unaffected. In vitro studies with isolated liver cells indicate that the association of alpha 2M-Me with endothelial cells is 21-fold higher per mg of cell protein than with parenchymal cells. The capacity of endothelial cells to degrade alpha 2M-Me appears to be 46 times higher than that of parenchymal cells. Competition studies show that poly(inosinic acid) or acetylated low-density lipoprotein effectively competes with the association of alpha 2M-Me with endothelial and Kupffer cells, but association with parenchymal cells is unaffected. It is suggested that activation of alpha 2M by methylamine induces a charge distribution on the protein which triggers specific uptake by the scavenger receptor on endothelial cells. It is concluded that the uptake of alpha 2M-Me by the scavenger receptor might function as an additional system for the uptake of activated alpha 2M. Images Fig. 11. PMID:1280102

  8. Activated α2 -Macroglobulin Induces Mesenchymal Cellular Migration Of Raw264.7 Cells Through Low-Density Lipoprotein Receptor-Related Protein 1.

    PubMed

    Ferrer, Darío G; Dato, Virginia Actis; Fincati, Javier R Jaldín; Lorenc, Valeria E; Sánchez, María C; Chiabrando, Gustavo A

    2016-12-24

    Distinct modes of cell migration contribute to diverse types of cell movements. The mesenchymal mode is characterized by a multistep cycle of membrane protrusion, the formation of focal adhesion, and the stabilization at the leading edge associated with the degradation of extracellular matrix (ECM) components and with regulated extracellular proteolysis. Both α2 -Macroglobulin (α2 M) and its receptor, low density lipoprotein receptor-related protein 1 (LRP1), play important roles in inflammatory processes, by controlling the extracellular activity of several proteases. The binding of the active form of α2 M (α2 M*) to LRP1 can also activate different signaling pathways in macrophages, thus inducing extracellular matrix metalloproteinase-9 (MMP-9) activation and cellular proliferation. In the present study, we investigated whether the α2 M*/LRP1 interaction induces cellular migration of the macrophage-derived cell line, Raw264.7. By using the wound-scratch migration assay and confocal microscopy, we demonstrate that α2 M* induces LRP1-mediated mesenchymal cellular migration. This migration exhibits the production of enlarged cellular protrusions, MT1-MMP distribution to these leading edge protrusions, actin polymerization, focal adhesion formation, and increased intracellular LRP1/β1-integrin colocalization. Moreover, the presence of calphostin-C blocked the α2 M*-stimulated cellular protrusions, suggesting that the PKC activation is involved in the cellular motility of Raw264.7 cells. These findings could constitute a therapeutic target for inflammatory processes with deleterious consequences for human health, such as rheumatoid arthritis, atherosclerosis and cancer. J. Cell. Biochem. 9999: 1-9, 2017. © 2016 Wiley Periodicals, Inc.

  9. Peroxisome proliferator-activated receptor alpha (PPARalpha) agonists down-regulate alpha2-macroglobulin expression by a PPARalpha-dependent mechanism.

    EPA Science Inventory

    Peroxisome proliferator-activated receptor alpha (PPARα) regulates transcription of genes involved both in lipid and glucose metabolism as well as inflammation. Fibrates are PPARα ligands used to normalize lipid and glucose parameters and exert anti-inflammatory effects. Fibrates...

  10. Characterization of alpha-2-macroglobulin from groupers.

    PubMed

    Chuang, Wen-Hsiao; Lee, Kuo-Kau; Liu, Ping-Chung

    2013-08-01

    Alpha-2-macroglobulin (α-2-M) is a protease inhibitor broadly present in the plasma of vertebrates and invertebrates, and is an important non-specific humoral factor in defence system of the animals. This study conducted the immuno-analysis and mass spectrometric analysis methods to investigate the characteristics of the protease inhibitor, α-2-M, among groupers and related species. Rabbit antiserum to the purified α-2-M of Epinephelus coioides was used in different immunological methods to determine the immune cross-reactions of the α-2-M in samples. Plasma of Epinephelus bruneus, Epinephelus fuscoguttatus, Epinephelus lanceolatus, and Epinephelus quoyanus exhibited high protease inhibitory activities by BAPNA-trypsin assay. To purify the α-2-M protein, plasma protein of grouper E. coioides was first precipitated by using PEG 6000, then Blue Sepharose 6 Fast Flow, DEAE Sephacel, Con A Separose 4B and Phenyl Sepharose High Performance columns were used on FPLC system for purification. The molecular mass of grouper plasma α-2-M was determined as a 180 kDa protein on non-reduced SDS-PAGE. In addition, it was determined as 97 and 80 kDa protein on reduced SDS-PAGE. Enzymatic and chemical deglycosylation of glycogen revealed that the contents of glycogen in 97 and 80 kDa subunits were 12.4% and 15%, respectively, and were all belonging to N-linked type. Only one precipitation arc was visualized in all plasma of Epinephelus spp. using the rabbit antiserum to the purified α-2-M of E. coioides, on crossed immunoelectrophoresis (CIE) gels. The plasma of Epinephelus spp. and seawater fish species showed stronger responses than freshwater fish species while that of other animal species showed no response by dot-blot assay. One single band was detected on Native PAGE-Western blotting assay, one single 180 kDa band was detected on non-reduced SDS-PAGE-Western blotting, and four bands (80, 97, 160, 250 kDa) were detected on reduced SDS-PAGE when various grouper plasma

  11. [Alpha-2 macroglobulin from cerebrospinal fluid in neurosurgical diseases].

    PubMed

    Vasil'eva, T G; Dobrogorskaia, L N; Kraeva, L N; Goncharova, V P

    1989-01-01

    Content of alpha 2-macroglobulin (alpha 2-MG) was estimated in cerebrospinal fluid (CSF) of patients with neurosurgical impairments. Minimal content of the globulin was found in patients with brain concussion (0.011 +/- 0.001 g/L, control group), maximal concentration--in severe craniocerebral trauma with brain contraction (0.056 +/- 0.007 g/L) and moderately increased content of alpha 2-MG was detected in intracranial tumors and drug-resistant epilepsy, 0.028 +/- 0.004 g/L and 0.025 +/- 0.004 g/L, respectively. Alteration in content of alpha 2-MG during postoperational period corresponded to clinical state of patients. Estimation of alpha 2-MG in CSF might be used as a criterion of brain impairment severity as well as for monitoring the treatment course.

  12. Degradation of Human Fibrinogen by Plasma α2-Macroglobulin-Enzyme Complexes

    PubMed Central

    Harpel, Peter C.; Mosesson, Michael W.

    1973-01-01

    This study demonstrates that human plasma α2-macroglobulin preparations possess an enzymic activity that degrades fibrinogen, resulting in the formation of products whose structure resembles that of circulating fibrinogen catabolites. The sequence of degradation is similar to that observed in plasmin-catalyzed digests, in that Aα-chain fragmentation precedes that of Bβ-chain. The addition of plasminogen activators to plasma induced an increase in the N-α-tosyl-l-arginine methyl ester HCl esterase and fibrinogenolytic activity associated with α2-macroglobulin purified from this plasma, indicating that the enzymic activity of the complex was preserved and could be increased in the presence of other plasma enzyme inhibitors. Immunochemical studies demonstrated that an α2-macroglobulin-plasmin complex had formed in urokinase-treated plasma. This α2-macroglobulin preparation manifested an esterolytic profile like that of a complex prepared from plasmin and purified α2-macroglobulin. After complex formation with α2-macroglobulin in plasma, plasmin retained less than 0.1% of its fibrinogenolytic activity. That plasmin expressed its activity while bound to α2-macroglobulin was suggested by immunoprecipitation of this activity with α2-macroglobulin antibody and by the demonstration that pancreatic trypsin inhibitor did not effectively inhibit its fibrinogenolytic or esterolytic activity. These results raise the possibility that, in addition to its activity as a major plasma proteolytic enzyme inhibitor, α2-macroglobulin may modulate enzyme-substrate interactions, such as those resulting in the formation of circulating fibrinogen catabolites, by providing a mechanism for the preservation and protection of a portion of the enzymic activity in the presence of other circulating inhibitors. Images PMID:4269529

  13. Physical and chemical properties of human plasma alpha2-macroglobulin.

    PubMed Central

    Hall, P K; Roberts, R C

    1978-01-01

    Alpha2-M (alpha2-macroglobulin) was purified from human plasma by two different procedures. As well as having no detectable impurities by the usual criteria for testing the homogeneity of protein preparations, these alpha2M preparations showed a single component, after reduction in urea, of 185000 daltons by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. The molecular weight of the alpha2M was found to be 718000 by sedimentation equilibrium experiments using the gravimetrically determined -v of 0.731 ml/g. The interaction of several proteinases with alpha2M was studied by using a novel discontinuous polyacrylamide-gel system, which showed clear separation of the enzyme-complexed alpha2M from the free alpha2M. These studies indicated that urokinase, as well as trypsin, chymotrypsin, plasmin and thrombin forms complexes with alphaM. The cleavage of the 185000-dalton subunit to a 85000-dalton species on interaction of trypsin with alpha2M was demonstrated by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis after reduction of the alpha2M-trypsin complex in urea. The amino acid composition, carbohydrate content, absorption coefficient at 280 nm, the specific refractive increment and the sedimentation coefficient for these alpha2M preparations were measured. The stability of the trypsin-binding activity of the alpha2M preparations was also studied under several storage situations. Images Fig. 1. Fig. 2. Fig. 3. Fig. 6. PMID:80217

  14. [Expression and secretion of alpha-2-macroglobulin by dust-stimulated alveolar macrophages].

    PubMed

    Ossege, L M; Voss, B; Müller, K M

    1994-04-01

    In dust-induced, fibrosing lung processes macrophages are increased activated by foreign body reactions. The release of monokines and proteolytic enzymes, which depends on phagocytosis, may lead to destruction of the extracellular matrix with the consequence of degradation and restitution. Also the transcellular signaling or cell-matrix-interaction may finally result in development of fibrosis. However the proteolytic effect of elastase and collagenase can be inhibited by alpha-2-macroglobulin. Alpha-2-macroglobulin is a protease-inhibitor, which is synthesized by macrophages and has a wide spectrum of inhibitory abilities. Our interest was focused on observation of the production of alpha-2-macroglobulin by alveolar macrophages after stimulation with inorganic dusts of different chemical and physical properties. Rat alveolar macrophages were isolated by bronchoalveolar lavage and exposed to crocidolite, quarz or welder steam dust in vitro. The expression and secretion of alpha-2-macroglobulin was examined by non radioactive in situ hybridization, indirect immunofluorescence and radial immunodiffusion according to Mancini. The stimulated rat alveolar macrophages showed an increased expression of alpha-2-macroglobulin-mRNA and also an enhanced synthesis of alpha-2-macroglobulin-protein. Besides only small differences between the substances used for stimulation were demonstrated.

  15. Alpha-2-macroglobulin as a radioprotective agent: a review

    PubMed Central

    Chen, Xueying; Kong, Xiangbo; Zhang, Zhaoqiang; Chen, Wei; Chen, Jieyu; Li, Huanyang; Cao, Wanting; Ge, Yaping

    2014-01-01

    Radiation is an important modality in cancer treatment, and eighty percent of cancer patients need radiotherapy at some point during their clinical management. However, radiation-induced damage to normal tissues restricts the therapeutic doses of radiation that can be delivered to tumours and thereby limits the effectiveness of the treatment. The use of radioprotectors represents an obvious strategy to obtain better tumour control using a higher dose in radiotherapy. However, most of the synthetic radioprotective compounds studied have shown inadequate clinical efficacy owing to their inherent toxicity and high cost. Hence, the development of radioprotective agents with lower toxicity and an extended window of protection has attracted a great deal of attention, and the identification of alternative agents that are less toxic and highly effective is an absolute necessity. Recent studies have shown that alpha-2-macroglobulin (α2M) possesses radioprotective effects. α2M is a tetrameric, disulfide-rich plasma glycoprotein that functions as a non-selective inhibitor of different types of non-specific proteases and as a carrier of cytokines, growth factors, and hormones. α2M induces protein factors whose interplay underlies radioprotection, which supports the idea that α2M is the central effector of natural radioprotection in the rat. Pretreatment with α2M has also induced a significant reduction of irradiation-induced DNA damage and the complete restoration of liver and body weight. Mihailović et al. concluded that the radioprotection provided by α2M was in part mediated through cytoprotection of new blood cells produced in the bone marrow; these authors also indicated that an important aspect of the radioprotective effect of amifostine was the result of the induction of the endogenous cytoprotective capability of α2M. The radioprotective effects of α2M are possibly due to antioxidant, anti-fibrosis, and anti-inflammatory functions, as well as the maintenance

  16. Partial primary structure of human pregnancy zone protein: extensive sequence homology with human alpha 2-macroglobulin.

    PubMed Central

    Sottrup-Jensen, L; Folkersen, J; Kristensen, T; Tack, B F

    1984-01-01

    Human pregnancy zone protein (PZP) is a major pregnancy-associated protein. Its quaternary structure (two covalently bound 180-kDa subunits, which are further non-covalently assembled into a tetramer of 720 kDa) is similar to that of human alpha 2-macroglobulin (alpha 2M). Here we show, from the results of complete or partial sequence determination of a random selection of 38 tryptic peptides covering 685 residues of the subunit of PZP, that PZP and alpha 2M indeed are extensively homologous. In the stretches of PZP sequenced so far, the degree of identically placed residues in the two proteins is 68%, indicating a close evolutionary relationship between PZP and alpha 2M. Although the function of PZP in pregnancy is largely unknown, its close structural relationship to alpha 2M suggests analogous proteinase binding properties and a potential for being taken up in cells by receptor-mediated endocytosis. In this regard our studies indicate a bait region in PZP significantly different from that present in alpha 2M. PZP could be the human equivalent of the acute-phase alpha-macroglobulins (e.g., rat alpha 2M and rabbit alpha 1M) described earlier. PMID:6209714

  17. BKCa channel regulates calcium oscillations induced by alpha-2-macroglobulin in human myometrial smooth muscle cells

    PubMed Central

    Wakle-Prabagaran, Monali; Lorca, Ramón A.; Ma, Xiaofeng; Stamnes, Susan J.; Amazu, Chinwendu; Hsiao, Jordy J.; Hyrc, Krzysztof L.; Wright, Michael E.; England, Sarah K.

    2016-01-01

    The large-conductance, voltage-gated, calcium (Ca2+)-activated potassium channel (BKCa) plays an important role in regulating Ca2+ signaling and is implicated in the maintenance of uterine quiescence during pregnancy. We used immunopurification and mass spectrometry to identify proteins that interact with BKCa in myometrium samples from term pregnant (≥37 wk gestation) women. From this screen, we identified alpha-2-macroglobulin (α2M). We then used immunoprecipitation followed by immunoblot and the proximity ligation assay to confirm the interaction between BKCa and both α2M and its receptor, low-density lipoprotein receptor-related protein 1 (LRP1), in cultured primary human myometrial smooth muscle cells (hMSMCs). Single-channel electrophysiological recordings in the cell-attached configuration demonstrated that activated α2M (α2M*) increased the open probability of BKCa in an oscillatory pattern in hMSMCs. Furthermore, α2M* caused intracellular levels of Ca2+ to oscillate in oxytocin-primed hMSMCs. The initiation of oscillations required an interaction between α2M* and LRP1. By using Ca2+-free medium and inhibitors of various Ca2+ signaling pathways, we demonstrated that the oscillations required entry of extracellular Ca2+ through store-operated Ca2+ channels. Finally, we found that the specific BKCa blocker paxilline inhibited the oscillations, whereas the channel opener NS11021 increased the rate of these oscillations. These data demonstrate that α2M* and LRP1 modulate the BKCa channel in human myometrium and that BKCa and its immunomodulatory interacting partners regulate Ca2+ dynamics in hMSMCs during pregnancy. PMID:27044074

  18. Mapping of structure-function relationships in proteins with a panel of monoclonal antibodies. A study on human alpha 2 macroglobulin.

    PubMed

    Van Leuven, F; Marynen, P; Cassiman, J J; Van den Berghe, H

    1988-06-28

    Monoclonal antibody (Mab) F2B2, directed to the receptor-recognition site of human alpha 2 macroglobulin (alpha 2M), has been instrumental in the characterization of that site and in the isolation of the receptor-binding domain. We have now prepared a panel of Mab to study the structure-function relationships in alpha 2 M, and in particular the expression of the receptor-recognition site. Reversed dot-blotting was very effective to screen hybridoma supernatant for specificities to either the native or complex form of alpha 2M. Reaction with the isolated 20 kDa receptor-binding domain of alpha 2M and cross-reaction with pregnancy zone protein was detected by the same technique. Eventually, a panel of 45 Mab was constructed consisting of essentially five types of specificities, although in fact no two Mab reacted with complete identity in all assays. In addition to the assays already mentioned, the Mab were tested for interference with binding of alpha 2M-trypsin to the cellular receptor, for competition with F2B2 for alpha 2M-trypsin and for inhibition of trypsin by alpha 2M. Finally, Western blotting was used as a first approximate mapping of the epitope relative to the internal thiolesters by exploiting the heat-induced fragmentation of alpha 2M at this site. The five categories of Mab thus detected were: (i) five Mab that react with native alpha 2M and not with alpha 2M trypsin; (ii) 18 Mab that react with both native alpha 2M and with alpha 2M-trypsin; (iii) 12 Mab, including F2B2 and F12A3, that react with the receptor-binding domain, neo-antigenically expressed on alpha 2M-trypsin, (iv)O six Mab that are also specific for alpha 2M-trypsin but map outside the receptor-binding domain; (v) three Mab that define hidden determinants, not expressed on undenatured alpha 2M. For completeness, the panel includes the Mab obtained against pregnancy zone protein.

  19. Alpha(2)-macroglobulin levels are high in adult patients with congenital antithrombin deficiency.

    PubMed

    Tripodi, A; Chantarangkul, V; De Stefano, V; Mannucci, P

    2000-04-15

    Antithrombin is responsible for about 80% of the progressive inhibitory activity of thrombin in human plasma. The role of other protease inhibitors known to inhibit thrombin is not completely clarified. However, their contribution may become relevant when antithrombin is low. We elected to investigate adult patients with congenital antithrombin deficiency to assess the concentration of other naturally occurring thrombin inhibitors such as alpha(2)-macroglobulin, alpha(1)-antitrypsin, heparin cofactor II, and C(1)-inhibitor. The study included 59 patients with congenital antithrombin deficiency with and without a previous history of thrombosis, together with an equal number of control subjects matched for age and sex. Statistically significant differences (patients vs. controls) were observed only for alpha(2)-macroglobulin (i.e., 120 vs. 102%, p<0.01). Further analysis of antithrombin-deficient carriers with and without a past history of thrombosis showed that alpha(2)-macroglobulin levels were higher than the 90th percentile of control distribution more often in asymptomatic than symptomatic men (odds ratio=0.04; confidence interval=0.003-0.60), but not in women (odds ratio=2.14; confidence interval=0.35-13.1). In conclusion, results from this cross sectional study showed that alpha(2)-macroglobulin levels were high in patients with congenital antithrombin deficiency. Furthermore, the high levels were found more often in asymptomatic than symptomatic men. Whether this increase provides protection against thrombosis should be evaluated in a prospective study.

  20. Albumin bound and alpha 2-macroglobulin bound zinc concentrations in the sera of healthy adults.

    PubMed

    Foote, J W; Delves, H T

    1984-09-01

    Reference ranges for albumin bound and alpha 2-macroglobulin bound zinc concentrations have been determined in a study of sera obtained from 134 healthy adults. The concentrations of zinc bound to alpha 2-macroglobulin were remarkably constant with a mean (+/-SD) of 2.4 +/- 0.6 mumol/l; the variations in total serum zinc concentrations were almost entirely accounted for by variations in the zinc associated with albumin. There were no sex related differences in the transport of zinc in serum; neither was this sensitive to the use of oral contraceptives. These data provide a baseline for further investigations into the effects of zinc deficiency on the serum transport of the metal.

  1. Selective precipitation of haptoglobin and alpha2-macroglobulin from human serum using Alocasia macrorhiza tuber protein.

    PubMed

    Nayak, B Shivananda; Ulloor, N Jagadish; Shivaraj, B

    2002-12-01

    Treatment of human serum with ammonium sulfate fraction (0-50%) of Alocasia macrorhiza tuber extract resulted in precipitation at neutral pH. The precipitate was dissolved at pH 10.5 and chromatographed on Sephadex G-100 column. Two protein peaks were resolved. While the first peak represented alpha2-macroglobulin and haptoglobin, the second peak accounted for specific Alocasia protein. Incidentally the Alocasia protein was shown to be responsible for selective and specific precipitation of alpha2-macroglobulin and haptoglobin from serum. Thus the plant protein in its pure form or in crude stage could be used for the rapid isolation of two of the prominent alpha2-globulins.

  2. Structure of protease-cleaved Escherichia coli α-2-macroglobulin reveals a putative mechanism of conformational activation for protease entrapment

    PubMed Central

    Fyfe, Cameron D.; Grinter, Rhys; Josts, Inokentijs; Mosbahi, Khedidja; Roszak, Aleksander W.; Cogdell, Richard J.; Wall, Daniel M.; Burchmore, Richard J. S.; Byron, Olwyn; Walker, Daniel

    2015-01-01

    Bacterial α-2-macroglobulins have been suggested to function in defence as broad-spectrum inhibitors of host proteases that breach the outer membrane. Here, the X-ray structure of protease-cleaved Escherichia coli α-2-macroglobulin is described, which reveals a putative mechanism of activation and conformational change essential for protease inhibition. In this competitive mechanism, protease cleavage of the bait-region domain results in the untethering of an intrinsically disordered region of this domain which disrupts native interdomain interactions that maintain E. coli α-2-macroglobulin in the inactivated form. The resulting global conformational change results in entrapment of the protease and activation of the thioester bond that covalently links to the attacking protease. Owing to the similarity in structure and domain architecture of Escherichia coli α-2-macroglobulin and human α-2-macro­globulin, this protease-activation mechanism is likely to operate across the diverse members of this group. PMID:26143919

  3. α-2-Macroglobulin in Saliva Is Associated with Glycemic Control in Patients with Type 2 Diabetes Mellitus

    PubMed Central

    Aitken, Juan Pablo; Ortiz, Carolina; Morales-Bozo, Irene; Rojas-Alcayaga, Gonzalo; Baeza, Mauricio; Beltran, Caroll

    2015-01-01

    Background. Subjects with type 2 diabetes mellitus (DM2) require an adequate glycemic control to avoid diabetic complications. Currently, saliva biomarkers are used as a diagnostic tool and can be indicative of the degree of progression and control of various diseases. Several studies indicate that α-2-macroglobulin levels are elevated in diabetic patients. Methods. 120 subjects with DM2 were enrolled and classified into two groups according to their glycemic control (percentage of glycated hemoglobin-A1c (HbA1c), <7% adequate glycemic control group; >7% inadequate glycemic control group). The relationship between α-2-macroglobulin levels from saliva samples and HbA1c was subsequently evaluated. Results. We found a positive correlation between α-2-macroglobulin and HbA1c (r = 0.778 and P < 0.0001). Area under the receivers operating characteristic (ROC) curve of α-2-macroglobulin indicated a positive discrimination threshold of α-2-macroglobulin (AUC = 0.903, CI 95%: 0.847–0.959, P < 0.0001) to diagnose glycemic control. Conclusions. Our data strongly suggest that the level of saliva α-2-macroglobulin is an indicator for the degree of glycemic control in diabetic patients and represents a promising alternative method to evaluate this parameter. PMID:25821337

  4. Structure of protease-cleaved Escherichia coli α-2-macroglobulin reveals a putative mechanism of conformational activation for protease entrapment

    SciTech Connect

    Fyfe, Cameron D.; Grinter, Rhys; Josts, Inokentijs; Mosbahi, Khedidja; Roszak, Aleksander W.; Cogdell, Richard J.; Wall, Daniel M.; Burchmore, Richard J. S.; Byron, Olwyn; Walker, Daniel

    2015-06-30

    The X-ray structure of protease-cleaved E. coli α-2-macroglobulin is described, which reveals a putative mechanism of activation and conformational change essential for protease inhibition. Bacterial α-2-macroglobulins have been suggested to function in defence as broad-spectrum inhibitors of host proteases that breach the outer membrane. Here, the X-ray structure of protease-cleaved Escherichia coli α-2-macroglobulin is described, which reveals a putative mechanism of activation and conformational change essential for protease inhibition. In this competitive mechanism, protease cleavage of the bait-region domain results in the untethering of an intrinsically disordered region of this domain which disrupts native interdomain interactions that maintain E. coli α-2-macroglobulin in the inactivated form. The resulting global conformational change results in entrapment of the protease and activation of the thioester bond that covalently links to the attacking protease. Owing to the similarity in structure and domain architecture of Escherichia coli α-2-macroglobulin and human α-2-macroglobulin, this protease-activation mechanism is likely to operate across the diverse members of this group.

  5. Effects of inhaled histamine, methacholine and capsaicin on sputum levels of alpha 2-macroglobulin

    PubMed Central

    Halldorsdottir, H.; Greiff, L.; Wollmer, P.; Andersson, M.; Svensson, C.; Alkner, U.; Persson, C. G.

    1997-01-01

    BACKGROUND: Plasma exudation-derived proteins and peptides contribute significantly to inflammation in the airway mucosa in vivo. In the guinea pig trachea both histamine and the neurogenic stimulant capsaicin produce acute mucosal tissue distribution and luminal entry of bulk plasma, whereas cholinergic agonists fail to produce this effect. Of these agents, only histamine induces mucosal exudation of plasma in human nasal airways. The exudative effect of the above agents on human bronchi remains unknown. METHODS: The bronchial exudative responses to inhalation of histamine, methacholine, and capsaicin were examined in two groups of healthy volunteers. Sputum was induced on three occasions in each study group by inhalation of hypertonic saline (4.5%) given as an aerosol for 40 minutes using an ultrasonic nebuliser. The second and third occasions were preceded by histamine and capsaicin challenges in the first study group, and by histamine and methacholine challenges in the second study group. Histamine and methacholine were given in cumulative doses (total doses 3160 micrograms, respectively) or until a 20% reduction in forced expiratory volume in one second (FEV1) was achieved. Cumulative doses of capsaicin were inhaled until coughing prevented the subjects from drawing a full breath. Sputum levels of alpha 2-macroglobulin (729 kDa) were measured as an index of mucosal exudation of bulk plasma. RESULTS: Histamine increased mean (SE) sputum levels of alpha 2-macroglobulin from 2.72 (1.01) micrograms/ml (95% confidence interval (CI) 0.49 to 4.94) to 18.38 (8.03) micrograms/ml (95% CI 0.49 to 36.27) in the first group, and from 1.66 (0.84) micrograms/ml (95% CI -0.18 to 3.49) to 9.43 (3.63) micrograms/ml (95% CI 1.59 to 17.27) in the second group. In contrast, capsaicin evoked no exudation (sputum levels of alpha 2- macroglobulin 1.21 (0.28) micrograms/ml (95% CI 0.59 to 1.83)) and methacholine produced a minor increase in sputum levels of alpha 2- macroglobulin (2

  6. FTIR spectroscopy and sequence prediction: Structure of human α2-macroglobulin

    NASA Astrophysics Data System (ADS)

    Dukor, Rina K.; Liebman, Michael N.; Yuan, Anna I.; Feinman, Richard D.

    1998-06-01

    The structure of a plasma proteinase inhibitor α2-Macroglobulin (α2m) is determined by FTIR spectroscopy and a number of sequence-structure prediction algorithms. In addition, α2M dimers and complexes with methylamine and trypsin are examined. Our FTIR results estimate a helix content of 5-15% and a β-sheet content of 28-36%. None of the sequence prediction algorithms used in this study predicted values close to experimental data. Considerable differences in the FTIR spectra of α2M dimer are observed and somewhat smaller changes are seen upon reaction of α2M with methylamine and dithiodipyridine (DTP).

  7. [Alpha 1-antitrypsin, alpha 2-macroglobulin and reactive protein C in gastric cancer].

    PubMed

    Dumitraşcu, D; Radu, D; Stanciu, L; Ioniţă, A; Petcovici, M

    1989-01-01

    Proteins of acute phase: alpha 1-antitrypsin (AAT), alpha 2-macroglobulin (AMG), reactive C protein (RCP) were determined in the serum of 50 patients with gastric cancer. The Mancini, simple radial immunodiffusion method was used (SRID). The concentration of these proteins increased at 32/50 (64%) for AAT; 30/50 (60%) for AMG and 33/50 (66%) for RCP. By cumulative evaluation, the positivity of serum markers increased to 88%. The importance of differential diagnosis with regard to the benign gastric lesions (adenoma, ulcer, segmentary fibrosis, before receiving the bioptic result, is emphasized.

  8. [Levels of alpha-2-macroglobulin in blood serum of women giving birth to hypotrophic and eutrophic newborns].

    PubMed

    Wasiluk, A; Dabrowska, M; Jaworski, S; Prokopowicz, J

    2000-01-01

    We tested the concentration of alpha-2-macroglobulin in sera of 33 women bearing eutrophic newborns, 36 women bearing hypotrophic newborns and 30 healthy not pregnant women in reproductive age. The concentration of this inhibitor was measured using radial immunodiffusion method according to Mancini et al. We found distinct decrease of alpha-2-macroglobulin concentration in sera of bearing women. In women bearing eutrophic newborns we found 154 mg/dl, in women bearing hypo-trophic newborns 171 mg/dl whereas in controls 250 mg/dl. We have noted statistically significant differences between tested groups to controls and between investigated groups. Taking under consideration the role of alpha-2-macroglobulin as the modulator of immune system as well as the activity of several cytokins, therefore one can suppose that alpha-2-M may affect on cellular growth developed foetus in intrauterine.

  9. Evolution of alpha 2-macroglobulin. The demonstration in a variety of vertebrate species of a protein resembling human alpha 2-macroglobulin.

    PubMed Central

    Starkey, P M; Barrett, A J

    1982-01-01

    Plasma or serum samples from a large number of vertebrate species were screened for the presence of a papain-binding protein resembling human alph a 2-macroglobulin (alpha 2M). The screening method depended on the unique property of alpha 2M of binding proteinases in such a way that the enzyme retains partial activity against low-molecular-weight substrates. A papain-binding protein was detected in serum from members of all the major vertebrate taxa. In mammals, birds, reptiles and amphibians the protein had an Mr similar to that of human alpha 2M (725 000), but in fish, including dipnoans, actinopterygians, elasmobranchs and cyclostomes, the papain-binding protein was of Mr about 360 000. Of the invertebrate species tested, all of which were arthropods, two were negative, but the horseshoe crab, an arachnid, did possess a papain-binding protein, although this was heterogeneous in electrophoresis and differed from alpha 2M in resisting inactivation by methylamine. From the results, and a detailed study of the properties of the fish papain-binding protein described in an accompanying paper [Starkey, Fletcher & Barrett (1982) Biochem. J. 205, 97-104], it seems that alpha 2M first appeared in an ancestor of all modern vertebrates as a protein of Mr 360 000 and that the larger macroglobulin (Mr 725 000) first appeared in an ancestor of the tetrapods. Images Fig. 1. PMID:6181778

  10. Molecular cloning of α-2-macroglobulin from hemocytes of common periwinkle Littorina littorea.

    PubMed

    Borisova, Elena A; Gorbushin, Alexander M

    2014-08-01

    We report the sequence of the proteinase inhibitor with a wide inhibition spectrum, α-2-macroglobulin (α2M), belonging to the thioester superfamily of proteins. This is the first α2M sequence from coenogastropod prosobranch snails. The full-length cDNA was cloned by RACE method, spans 7897 bp and contains an open reading frame of 5460 bp. The ORF encodes a protein of 1819 amino acids. The deduced mature protein contains 1795 amino acids with a molecular weight of 200 kDa and isoelectric point of 5.00. Littorina littorea α2M bears 4 conserved α2M domains and one internal thioester. Phylogenetic analysis showed that the sequence forms well supported cluster with Mollusca species and other representatives of Lophotrochozoa.

  11. Immunodetection of hemocytes, peneidins and α2-macroglobulin in the lymphoid organ of white spot syndrome virus infected shrimp.

    PubMed

    Rodríguez, Jenny; Ruiz, Julio; Maldonado, Martha; Echeverría, Fabrizio

    2012-08-01

    Viral diseases restrict the development of the world shrimp industry and there are few studies on cell response to the presence of viral infections. We performed immunohistochemistry assays to characterize hemocytes subpopulations involved in the immune process occurring in the LO of Litopenaeus vannamei shrimp. Tissue sections of animals that increased their LO spheroids and hemocytes infiltration after WSSV induced infection, were used. Three MABs namely, 40E10 (recognizing small granule hemocytes), 40E2 (recognizing large granule hemocytes), and 41B12, which recognize α(2)-macroglobulin were used. Additionally one polyclonal antibody was used against the penaeidins antimicrobial peptides, and to detect WSSV a commercial immunohistochemistry kit (DiagXotics) was used. Numerous small granule hemocytes were detected in the stromal matrix of LO tubules, whereas large granule hemocytes were less numerous and located mainly in hemal sinuses. The exocytosis of two molecules, which have been related to the phagocytosis process, i.e. penaeidins, and α(2)-macroglobulin, was detected in the external stromal matrix and the outer tubule walls. α(2) -macroglobulin inhibits phenoloxidase activity and its strong release in LO tissue may explain the absence of melanization in the immune processes occurring in it. The immunolabeling of vesicles within the LO spheroids with MABs 41B12 40E10 and antipenaedin antibody suggests that LOS are formed by phagocytic cells derived from small granule and hyaline hemocytes, with a possible role of peneidins and α(2)-macroglobulin acting as opsonines.

  12. Pattern recognition receptors in antifungal immunity.

    PubMed

    Plato, Anthony; Hardison, Sarah E; Brown, Gordon D

    2015-03-01

    Receptors of the innate immune system are the first line of defence against infection, being able to recognise and initiate an inflammatory response to invading microorganisms. The Toll-like (TLR), NOD-like (NLR), RIG-I-like (RLR) and C-type lectin-like receptors (CLR) are four receptor families that contribute to the recognition of a vast range of species, including fungi. Many of these pattern recognition receptors (PRRs) are able to initiate innate immunity and polarise adaptive responses upon the recognition of fungal cell wall components and other conserved molecular patterns, including fungal nucleic acids. These receptors induce effective mechanisms of fungal clearance in normal hosts, but medical interventions, immunosuppression or genetic predisposition can lead to susceptibility to fungal infections. In this review, we highlight the importance of PRRs in fungal infection, specifically CLRs, which are the major PRR involved. We will describe specific PRRs in detail, the importance of receptor collaboration in fungal recognition and clearance, and describe how genetic aberrations in PRRs can contribute to disease pathology.

  13. Pattern recognitions receptors in immunodeficiency disorders.

    PubMed

    Mortaz, Esameil; Adcock, Ian M; Tabarsi, Payam; Darazam, Ilad Alavi; Movassaghi, Masoud; Garssen, Johan; Jamaati, Hamidreza; Velayati, Aliakbar

    2017-01-14

    Pattern recognition receptors (PRRs) recognize common microbial or host-derived macromolecules and have important roles in early activation and response of the immune system. Initiation of the innate immune response starts with the recognition of microbial structures called pathogen associated molecular patterns (PAMPs). Recognition of PAMPs is performed by germline-encoded receptors expressed mainly on immune cells termed pattern recognition receptors (PRRs). Several classes of pattern recognition receptors (PRRs) are involved in the pathogenesis of diseases, including Toll-like receptors (TLRs), C-type lectin receptors (CLRs), and Nod-like receptors (NLRs). Patients with primary immune deficiencies (PIDs) affecting TLR signaling can elucidate the importance of these proteins in the human immune system. Defects in interleukin-1 receptor-associated kinase-4 and myeloid differentiation factor 88 (MyD88) lead to susceptibility to infections with bacteria, while mutations in nuclear factor-κB essential modulator (NEMO) and other downstream mediators generally induce broader susceptibility to bacteria, viruses, and fungi. In contrast, TLR3 signaling defects are associated with susceptibility to herpes simplex virus type 1 encephalitis. Other PIDs induce functional alterations of TLR signaling pathways, such as common variable immunodeficiency in which plasmacytoid dendritic cell defects enhance defective responses of B cells to shared TLR agonists. Altered TLR responses to TLR2 and 4 agonists are seen in chronic granulomatous disease (CGD) and X-linked agammaglobulinemia (XLA). Enhanced TLR responses, meanwhile, are seen for TLRs 5 and 9 in CGD, TLRs 4, 7/8, and 9 in XLA, TLRs 2 and 4 in hyper IgE syndrome (HIES), and for most TLRs in adenosine deaminase deficiency. In this review we provide the reader with an update on the role of TLRs and downstream signaling pathways in PID disorders.

  14. Alpha-2-macroglobulin loaded microcapsules enhance human leukocyte functions and innate immune response

    PubMed Central

    Canova, Donata Federici; Pavlov, Anton M.; Norling, Lucy V.; Gobbetti, Thomas; Brunelleschi, Sandra; Le Fauder, Pauline; Cenac, Nicolas; Sukhorukov, Gleb B.; Perretti, Mauro

    2015-01-01

    Synthetic microstructures can be engineered to deliver bioactive compounds impacting on their pharmacokinetics and pharmacodynamics. Herein, we applied dextran-based layer-by-layer (LbL) microcapsules to deliver alpha-2-macroglobulin (α2MG), a protein with modulatory properties in inflammation. Extending recent observations made with dextran-microcapsules loaded with α2MG in experimental sepsis, we focused on the physical and chemical characteristics of these microstructures and determined their biology on rodent and human cells. We report an efficient encapsulation of α2MG into microcapsules, which enhanced i) human leukocyte recruitment to inflamed endothelium and ii) human macrophage phagocytosis: in both settings microcapsules were more effective than soluble α2MG or empty microcapsules (devoid of active protein). Translation of these findings revealed that intravenous administration of α2MG-microcapsules (but not empty microcapsules) promoted neutrophil migration into peritoneal exudates and augmented macrophage phagocytic functions, the latter response being associated with alteration of bioactive lipid mediators as assessed by mass spectrometry. The present study indicates that microencapsulation can be an effective strategy to harness the complex biology of α2MG with enhancing outcomes on fundamental processes of the innate immune response paving the way to potential future development in the control of sepsis. PMID:26385167

  15. Role of alpha-2-macroglobulin and bacterial elastase in guinea-pig pseudomonal septic shock.

    PubMed Central

    Khan, M. M.; Shibuya, Y.; Kambara, T.; Yamamoto, T.

    1995-01-01

    An essential role of alpha-2-macroglobulin (alpha 2M) was revealed in the prevention of septic shock induced in guinea-pigs by an elastase producing strain (IFO-3455) of Pseudomonas aeruginosa. When bacterial peritonitis was induced by inoculating fibrin-thrombin clot containing viable bacteria at a dose of 10(9) c.f.u./kg body weight, the guinea-pigs (n = 6) died within 7-8 hours due to septic shock. Prior to the shock, consumption of two-thirds of the circulating alpha 2M was observed. When circulating alpha 2M was depleted 4 hours after the bacterial inoculation, the guinea-pigs immediately developed shock and died within one hour. This shock was prevented either with a specific elastase inhibitor, HONHCOCH(CH2C6H5)CO-Ala-Gly-NH2, zincov (6 microM), or with human alpha 2M. Simultaneous depletion of circulating Hageman factor also prevented shock in the alpha 2M-depleted animals. These results indicate that septic shock was induced through activation of the Hageman factor dependent system by the bacteria-produced elastase which survived alpha 2M in the circulation. PMID:7537522

  16. Normal and pathological serum levels of alpha2-macroglobulins in men and mice.

    PubMed Central

    Tunstall, A M; Merriman, J M; Milne, I; James, K

    1975-01-01

    The serum levels of alpha2-macroglobulin have been measured in normal men and mice and in a number of immunopathological conditions. Normal human concentrations are high in youth, reach their minimum in middle age, and gradually increase with old age. In all age groups the mean is higher in the female than in the male. Conversely, in normal mice the alpha2M level is low in youth, maximum in middle age, and shows a slight depression with old age, and the levels are frequently higher in males than in females; there are also strain variations. In human immunopathological conditions, there are some deviations from the normal alpha2M level but these are seen to be changes from the normal distribution of values around the mean, rather than significant elevation or depression of mean values. In some disease states studied there are differences between the sexes in the deviation from normal. "Abnormal" strains of mice had alpha2M levels within the range exhibited by "normal" mice but changes in the levels are seen in mice with various myelomas. Images PMID:47865

  17. Inhibition of Aeromonas sobria serine protease (ASP) by α2-macroglobulin.

    PubMed

    Murakami, Yoji; Wada, Yoshihiro; Kobayashi, Hidetomo; Irie, Atsushi; Hasegawa, Makoto; Yamanaka, Hiroyasu; Okamoto, Keinosuke; Eto, Masatoshi; Imamura, Takahisa

    2012-10-01

    ASP is a serine protease secreted by Aeromonas sobria. ASP cleaves various plasma proteins, which is associated with onset of sepsis complications, such as shock and blood coagulation disorder. To investigate a host defense mechanism against this virulence factor, we examined the plasma for ASP inhibitor(s). Human plasma inhibited ASP activity for azocasein, which was almost completely abolished by treating plasma with methylamine, which inactivates α2-macroglobulin (α2-MG). The ASP-inhibitor complex in ASP-added plasma was not detected by immunoblotting using anti-ASP antibody; however, using gel filtration of the plasma ASP activity for an oligopeptide, the ASP substrate was eluted in the void fraction (Mw>200 000), suggesting ASP trapping by α2-MG. Indeed, human α2-MG inhibited ASP azocaseinolytic activity in a dose-dependent manner, rapidly forming a complex with the ASP. Fibrinogen degradation by ASP was completely inhibited in the presence of α2-MG. α1-Protease inhibitor, antithrombin, and α2-plasmin inhibitor neither inhibited ASP activity nor formed a complex with ASP. Surprisingly, ASP degraded these plasma serine protease inhibitors. Thus, α2-MG is the major ASP inhibitor in the human plasma and can limit ASP virulence activities in A. sobria infection sites. However, as shown by fluorescence correlation spectroscopy, slow ASP inhibition by α2-MG in plasma may indicate insufficient ASP control in vivo.

  18. Increased alpha 2-macroglobulin in diabetes: a hyperglycemia related phenomenon associated with reduced antithrombin III activity.

    PubMed

    Ceriello, A; Giugliano, D; Quatraro, A; Stante, A; Dello Russo, P; Torella, R

    1989-01-01

    Increased alpha 2-macroglobulin (alpha 2M) activity and concentration, and decreased antithrombin III (ATIII) plasma concentration are reported in diabetic subjects. In diabetes an inverse correlation between ATIII activity and blood glucose, HbA1, alpha 2M activity and alpha 2M concentration, and a direct correlation between both alpha 2M activity and alpha 2M concentration with blood glucose and HbA1 are found. Moreover, a direct correlation between alpha 2M activity and alpha 2M concentration fails. In both diabetic and normal subjects induced hyperglycemia increases alpha 2M activity and alpha 2M concentration reduces ATIII activity, while ATIII concentration is not affected. These data which show that hyperglycemia may increase alpha 2M molecule levels while altering only the biological function of ATIII, provide evidence that hyperglycemia may decrease, directly, the biological function of some proteins and may condition the levels of some risk factors for the development of diabetic complications such as alpha 2M.

  19. Activated α2-Macroglobulin Binding to Human Prostate Cancer Cells Triggers Insulin-like Responses

    PubMed Central

    Misra, Uma Kant; Pizzo, Salvatore Vincent

    2015-01-01

    Ligation of cell surface GRP78 by activated α2-macroglobulin (α2M*) promotes cell proliferation and suppresses apoptosis. α2M*-treated human prostate cancer cells exhibit a 2–3-fold increase in glucose uptake and lactate secretion, an effect similar to insulin treatment. In both α2M* and insulin-treated cells, the mRNA levels of SREBP1-c, SREBP2, fatty-acid synthase, acetyl-CoA carboxylase, ATP citrate lyase, and Glut-1 were significantly increased together with their protein levels, except for SREBP2. Pretreatment of cells with α2M* antagonist antibody directed against the carboxyl-terminal domain of GRP78 blocks these α2M*-mediated effects, and silencing GRP78 expression by RNAi inhibits up-regulation of ATP citrate lyase and fatty-acid synthase. α2M* induces a 2–3-fold increase in lipogenesis as determined by 6-[14C]glucose or 1-[14C]acetate incorporation into free cholesterol, cholesterol esters, triglycerides, free fatty acids, and phosphatidylcholine, which is blocked by inhibitors of fatty-acid synthase, PI 3-kinase, mTORC, or an antibody against the carboxyl-terminal domain of GRP78. We also assessed the incorporation of [14CH3]choline into phosphatidylcholine and observed similar effects. Lipogenesis is significantly affected by pretreatment of prostate cancer cells with fatostatin A, which blocks sterol regulatory element-binding protein proteolytic cleavage and activation. This study demonstrates that α2M* functions as a growth factor, leading to proliferation of prostate cancer cells by promoting insulin-like responses. An antibody against the carboxyl-terminal domain of GRP78 may have important applications in prostate cancer therapy. PMID:25720493

  20. Recognition of lactoferrin and aminopeptidase M-modified lactoferrin by the liver: involvement of proteoglycans and the remnant receptor.

    PubMed Central

    Ziere, G J; Kruijt, J K; Bijsterbosch, M K; van Berkel, T J

    1996-01-01

    1. Lactoferrin and aminopeptidase M-modified lactoferrin (APM-lactoferrin; which lacks its 14 N-terminal amino acids) inhibit the liver uptake of lipoprotein remnant. In the present study, the role of proteoglycans in the initial interaction of beta-migrating very-low-density lipoprotein (beta-VLDL), native and APM-lactoferrin with isolated rat parenchymal liver cells was investigated. Treatment of the cells with chondroitinase lowered the Kd of lactoferrin binding (from 10 to 2.4 microM), and the number of sites/cell (from 20 x 10(6) to 7 x 10(6)), while heparinase treatment did not affect the binding. The binding characteristics of APM-lactoferrin and beta-VLDL were not altered by treatment of the cells with chondroitinase or heparinase. It is concluded that proteoglycans are not involved in the initial binding of APM-lactoferrin and beta-VLDL to parenchymal cells, while chondroitin sulphate proteoglycans are mainly responsible for the massive, low-affinity binding of native lactoferrin..2. The binding of lactoferrin, APM-lactoferrin and beta-VLDL to parenchymal liver cells was not influenced by the glutathione S-transferase-receptor-associated protein (GST-RAP) (97.2% +/- 4.0%, 95.5 +/- 3.7% and 98.5% of the control binding), while the binding of alpha 2-macroglobulin was fully blocked at 10 micrograms/ml GST-RAP (1.8 +/- 0.5% of the control binding). Since GST-RAP blocks the binding of all the known ligands to the low-density lipoprotein (LDL)-receptor-related protein (LRP), it is concluded that LRP is not the initial primary recognition site for lactoferrin, APM-lactoferrin and beta-VLDL on parenchymal liver cells. 3. We showed earlier that.APM-lactoferrin, as compared with lactoferrin, is a more effective inhibitor of the liver uptake of lipoprotein remnants (49.4 +/- 4.0% versus 80.8 +/- 4.8% of the control at 500 micrograms/ml respectively). We found in the present study that beta-VLDL is able to inhibit the binding of APM-lactoferrin to parenchymal liver

  1. Hyperglycemia-conditioned increase in alpha-2-macroglobulin in healthy normal subjects: a phenomenon correlated with deficient antithrombin III activity.

    PubMed

    Ceriello, A; Quatraro, A; Dello Russo, P; Marchi, E; Barbanti, M; Giugliano, D

    1989-01-01

    Induced hyperglycemia in normal subjects increases alpha 2-macroglobulin (alpha 2M) activity and alpha 2M concentration and reduces antithrombin III (ATIII) activity, while it does not affect ATIII plasma concentration. Hyperglycemia-determined variations in ATIII activity and alpha 2M molecules are correlated in an inverse and parallel fashion. A compensatory role for the increase in alpha 2M in the regulation of the coagulation system may be hypothesized. Moreover, these data provide evidence that hyperglycemia may decrease, directly, the biological function of some proteins and may influence the levels of some risk factors for the development of complications in diabetes.

  2. Pattern recognition receptors in microbial keratitis

    PubMed Central

    Taube, M-A; del Mar Cendra, M; Elsahn, A; Christodoulides, M; Hossain, P

    2015-01-01

    Microbial keratitis is a significant cause of global visual impairment and blindness. Corneal infection can be caused by a wide variety of pathogens, each of which exhibits a range of mechanisms by which the immune system is activated. The complexity of the immune response to corneal infection is only now beginning to be elucidated. Crucial to the cornea's defences are the pattern-recognition receptors: Toll-like and Nod-like receptors and the subsequent activation of inflammatory pathways. These inflammatory pathways include the inflammasome and can lead to significant tissue destruction and corneal damage, with the potential for resultant blindness. Understanding the immune mechanisms behind this tissue destruction may enable improved identification of therapeutic targets to aid development of more specific therapies for reducing corneal damage in infectious keratitis. This review summarises current knowledge of pattern-recognition receptors and their downstream pathways in response to the major keratitis-causing organisms and alludes to potential therapeutic approaches that could alleviate corneal blindness. PMID:26160532

  3. An α2-macroglobulin-like protein is the cue to gregarious settlement of the barnacle Balanus amphitrite

    PubMed Central

    Dreanno, Catherine; Matsumura, Kiyotaka; Dohmae, Naoshi; Takio, Koji; Hirota, Hiroshi; Kirby, Richard R.; Clare, Anthony S.

    2006-01-01

    Many benthic marine invertebrates, like barnacles, have a planktonic larval stage whose primary purpose is dispersal. How these species colonize suitable substrata is fundamental to understanding their evolution, population biology, and wider community dynamics. Unlike larval dispersal, settlement occurs on a relatively small spatial scale and involves larval behavior in response to physical and chemical characteristics of the substratum. Biogenic chemical cues have been implicated in this process. Their identification, however, has proven challenging, no more so than for the chemical basis of barnacle gregariousness, which was first described >50 years ago. We now report that a biological cue to gregarious settlement, the settlement-inducing protein complex (SIPC), of the major fouling barnacle Balanus amphitrite is a previously undescribed glycoprotein. The SIPC shares a 30% sequence homology with the thioester-containing family of proteins that includes the α2-macroglobulins. The cDNA (5.2 kb) of the SIPC encodes a protein precursor comprising 1,547 aa with a 17-residue signal peptide region. A number of structural characteristics and the absence of a thioester bond in the SIPC suggest that this molecule is a previously undescribed protein that may have evolved by duplication from an ancestral α2-macroglobulin gene. Although the SIPC is regarded as an adult cue that is recognized by the cyprid at settlement, it is also expressed in the juvenile and in larvae, where it may function in larva–larva settlement interactions. PMID:16983086

  4. The electrophoretically 'slow' and 'fast' forms of the alpha 2-macroglobulin molecule.

    PubMed Central

    Barrett, A J; Brown, M A; Sayers, C A

    1979-01-01

    alpha 2-Macroglobulin (alpha 2M) was isolated from human plasma by a four-step procedure: poly(ethylene glyco) fractionation, gel chromatography, euglobulin precipitation and immunoadsorption. No contaminants were detected in the final preparations by electrophoresis or immunoprecipitation. The protein ran as a single slow band in gel electrophoresis, and was designated 'S-alpha 2M'. S-alpha 2M bound about 2 mol of trypsin/mol. Treatment of S-alpha 2M with a proteinase or ammonium salts produced a form of the molecule more mobile in electrophoresis, and lacking proteinase-binding activity (F-alpha 2M). The electrophoretic mobility of the F-alpha 2M resulting from reaction with NH4+ salts was identical with that of proteinase complexes. We attribute the change in electrophoretic mobility of the alpha 2M to a conformation change, but there was no evidence of a change in pI or Strokes radius. Electrophoresis of S-alpha 2M in the presence of sodium dodecylsulphate gave results consistent with the view that the alpha 2M molecule is a tetramer of identical subunits, assembled as a non-covalent pair of disulphide-linked dimers. Some of the subunits seemed to be 'nicked' into two-thires-length and one-third-length chains, however. This was not apparent with F-alpha 2M produced by ammonium salts. F-alpha 2M produced by trypsin showed two new bands attributable to cleavage of the subunit polypeptide chain near the middle. Immunoassays of F-alpha 2M gave 'rockets' 12-29% lower than those with S-alpha 2M. The nature of the interactions between subunits in S-alpha 2M and F-alpha 2M was investigated by treating each form with glutaraldehyde before electrophoresis in the presence of sodium dodecyl sulphate. A much greater degree of cross-linking was observed with the F-alpha 2M, indicating that the subunits interact most closely in this form of the molecule. Exposure of S-alpha 2M to 3 M-urea or pH3 resulted in dissociation to the disulphide-bonded half-molecules; these did not

  5. Pattern-Recognition Receptors and Gastric Cancer

    PubMed Central

    Castaño-Rodríguez, Natalia; Kaakoush, Nadeem O.; Mitchell, Hazel M.

    2014-01-01

    Chronic inflammation has been associated with an increased risk of several human malignancies, a classic example being gastric adenocarcinoma (GC). Development of GC is known to result from infection of the gastric mucosa by Helicobacter pylori, which initially induces acute inflammation and, in a subset of patients, progresses over time to chronic inflammation, gastric atrophy, intestinal metaplasia, dysplasia, and finally intestinal-type GC. Germ-line encoded receptors known as pattern-recognition receptors (PRRs) are critical for generating mature pro-inflammatory cytokines that are crucial for both Th1 and Th2 responses. Given that H. pylori is initially targeted by PRRs, it is conceivable that dysfunction within genes of this arm of the immune system could modulate the host response against H. pylori infection, and subsequently influence the emergence of GC. Current evidence suggests that Toll-like receptors (TLRs) (TLR2, TLR3, TLR4, TLR5, and TLR9), nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) (NOD1, NOD2, and NLRP3), a C-type lectin receptor (DC-SIGN), and retinoic acid-inducible gene (RIG)-I-like receptors (RIG-I and MDA-5), are involved in both the recognition of H. pylori and gastric carcinogenesis. In addition, polymorphisms in genes involved in the TLR (TLR1, TLR2, TLR4, TLR5, TLR9, and CD14) and NLR (NOD1, NOD2, NLRP3, NLRP12, NLRX1, CASP1, ASC, and CARD8) signaling pathways have been shown to modulate the risk of H. pylori infection, gastric precancerous lesions, and/or GC. Further, the modulation of PRRs has been suggested to suppress H. pylori-induced inflammation and enhance GC cell apoptosis, highlighting their potential relevance in GC therapeutics. In this review, we present current advances in our understanding of the role of the TLR and NLR signaling pathways in the pathogenesis of GC, address the involvement of other recently identified PRRs in GC, and discuss the potential implications of PRRs in GC immunotherapy

  6. Receptor recognition mechanisms of coronaviruses: a decade of structural studies.

    PubMed

    Li, Fang

    2015-02-01

    Receptor recognition by viruses is the first and essential step of viral infections of host cells. It is an important determinant of viral host range and cross-species infection and a primary target for antiviral intervention. Coronaviruses recognize a variety of host receptors, infect many hosts, and are health threats to humans and animals. The receptor-binding S1 subunit of coronavirus spike proteins contains two distinctive domains, the N-terminal domain (S1-NTD) and the C-terminal domain (S1-CTD), both of which can function as receptor-binding domains (RBDs). S1-NTDs and S1-CTDs from three major coronavirus genera recognize at least four protein receptors and three sugar receptors and demonstrate a complex receptor recognition pattern. For example, highly similar coronavirus S1-CTDs within the same genus can recognize different receptors, whereas very different coronavirus S1-CTDs from different genera can recognize the same receptor. Moreover, coronavirus S1-NTDs can recognize either protein or sugar receptors. Structural studies in the past decade have elucidated many of the puzzles associated with coronavirus-receptor interactions. This article reviews the latest knowledge on the receptor recognition mechanisms of coronaviruses and discusses how coronaviruses have evolved their complex receptor recognition pattern. It also summarizes important principles that govern receptor recognition by viruses in general.

  7. Human immunodeficiency virus type 1 proteinase is rapidly and efficiently inactivated in human plasma by alpha 2-macroglobulin.

    PubMed

    Kisselev, A F; von der Helm, K

    1994-10-01

    Human plasma impairs the activity of the human immunodeficiency virus (HIV-1) proteinase to cleave the HIV-1 gag-polyprotein precursor. The inhibition is due to the entrapment of the proteinase by plasma alpha 2-macroglobulin (alpha 2M). In methylamine-treated plasma, where alpha 2M is inactivated, HIV proteinase is not blocked. The interaction of alpha 2M and HIV-1 proteinase resulting in covalent complexes of proteinase and alpha 2M was demonstrated by immunoblotting with antiserum either to alpha 2M or to the HIV proteinase. We suggest if HIV-1 proteinase would be released in vivo from infected patients' cells, alpha 2M entrapment may prevent or minimize a conceivable cleavage of extracellular matrix or plasma proteins by the HIV-1 enzyme.

  8. Macrophage pattern recognition receptors in immunity, homeostasis and self tolerance.

    PubMed

    Mukhopadhyay, Subhankar; Plüddemann, Annette; Gordon, Siamon

    2009-01-01

    Macrophages, a major component of innate immune defence, express a large repertoire of different classes of pattern recognition receptors and other surface antigens which determine the immunologic and homeostatic potential of these versatile cells. In the light of present knowledge ofmacrophage surface antigens, we discuss self versus nonself recognition, microbicidal effector functions and self tolerance in the innate immune system.

  9. A bacterial tyrosine phosphatase inhibits plant pattern recognition receptor activation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Perception of pathogen-associated molecular patterns (PAMPs) by surface-localised pattern-recognition receptors (PRRs) is a key component of plant innate immunity. Most known plant PRRs are receptor kinases and initiation of PAMP-triggered immunity (PTI) signalling requires phosphorylation of the PR...

  10. Guinea pig macroalbumin. A major inhibitor of activated Hageman factor in plasma with an alpha 2-macroglobulin-like nature.

    PubMed Central

    Ishimatsu, T.; Yamamoto, T.; Kozono, K.; Kambara, T.

    1984-01-01

    A major inhibitor of the beta form of activated Hageman factor (beta-HFa) with an apparent molecular weight of 28,000, which was reported as a strong permeability factor (Yamamoto and Cochrane, Am J Pathol 1981, 105: 164-175), was purified from guinea pig plasma. When it was depleted in vitro, the plasma lost 71% of the total inhibitory activity toward beta-HFa. The inhibitor, termed "macroalbumin," with an apparent molecular weight of 720,000 and an apparent pI of 4.6, seemed to be an inhibitor similar to alpha 1- or alpha 2-macroglobulin of man and other mammalian species in physicochemical characteristics and in enzymologic properties. Though the inhibitory activity to beta-HFa was negligible in normal skin extract, a significant inhibitory activity appeared in extracts of permeability-enhanced skin sites which were induced by intradermal beta-HFa injection. The inhibitory activity that appeared was macroalbumin-dependent, with more than a 10-fold increase in the concentration. These results indicate the roles of macroalbumin as a negative feedback regulator in situ to the Hageman-factor-dependent pathway in a permeability enhancement system. Images Figure 4 Figure 5 PMID:6201075

  11. Alpha-2-macroglobulin as the major defence in acute pseudomonal septic shock in the guinea-pig model.

    PubMed Central

    Khan, M. M.; Shibuya, Y.; Nakagaki, T.; Kambara, T.; Yamamoto, T.

    1994-01-01

    An intravenous injection of 1.2 mg/kg of Pseudomonas aeruginosa elastase induces immediate lethal shock in guinea-pigs. In the present study, alpha-2-macroglobulin (alpha 2M) was shown to be the major factor in guinea-pig plasma that inhibits the enzymatic activity of elastase in vitro. Depletion of circulating alpha 2M by injecting anti-guinea-pig alpha 2M rabbit IgG F(ab')2 rendered the animals sensitive to a dose of elastase of 0.05 mg/kg. When the alpha 2M-depleted guinea-pigs were reconstituted with human alpha 2M, this sensitivity was reversed. Lethal shock did not occur in alpha 2M-depleted animals even at an elastase dose of 0.2 mg/kg when Hageman factor was simultaneously depleted, indicating that elastase induces shock through activation of the Hageman factor-dependent system. Similar results were obtained when the culture supernatants of an elastase-producing strain, IFO-3455, were used instead of the purified elastase, whereas no cardiovascular changes occurred, even in the alpha 2M-depleted guinea-pigs, when the culture supernatants were pretreated with an elastase specific inhibitor (zincov) or when the culture supernatants of an elastase non-producing strain, PA-103 were used. PMID:7524612

  12. Alpha-2-macroglobulin functions as an inhibitor of fibrinolytic, clotting, and neutrophilic proteinases in sepsis: studies using a baboon model.

    PubMed

    de Boer, J P; Creasey, A A; Chang, A; Abbink, J J; Roem, D; Eerenberg, A J; Hack, C E; Taylor, F B

    1993-12-01

    Alpha-2-macroglobulin (alpha 2M) may function as a proteinase inhibitor in vivo. Levels of this protein are decreased in sepsis, but the reason these levels are low is unknown. Therefore, we analyzed the behavior of alpha 2M in a baboon model for sepsis. Upon challenge with a lethal (4 baboons) or a sublethal (10 baboons) dose of Escherichia coli, levels of inactivated alpha 2M (i alpha 2M) steadily increased, the changes being more pronounced in the animals that received the lethal dose. The rise in i alpha 2M significantly correlated with the increase of thrombin-antithrombin III, plasmin-alpha 2-antiplasmin, and, to a lesser extent, with that of elastase-alpha 1-antitrypsin complexes, raising the question of involvement of fibrinolytic, clotting, and neutrophilic proteinases in the inactivation of alpha 2M. Experiments with chromogenic substrates confirmed that thrombin, plasmin, elastase, and cathepsin G indeed had formed complexes with alpha 2M. Changes in alpha 2M similar to those observed in the animals that received E. coli occurred in baboons challenged with Staphylococcus aureus, indicating that alpha 2M formed complexes with the proteinases just mentioned in gram-positive sepsis as well. We conclude that alpha 2M in this baboon model for sepsis is inactivated by formation of complexes with proteinases, derived from activated neutrophils and from fibrinolytic and coagulation cascades. We suggest that similar mechanisms may account for the decreased alpha 2M levels in clinical sepsis.

  13. The rat acute-phase protein α2-macroglobulin plays a central role in amifostine-mediated radioprotection.

    PubMed

    Mirjana, Mihailović; Goran, Poznanović; Nevena, Grdović; Melita, Vidaković; Svetlana, Dinić; Ilijana, Grigorov; Desanka, Bogojević

    2010-09-01

    Previously we reported that elevated circulating concentrations of the acute-phase (AP) protein α(2)-macroglobulin (α(2)M), either as typically occurring in pregnant female rats or after administration to male rats, provides radioprotection, displayed as 100% survival of experimental animals exposed to total-body irradiation with 6.7 Gy (LD(50/30)) x-rays, that is as effective as that afforded by the synthetic radioprotector amifostine. The finding that amifostine administration induces a 45-fold increase in α(2)M in the circulation led us to hypothesise that α(2)M assumes an essential role in both natural and amifostine-mediated radioprotection in the rat. In the present work we examined the activation of cytoprotective mechanisms in rat hepatocytes after the exogenous administration of α(2)M and amifostine. Our results showed that the IL6/JAK/STAT3 hepatoprotective signal pathway, described in a variety of liver-injury models, upregulated the α(2)M gene in amifostine-pretreated animals. In both α(2)M- and amifostine-pretreated rats we observed the activation of the Akt signalling pathways that mediate cellular survival. At the cellular level this was reflected as a significant reduction of irradiation-induced DNA damage that allowed for the rapid and complete restoration of liver mass and ultimately at the level of the whole organism the complete restoration of body weight. We conclude that the selective upregulation of α(2)M plays a central role in amifostine-provided radioprotection.

  14. The structural basis for receptor recognition of human interleukin-18

    DOE PAGES

    Tsutsumi, Naotaka; Kimura, Takeshi; Arita, Kyohei; ...

    2014-12-15

    Interleukin (IL)-18 is a proinflammatory cytokine that belongs to the IL-1 family and plays an important role in inflammation. The uncontrolled release of this cytokine is associated with severe chronic inflammatory disease. IL-18 forms a signalling complex with the IL-18 receptor α (Rα) and β (Rβ) chains at the plasma membrane, which induces multiple inflammatory cytokines. Here, we present a crystal structure of human IL-18 bound to the two receptor extracellular domains. Generally, the receptors’ recognition mode for IL-18 is similar to IL-1β; however, certain notable differences were observed. The architecture of the IL-18 receptor second domain (D2) is uniquemore » among the other IL-1R family members, which presumably distinguishes them from the IL-1 receptors that exhibit a more promiscuous ligand recognition mode. The structures and associated biochemical and cellular data should aid in developing novel drugs to neutralize IL-8 activity.« less

  15. The structural basis for receptor recognition of human interleukin-18

    SciTech Connect

    Tsutsumi, Naotaka; Kimura, Takeshi; Arita, Kyohei; Ariyoshi, Mariko; Ohnishi, Hidenori; Yamamoto, Takahiro; Zuo, Xiaobing; Maenaka, Katsumi; Park, Enoch Y.; Kondo, Naomi; Shirakawa, Masahiro; Tochio, Hidehito; Kato, Zenichiro

    2014-12-15

    Interleukin (IL)-18 is a proinflammatory cytokine that belongs to the IL-1 family and plays an important role in inflammation. The uncontrolled release of this cytokine is associated with severe chronic inflammatory disease. IL-18 forms a signalling complex with the IL-18 receptor α (Rα) and β (Rβ) chains at the plasma membrane, which induces multiple inflammatory cytokines. Here, we present a crystal structure of human IL-18 bound to the two receptor extracellular domains. Generally, the receptors’ recognition mode for IL-18 is similar to IL-1β; however, certain notable differences were observed. The architecture of the IL-18 receptor second domain (D2) is unique among the other IL-1R family members, which presumably distinguishes them from the IL-1 receptors that exhibit a more promiscuous ligand recognition mode. The structures and associated biochemical and cellular data should aid in developing novel drugs to neutralize IL-8 activity.

  16. The role of pattern recognition receptors in the innate recognition of Candida albicans

    PubMed Central

    Zheng, Nan-Xin; Wang, Yan; Hu, Dan-Dan; Yan, Lan; Jiang, Yuan-Ying

    2015-01-01

    Candida albicans is both a commensal microorganism in healthy individuals and a major fungal pathogen causing high mortality in immunocompromised patients. Yeast-hypha morphological transition is a well known virulence trait of C. albicans. Host innate immunity to C. albicans critically requires pattern recognition receptors (PRRs). In this review, we summarize the PRRs involved in the recognition of C. albicans in epithelial cells, endothelial cells, and phagocytic cells separately. We figure out the differential recognition of yeasts and hyphae, the findings on PRR-deficient mice, and the discoveries on human PRR-related single nucleotide polymorphisms (SNPs). PMID:25714264

  17. Pathogen recognition or homeostasis? APC receptor functions in innate immunity.

    PubMed

    Gordon, Siamon

    2004-06-01

    Myeloid cells (macrophages, neutrophils, dendritic cells) express a repertoire of plasma membrane receptors able to recognize all classes of macromolecules. The concept of pattern recognition has emphasized microbial ligands and host defence. However, these receptors play a broader role in tissue homeostasis within multicellular hosts, clearing the extracellular environment of potential undesirable ligands arising endogenously as well as from without. This article will evaluate one of the paradigms that underlie innate immunity.

  18. The structural role of receptor tyrosine sulfation in chemokine recognition

    PubMed Central

    Ludeman, Justin P; Stone, Martin J

    2014-01-01

    Tyrosine sulfation is a post-translational modification of secreted and transmembrane proteins, including many GPCRs such as chemokine receptors. Most chemokine receptors contain several potentially sulfated tyrosine residues in their extracellular N-terminal regions, the initial binding site for chemokine ligands. Sulfation of these receptors increases chemokine binding affinity and potency. Although receptor sulfation is heterogeneous, insights into the molecular basis of sulfotyrosine (sTyr) recognition have been obtained using purified, homogeneous sulfopeptides corresponding to the N-termini of chemokine receptors. Receptor sTyr residues bind to a shallow cleft defined by the N-loop and β3-strand elements of cognate chemokines. Tyrosine sulfation enhances the affinity of receptor peptides for cognate chemokines in a manner dependent on the position of sulfation. Moreover, tyrosine sulfation can alter the selectivity of receptor peptides among several cognate chemokines for the same receptor. Finally, binding to receptor sulfopeptides can modulate the oligomerization state of chemokines, thereby influencing the ability of a chemokine to activate its receptor. These results increase the motivation to investigate the structural basis by which tyrosine sulfation modulates chemokine receptor activity and the biological consequences of this functional modulation. Linked ArticlesThis article is part of a themed section on Molecular Pharmacology of GPCRs. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2014.171.issue-5 PMID:24116930

  19. [Recognition of commensal microflora by pattern recognition receptors in human physiology and pathology].

    PubMed

    Bondarenko, V M; Likhoded, V G

    2012-01-01

    Contemporary data on the interaction of commensal microflora and Toll-like pattern recognition receptors are presented. These receptors recognize normal intestine microflora in physiological conditions, and this interaction is necessary for the maintenance of homeostasis and damage reparation of the intestine, for the induction of heat shock cytoprotective proteins. As a side effect in disruption of immunologic tolerance and misbalance of protective immunological mechanisms, multiorgan pathologic changes of organs and tissues may develop, including chronic inflammation processes of various localization.

  20. Pattern Recognition Receptors in Innate Immunity, Host Defense, and Immunopathology

    ERIC Educational Resources Information Center

    Suresh, Rahul; Mosser, David M.

    2013-01-01

    Infection by pathogenic microbes initiates a set of complex interactions between the pathogen and the host mediated by pattern recognition receptors. Innate immune responses play direct roles in host defense during the early stages of infection, and they also exert a profound influence on the generation of the adaptive immune responses that ensue.…

  1. Perirhinal Cortex Muscarinic Receptor Blockade Impairs Taste Recognition Memory Formation

    ERIC Educational Resources Information Center

    Gutierrez, Ranier; De la Cruz, Vanesa; Rodriguez-Ortiz, Carlos J.; Bermudez-Rattoni, Federico

    2004-01-01

    The relevance of perirhinal cortical cholinergic and glutamatergic neurotransmission for taste recognition memory and learned taste aversion was assessed by microinfusions of muscarinic (scopolamine), NMDA (AP-5), and AMPA (NBQX) receptor antagonists. Infusions of scopolamine, but not AP5 or NBQX, prevented the consolidation of taste recognition…

  2. The conformational changes of alpha 2-macroglobulin induced by methylamine or trypsin. Characterization by extrinsic and intrinsic spectroscopic probes.

    PubMed Central

    Larsson, L J; Lindahl, P; Hallén-Sandgren, C; Björk, I

    1987-01-01

    The conformational changes around the thioester-bond region of human or bovine alpha 2M (alpha 2-macroglobulin) on reaction with methylamine or trypsin were studied with the probe AEDANS [N-(acetylaminoethyl)-8-naphthylamine-1-sulphonic acid], bound to the liberated thiol groups. The binding affected the fluorescence emission and lifetime of the probe in a manner indicating that the thioester-bond region is partially buried in all forms of the inhibitor. In human alpha 2M these effects were greater for the trypsin-treated than for the methylamine-treated inhibitor, which both have undergone similar, major, conformational changes. This difference may thus be due to a close proximity of the thioester region to the bound proteinase. Reaction of trypsin with thiol-labelled methylamine-treated bovine alpha 2M, which retains a near-native conformation and inhibitory activity, indicated that the major conformational change accompanying the binding of proteinases involves transfer of the thioester-bond region to a more polar environment without increasing the exposure of this region at the surface of the protein. Labelling of the transglutaminase cross-linking site of human alpha 2M with dansylcadaverine [N-(5-aminopentyl)-5-dimethylaminonaphthalene-1-sulphonamide] suggested that this site is in moderately hydrophobic surroundings. Reaction of the labelled inhibitor with methylamine or trypsin produced fluorescence changes consistent with further burial of the cross-linking site. These changes were more pronounced for trypsin-treated than for methylamine-treated alpha 2M, presumably an effect of the cleavage of the adjacent 'bait' region. Solvent perturbation of the u.v. absorption and iodide quenching of the tryptophan fluorescence of human alpha 2M showed that one or two tryptophan residues in each alpha 2M monomer are buried on reaction with methylamine or trypsin, with no discernible change in the exposure of tyrosine residues. Together, these results indicate an

  3. Pharmacophore model for bile acids recognition by the FPR receptor

    NASA Astrophysics Data System (ADS)

    Ferrari, Cristina; Macchiarulo, Antonio; Costantino, Gabriele; Pellicciari, Roberto

    2006-05-01

    Formyl-peptide receptors (FPRs) belong to the family A of the G-protein coupled receptor superfamily and include three subtypes: FPR, FPR-like-1 and FPR-like-2. They have been involved in the control of␣many inflammatory processes promoting the recruitment and infiltration of leukocytes in regions of inflammation through the molecular recognition of chemotactic factors. A large number of structurally diverse chemotypes modulate the activity of FPRs. Newly identified antagonists include bile acids deoxycholic acid (DCA) and chenodeoxycholic acid (CDCA). The molecular recognition of these compounds at FPR receptor was computationally investigated using both ligand- and structure-based approaches. Our findings suggest that all antagonists bind at the first third of the seven helical bundles. A closer inspection of bile acid interaction reveals a number of unexploited anchor points in the binding site that may be used to aid the design of new potent and selective bile acids derivatives at FPR.

  4. Receptor recognition and cross-species infections of SARS coronavirus.

    PubMed

    Li, Fang

    2013-10-01

    Receptor recognition is a major determinant of the host range, cross-species infections, and pathogenesis of the severe acute respiratory syndrome coronavirus (SARS-CoV). A defined receptor-binding domain (RBD) in the SARS-CoV spike protein specifically recognizes its host receptor, angiotensin-converting enzyme 2 (ACE2). This article reviews the latest knowledge about how RBDs from different SARS-CoV strains interact with ACE2 from several animal species. Detailed research on these RBD/ACE2 interactions has established important principles on host receptor adaptations, cross-species infections, and future evolution of SARS-CoV. These principles may apply to other emerging animal viruses, including the recently emerged Middle East respiratory syndrome coronavirus (MERS-CoV). This paper forms part of a series of invited articles in Antiviral Research on "From SARS to MERS: 10years of research on highly pathogenic human coronaviruses".

  5. Gibberellin Perception by the Gibberellin Receptor and its Effector Recognition

    NASA Astrophysics Data System (ADS)

    Hakoshima, Toshio; Murase, Kohji; Hirano, Yoshinori; Sun, Tai-Ping

    Gibberellins control a diverse range of growth and developmental processes in higher plants and have been widely utilized in the agricultural industry. By binding to a nuclear receptor GIBBERELLIN INSENSITIVE DWARF1 (GID1), gibberellins regulate gene expression by promoting degradation of the transcriptional regulator DELLA proteins. The precise manner in which GID1 discriminates and becomes activated by bioactive gibberellins for specific binding to DELLA proteins remains unclear. We present the crystal structure of a ternary complex of Arabidopsis thaliana GID1A, a bioactive gibberellin and the N-terminal DELLA domain of GAI. In this complex, GID1a occludes gibberellin in a deep binding pocket covered by its N-terminal helical switch region, which in turn interacts with the DELLA domain containing DELLA, VHYNP and LExLE motifs. Our results establish a structural model of a plant hormone receptor which is distinct from the hormone-perception mechanism and effector recognition of the known auxin receptors.

  6. A bacterial tyrosine phosphatase inhibits plant pattern recognition receptor activation.

    PubMed

    Macho, Alberto P; Schwessinger, Benjamin; Ntoukakis, Vardis; Brutus, Alexandre; Segonzac, Cécile; Roy, Sonali; Kadota, Yasuhiro; Oh, Man-Ho; Sklenar, Jan; Derbyshire, Paul; Lozano-Durán, Rosa; Malinovsky, Frederikke Gro; Monaghan, Jacqueline; Menke, Frank L; Huber, Steven C; He, Sheng Yang; Zipfel, Cyril

    2014-03-28

    Innate immunity relies on the perception of pathogen-associated molecular patterns (PAMPs) by pattern-recognition receptors (PRRs) located on the host cell's surface. Many plant PRRs are kinases. Here, we report that the Arabidopsis receptor kinase EF-TU RECEPTOR (EFR), which perceives the elf18 peptide derived from bacterial elongation factor Tu, is activated upon ligand binding by phosphorylation on its tyrosine residues. Phosphorylation of a single tyrosine residue, Y836, is required for activation of EFR and downstream immunity to the phytopathogenic bacterium Pseudomonas syringae. A tyrosine phosphatase, HopAO1, secreted by P. syringae, reduces EFR phosphorylation and prevents subsequent immune responses. Thus, host and pathogen compete to take control of PRR tyrosine phosphorylation used to initiate antibacterial immunity.

  7. Interaction of Pattern Recognition Receptors with Mycobacterium Tuberculosis.

    PubMed

    Mortaz, Esmaeil; Adcock, Ian M; Tabarsi, Payam; Masjedi, Mohammad Reza; Mansouri, Davood; Velayati, Ali Akbar; Casanova, Jean-Laurent; Barnes, Peter J

    2015-01-01

    Tuberculosis (TB) is considered a major worldwide health problem with 10 million new cases diagnosed each year. Our understanding of TB immunology has become greater and more refined since the identification of Mycobacterium tuberculosis (MTB) as an etiologic agent and the recognition of new signaling pathways modulating infection. Understanding the mechanisms through which the cells of the immune system recognize MTB can be an important step in designing novel therapeutic approaches, as well as improving the limited success of current vaccination strategies. A great challenge in chronic disease is to understand the complexities, mechanisms, and consequences of host interactions with pathogens. Innate immune responses along with the involvement of distinct inflammatory mediators and cells play an important role in the host defense against the MTB. Several classes of pattern recognition receptors (PRRs) are involved in the recognition of MTB including Toll-Like Receptors (TLRs), C-type lectin receptors (CLRs) and Nod-like receptors (NLRs) linked to inflammasome activation. Among the TLR family, TLR1, TLR2, TLR4, and TLR9 and their down-stream signaling proteins play critical roles in the initiation of the immune response in the pathogenesis of TB. The inflammasome pathway is associated with the coordinated release of cytokines such as IL-1β and IL-18 which also play a role in the pathogenesis of TB. Understanding the cross-talk between these signaling pathways will impact on the design of novel therapeutic strategies and in the development of vaccines and immunotherapy regimes. Abnormalities in PRR signaling pathways regulated by TB will affect disease pathogenesis and need to be elucidated. In this review we provide an update on PRR signaling during M. tuberculosis infection and indicate how greater knowledge of these pathways may lead to new therapeutic opportunities.

  8. Strategies for carbohydrate recognition by the mannose 6-phosphate receptors

    PubMed Central

    Dahms, Nancy M; Olson, Linda J; Kim, Jung-Ja P

    2008-01-01

    The two members of the P-type lectin family, the 46 kDa cation-dependent mannose 6-phosphate receptor (CD-MPR) and the 300 kDa cation-independent mannose 6-phosphate receptor (CI-MPR), are ubiquitously expressed throughout the animal kingdom and are distinguished from all other lectins by their ability to recognize phosphorylated mannose residues. The best-characterized function of the MPRs is their ability to direct the delivery of ∼60 different newly synthesized soluble lysosomal enzymes bearing mannose 6-phosphate (Man-6-P) on their N-linked oligosaccharides to the lysosome. In addition to its intracellular role in lysosome biogenesis, the CI-MPR, but not the CD-MPR, participates in a number of other biological processes by interacting with various molecules at the cell surface. The list of extracellular ligands recognized by this multifunctional receptor has grown to include a diverse spectrum of Man-6-P-containing proteins as well as several non-Man-6-P-containing ligands. Recent structural studies have given us a clearer view of how these two receptors use related, but yet distinct, approaches in the recognition of phosphomannosyl residues. PMID:18621992

  9. Pharmacology and therapeutic potential of pattern recognition receptors.

    PubMed

    Paul-Clark, M J; George, P M; Gatheral, T; Parzych, K; Wright, W R; Crawford, D; Bailey, L K; Reed, D M; Mitchell, J A

    2012-08-01

    Pharmacologists have used pathogen-associated molecular patterns (PAMPs), such as lipopolysaccharide (LPS) for decades as a stimulus for studying mediators involved in inflammation and for the screening of anti-inflammatory compounds. However, in the view of immunologists, LPS was too non-specific for studying the mechanisms of immune signalling in infection and inflammation, as no receptors had been identified. This changed in the late 1990s with the discovery of the Toll-like receptors. These 'pattern recognition receptors' (PRRs) were able to recognise highly conserved sequences, the so called pathogen associated molecular patterns (PAMPs) present in or on pathogens. This specificity of particular PAMPs and their newly defined receptors provided a common ground between pharmacologists and immunologists for the study of inflammation. PRRs also recognise endogenous agonists, the so called danger-associated molecular patterns (DAMPs), which can result in sterile inflammation. The signalling pathways and ligands of many PRRs have now been characterised and there is no doubt that this rich vein of research will aid the discovery of new therapeutics for infectious conditions and chronic inflammatory disease.

  10. Molecular Recognition of Insulin by a Synthetic Receptor

    SciTech Connect

    Chinai, Jordan M.; Taylor, Alexander B.; Ryno, Lisa M.; Hargreaves, Nicholas D.; Morris, Christopher A.; Hart, P. John; Urbach, Adam R.

    2011-08-29

    The discovery of molecules that bind tightly and selectively to desired proteins continues to drive innovation at the interface of chemistry and biology. This paper describes the binding of human insulin by the synthetic receptor cucurbit[7]uril (Q7) in vitro. Isothermal titration calorimetry and fluorescence spectroscopy experiments show that Q7 binds to insulin with an equilibrium association constant of 1.5 x 10{sup 6} M{sup -1} and with 50-100-fold selectivity versus proteins that are much larger but lack an N-terminal aromatic residue, and with >1000-fold selectivity versus an insulin variant lacking the N-terminal phenylalanine (Phe) residue. The crystal structure of the Q7{center_dot}insulin complex shows that binding occurs at the N-terminal Phe residue and that the N-terminus unfolds to enable binding. These findings suggest that site-selective recognition is based on the properties inherent to a protein terminus, including the unique chemical epitope presented by the terminal residue and the greater freedom of the terminus to unfold, like the end of a ball of string, to accommodate binding. Insulin recognition was predicted accurately from studies on short peptides and exemplifies an approach to protein recognition by targeting the terminus.

  11. Recognition of Bacterial Signal Peptides by Mammalian Formyl Peptide Receptors

    PubMed Central

    Bufe, Bernd; Schumann, Timo; Kappl, Reinhard; Bogeski, Ivan; Kummerow, Carsten; Podgórska, Marta; Smola, Sigrun; Hoth, Markus; Zufall, Frank

    2015-01-01

    Formyl peptide receptors (FPRs) are G-protein-coupled receptors that function as chemoattractant receptors in innate immune responses. Here we perform systematic structure-function analyses of FPRs from six mammalian species using structurally diverse FPR peptide agonists and identify a common set of conserved agonist properties with typical features of pathogen-associated molecular patterns. Guided by these results, we discover that bacterial signal peptides, normally used to translocate proteins across cytoplasmic membranes, are a vast family of natural FPR agonists. N-terminally formylated signal peptide fragments with variable sequence and length activate human and mouse FPR1 and FPR2 at low nanomolar concentrations, thus establishing FPR1 and FPR2 as sensitive and broad signal peptide receptors. The vomeronasal receptor mFpr-rs1 and its sequence orthologue hFPR3 also react to signal peptides but are much more narrowly tuned in signal peptide recognition. Furthermore, all signal peptides examined here function as potent activators of the innate immune system. They elicit robust, FPR-dependent calcium mobilization in human and mouse leukocytes and trigger a range of classical innate defense mechanisms, such as the production of reactive oxygen species, metalloprotease release, and chemotaxis. Thus, bacterial signal peptides constitute a novel class of immune activators that are likely to contribute to mammalian immune defense against bacteria. This evolutionarily conserved detection mechanism combines structural promiscuity with high specificity and enables discrimination between bacterial and eukaryotic signal sequences. With at least 175,542 predicted sequences, bacterial signal peptides represent the largest and structurally most heterogeneous class of G-protein-coupled receptor agonists currently known for the innate immune system. PMID:25605714

  12. Domain architecture evolution of pattern-recognition receptors

    PubMed Central

    Zhang, Qing; Zmasek, Christian M.

    2010-01-01

    In animals, the innate immune system is the first line of defense against invading microorganisms, and the pattern-recognition receptors (PRRs) are the key components of this system, detecting microbial invasion and initiating innate immune defenses. Two families of PRRs, the intracellular NOD-like receptors (NLRs) and the transmembrane Toll-like receptors (TLRs), are of particular interest because of their roles in a number of diseases. Understanding the evolutionary history of these families and their pattern of evolutionary changes may lead to new insights into the functioning of this critical system. We found that the evolution of both NLR and TLR families included massive species-specific expansions and domain shuffling in various lineages, which resulted in the same domain architectures evolving independently within different lineages in a process that fits the definition of parallel evolution. This observation illustrates both the dynamics of the innate immune system and the effects of “combinatorially constrained” evolution, where existence of the limited numbers of functionally relevant domains constrains the choices of domain architectures for new members in the family, resulting in the emergence of independently evolved proteins with identical domain architectures, often mistaken for orthologs. Electronic supplementary material The online version of this article (doi:10.1007/s00251-010-0428-1) contains supplementary material, which is available to authorized users. PMID:20195594

  13. [New immunology--immunology of pattern recognition receptors].

    PubMed

    Lebedev, K A; Poniakina, I D

    2006-01-01

    Pattern recognition receptors (PRRs) have been found on all cells of the body--cells of the innate and adaptive immune systems, epithelial and endothelial cells, keratinocytes, etc. PRRs can recognize specific molecular structures of microorganisms as well as allergens and other substances. The interaction with ligands of foreign microorganisms activates PRRs, after which host cells start to produce cytokines to both specifically activate innate immunity and to control adaptive immune reactions. On the other hand, no immune response develops against microorganisms of the normal microflora. Practically, the development of all immune responses is controlled by PRRs. These responses start in epithelial cells, skin cells, and vascular epithelial cells, which meet alien first. The immune system uses these cells to control the composition of normal microflora. Accordingly, the definition of immune system functions should be complemented by the regulation of body's microflora in addition to the protection from alien and altered self.

  14. Fungal recognition enhances mannose receptor shedding through dectin-1 engagement.

    PubMed

    Gazi, Umut; Rosas, Marcela; Singh, Sonali; Heinsbroek, Sigrid; Haq, Imran; Johnson, Simon; Brown, Gordon D; Williams, David L; Taylor, Philip R; Martinez-Pomares, Luisa

    2011-03-11

    The mannose receptor (MR) is an endocytic type I membrane molecule with a broad ligand specificity that is involved in both hemostasis and pathogen recognition. Membrane-anchored MR is cleaved by a metalloproteinase into functional soluble MR (sMR) composed of the extracellular domains of intact MR. Although sMR production was initially considered a constitutive process, enhanced MR shedding has been observed in response to the fungal pathogen Pneumocystis carinii. In this work, we have investigated the mechanism mediating enhanced MR shedding in response to fungi. We show that other fungal species, including Candida albicans and Aspergillus fumigatus, together with zymosan, a preparation of the cell wall of Saccharomyces cerevisiae, mimic the effect of P. carinii on sMR production and that this effect takes place mainly through β-glucan recognition. Additionally, we demonstrate that MR cleavage in response to C. albicans and bioactive particulate β-glucan requires expression of dectin-1. Our data, obtained using specific inhibitors, are consistent with the canonical Syk-mediated pathway triggered by dectin-1 being mainly responsible for inducing MR shedding, with Raf-1 being partially involved. As in the case of steady-state conditions, MR shedding in response to C. albicans and β-glucan particles requires metalloprotease activity. The induction of MR shedding by dectin-1 has clear implications for the role of MR in fungal recognition, as sMR was previously shown to retain the ability to bind fungal pathogens and can interact with numerous host molecules, including lysosomal hydrolases. Thus, MR cleavage could also impact on the magnitude of inflammation during fungal infection.

  15. The Role of Pattern Recognition Receptors in Intestinal Inflammation

    PubMed Central

    Fukata, Masayuki; Arditi, Moshe

    2013-01-01

    Recognition of microorganisms by pattern recognition receptors (PRRs) is the primary component of innate immunity that is responsible for the maintenance of host-microbial interactions in intestinal mucosa. Disregulation in host-commensal interactions has been implicated as the central pathogenesis of inflammatory bowel disease (IBD), which predisposes to developing colorectal cancer. Recent animal studies have begun to outline some unique physiology and pathology involving each PRR signaling in the intestine. The major roles played by PRRs in the gut appear to be regulation of the number and the composition of commensal bacteria, epithelial proliferation and mucosal permiability in response to epithelial injury. In addition, PRR signaling in lamina propria immune cells may be involved in induction of inflammation in response to invasion of pathogens. Because some PRR-deficient mice have shown variable susceptibility to colitis, the outcome of intestinal inflammation may be modified depending on PRR signaling in epithelial cells, immune cells, and the composition of commensal flora. Through recent findings in animal models of IBD, this review will discuss how abnormal PRR signaling may contribute to the pathogenesis of inflammation and inflammation-associated tumorigenesis in the intestine. PMID:23515136

  16. A 16-amino acid peptide from human alpha2-macroglobulin binds transforming growth factor-beta and platelet-derived growth factor-BB.

    PubMed Central

    Webb, D. J.; Roadcap, D. W.; Dhakephalkar, A.; Gonias, S. L.

    2000-01-01

    Alpha2-macroglobulin (alpha2M) is a major carrier of transforming growth factor-beta (TGF-beta) in vitro and in vivo. By screening glutathione S-transferase (GST) fusion proteins with overlapping sequences, we localized the TGFbeta-binding site to aa 700-738 of the mature human alpha2M subunit. In separate experiments, we screened overlapping synthetic peptides corresponding to aa 696-777 of alpha2M and identified a single 16-mer (718-733) that binds TGF-beta1. Platelet-derived growth factor-BB (PDGF-BB) bound to the same peptide, even though TGF-beta and PDGF-BB share almost no sequence identity. The sequence of the growth factor-binding peptide, WDLVVVNSAGVAEVGV, included a high proportion of hydrophobic amino acids. The analogous peptide from murinoglobulin, a human alpha2M homologue that does not bind growth factors, contained only three nonconservative amino acid substitutions; however, the MUG peptide failed to bind TGF-beta1 and PDGF-BB. These results demonstrate that a distinct and highly-restricted site in alpha2M, positioned near the C-terminal flank of the bait region, mediates growth factor binding. At least part of the growth factor-binding site is encoded by exon 18 of the alpha2M gene, which is notable for a 5' splice site polymorphism that has been implicated in Alzheimer's Disease. PMID:11106172

  17. Hampered cumulus expansion of porcine cumulus-oocyte complexes by excessive presence of alpha2 -macroglobulin is likely mediated via inhibition of zinc-dependent metalloproteases.

    PubMed

    Appeltant, Ruth; Beek, Josine; Maes, Dominiek; Bijttebier, Jo; Van Steendam, Katleen; Nauwynck, Hans; Van Soom, Ann

    2017-01-26

    In vitro maturation (IVM) in serum causes hampered expansion of porcine cumulus-oocyte complexes (COCs) due to excessive alpha2 -macroglobulin (A2M). This study investigated two hypotheses that could explain the effect of A2M: (i) binding of epidermal growth factor (EGF) to A2M, followed by its decreased availability; and (ii) inhibition of zinc-dependent metalloproteases. Cumulus expansion was evaluated based on the diameter of the COCs, the proportion of COCs participating in a floating cloud and the proportion of COCs with loss of cumulus cells. The first hypothesis of decreased EGF availability was tested by increasing the EGF concentration (20 and 50 ng/mL vs. 10 ng/mL), but was not confirmed because cumulus expansion did not improve. To verify the second hypothesis of inhibited zinc-dependent metalloproteases, the effect of tissue inhibitor of metalloproteases-3 (TIMP-3) on cumulus expansion during IVM with and without A2M was investigated. To immuno-neutralize A2M, serum was pre-incubated with A2M antibodies. Impaired cumulus expansion because of TIMP-3 could only be observed during IVM in 10% of serum with A2M antibodies. No effect of TIMP-3 was observed in medium without A2M antibodies. These results indicate that A2M and TIMP-3 share a common target, a zinc-dependent metalloprotease. Future research is directed toward the identification of the protease involved.

  18. Alpha 2 macroglobulin is a maternally-derived immune factor in amphioxus embryos: New evidence for defense roles of maternal immune components in invertebrate chordate.

    PubMed

    Pathirana, Anjalika; Diao, Mingyue; Huang, Shibo; Zuo, Lingling; Liang, Yujun

    2016-03-01

    In fish, a series of maternal derived immune components have been identified in their eggs or embryos at very early stages, which are proposed to provide protections to themselves against pathogenic attacks from hostile environment. The phenomenon of maternal immunity has been also recorded in several invertebrate species, however, so far, very limited information about the maternal immune molecules are available. In this study, it was demonstrated maternal alpha2 macroglobulin (A2m) protein, an important innate immune factor, exists in the fertilized eggs of amphioxus Branchiostoma japonicum, an invertebrate chordate. Maternal mRNA of A2m was also detected in amphioxus embryos at very early developing stages. In addition, it was recorded that the egg lysate prepared from the newly fertilized eggs can inhibit the growth of both Gram-negative bacterium Escherichia coli and Gram-positive bacterium Staphylococcus aureus in a concentration dependent manner. The bacteriostatic activity can be reduced notably after precipitated A2m with anti-A2m antibody. Thus maternal A2m is partly attributed to the bacteriostatic activity. It was further demonstrated that recombinant A2m can bind to E. coli cells directly. All these points come to a result that A2m is a maternal immune factor existing in eggs of invertebrate chordate, which may be involved in defense their embryos against harmful microbes' attacks.

  19. Lectin-based protein microarray analysis of differences in serum alpha-2-macroglobulin glycosylation between patients with colorectal cancer and persons without cancer.

    PubMed

    Šunderić, Miloš; Šedivá, Alena; Robajac, Dragana; Miljuš, Goran; Gemeiner, Peter; Nedić, Olgica; Katrlík, Jaroslav

    2016-07-01

    Glycosylation is co- and posttranslational modifications affecting proteins. The glycopattern changes are associated with changes in biological function and are involved in many diseases including cancer. We present the lectin-based protein microarray method enabling determination of differences in protein glycosylation. The method involves isolation of targeted protein from samples by immunoprecipitation, spotting of protein from multiple samples into arrays on a microarray slide, incubation with set of biotinylated lectins, the reaction with fluorescent conjugate of streptavidin, and detection of fluorescent intensities by microarray scanner. Lectin-based protein microarray was applied in investigation of differences in alpha-2-macroglobulin (α2M) glycosylation isolated from sera samples of healthy persons and patients with colorectal cancer (CC). From 14 lectins used in analysis, statistically significant differences (Student's t-test, P < 0.05) between two groups of samples (persons without cancer and CC patients) were found for 5 of them. α2M molecules isolated from sera of CC patients have higher content of α2,6 sialic acid, N-acetylglucosamine and mannose residues, and tri-/tetraantennary complex type high-mannose N-glycans. A novel lectin-based protein microarray developed and described can serve as a suitable analytical technique for sensitive, simple, fast, and high-throughput determination of differences in protein glycosylation isolated from serum or other samples.

  20. Inactivation of viruses by pasteurization at 60 °C for 10 h with and without 40% glucose as stabilizer during a new manufacturing process of α2-Macroglobulin from Cohn Fraction IV.

    PubMed

    Huangfu, Chaoji; Ma, Yuyuan; Jia, Junting; Lv, Maomin; Zhu, Fengxuan; Ma, Xiaowei; Zhao, Xiong; Zhang, Jingang

    2017-03-01

    Pasteurization is regularly used to inactivate viruses for the safety of plasma derivatives. Influence of pasteurization at 60 °C for 10 h on α2-Macroglobulin activity and virus inactivation were studied. With 40% sugar as stabilizers more than 70% α2-Macroglobulin activity was reserved after pasteurization compared with 20% in control. Glucose presented a better activity protection effect than sucrose and maltose. By pasteurization without stabilizer the virus titers of pseudorabies virus, Sindbis virus, porcine parvovirus and encephalomyocarditis virus were reduced more than 5.88 log10, 7.50 log10, 4.88 log10, and 5.63 log10 respectively within 2 h. By pasteurization with 40% glucose vesicular stomatitis virus was inactivated more than 5.88 log10 within 1 h. Only 2.71 log10 reduction was achieved for encephalomyocarditis virus after 10 h. 40% glucose protected α2-M activity and viruses simultaneously from pasteurization. Other viral inactivation methods need to be incorporated to ensure viral safety of this manufacturing process of α2-Macroglobulin.

  1. Potent neutralization of hepatitis A virus reveals a receptor mimic mechanism and the receptor recognition site

    PubMed Central

    Wang, Xiangxi; Zhu, Ling; Dang, Minghao; Hu, Zhongyu; Gao, Qiang; Yuan, Shuai; Sun, Yao; Zhang, Bo; Ren, Jingshan; Kotecha, Abhay; Walter, Thomas S.; Wang, Junzhi; Fry, Elizabeth E.; Stuart, David I.; Rao, Zihe

    2017-01-01

    Hepatitis A virus (HAV) infects ∼1.4 million people annually and, although there is a vaccine, there are no licensed therapeutic drugs. HAV is unusually stable (making disinfection problematic) and little is known of how it enters cells and releases its RNA. Here we report a potent HAV-specific monoclonal antibody, R10, which neutralizes HAV infection by blocking attachment to the host cell. High-resolution cryo-EM structures of HAV full and empty particles and of the complex of HAV with R10 Fab reveal the atomic details of antibody binding and point to a receptor recognition site at the pentamer interface. These results, together with our observation that the R10 Fab destabilizes the capsid, suggest the use of a receptor mimic mechanism to neutralize virus infection, providing new opportunities for therapeutic intervention. PMID:28074040

  2. Potent neutralization of hepatitis A virus reveals a receptor mimic mechanism and the receptor recognition site.

    PubMed

    Wang, Xiangxi; Zhu, Ling; Dang, Minghao; Hu, Zhongyu; Gao, Qiang; Yuan, Shuai; Sun, Yao; Zhang, Bo; Ren, Jingshan; Kotecha, Abhay; Walter, Thomas S; Wang, Junzhi; Fry, Elizabeth E; Stuart, David I; Rao, Zihe

    2017-01-24

    Hepatitis A virus (HAV) infects ∼1.4 million people annually and, although there is a vaccine, there are no licensed therapeutic drugs. HAV is unusually stable (making disinfection problematic) and little is known of how it enters cells and releases its RNA. Here we report a potent HAV-specific monoclonal antibody, R10, which neutralizes HAV infection by blocking attachment to the host cell. High-resolution cryo-EM structures of HAV full and empty particles and of the complex of HAV with R10 Fab reveal the atomic details of antibody binding and point to a receptor recognition site at the pentamer interface. These results, together with our observation that the R10 Fab destabilizes the capsid, suggest the use of a receptor mimic mechanism to neutralize virus infection, providing new opportunities for therapeutic intervention.

  3. Identification of Alpha 2 Macroglobulin (A2M) as a master inhibitor of cartilage degrading factors that attenuates post-traumatic osteoarthritis progression

    PubMed Central

    Wang, Shaowei; Wei, Xiaochun; Zhou, Jingming; Zhang, Jing; Li, Kai; Chen, Qian; Terek, Richard; Fleming, Braden C.; Goldring, Mary B.; Ehrlich, Michael G.; Zhang, Ge; Wei, Lei

    2014-01-01

    Objective To determine if supplemental intra-articular alpha-2 macroglobulin (A2M) has a chondroprotective effect in a rat OA model. Methods A2M was identified as a potential therapeutic agent by comparing A2M concentrations in serum, synovial fluid (SF), and cartilage from normal and osteoarthritic (OA) patients by Western blotting, mass spectrometry, ELISA, and immunohistochemistry (IHC). The effects of A2M on IL-1-induced cartilage catabolic enzymes were evaluated by Luminex and ELISA in cultured chondrocytes. In vivo effects on cartilage degeneration and MMP-13 concentration were evaluated in male rats (N=120) randomized to four treatments: (1) CLT+saline, (2) ACLT+A2M (1IU/kg), (3) ACLT+A2M (2IU/kg) or (4) sham surgery+saline. Intra-articular injections were given for 6 weeks. The concentration of MMP-13 in SF lavages was measured using ELISA. OA-related gene expression was quantified by RT-qPCR. Histology was performed to grade OA. Results In both normal and OA patients, the levels of A2M were lower in SF compared to serum, and MMP-13 was higher in SF than serum of OA patients. In vitro, A2M inhibited the induction of MMP-13 by IL-1 in a dose-dependent manner in human chondrocytes. In the rat ACLT OA model, supplemental intra-articular injection of A2M reduced the concentration of MMP-13 in SF, had a favorable effect on OA-related gene expression, and attenuated OA progression. Conclusion A2M is a plasma protease inhibitor that is not present in sufficient concentrations to inactivate the high concentrations of catabolic factors found in OA SF. Our findings suggest that supplemental intra-articular A2M provides chondral protection for post traumatic OA. PMID:24578232

  4. Administration of rat acute-phase protein α(2)-macroglobulin before total-body irradiation initiates cytoprotective mechanisms in the liver.

    PubMed

    Bogojević, Desanka; Poznanović, Goran; Grdović, Nevena; Grigorov, Ilijana; Vidaković, Melita; Dinić, Svetlana; Mihailović, Mirjana

    2011-03-01

    Previously, we showed that administration of the acute-phase protein α(2)-macroglobulin (α(2)M) to rats before total-body irradiation with 6.7 Gy (LD(50/30)) of X-rays provides the same level of radioprotection as amifostine. Here, we compare the cytoprotective effects of α(2)M and amifostine on rat liver. The potential of the liver to replenish cells destroyed by ionizing radiation was assessed by immunoblot analysis with antibody to proliferating cell nuclear antigen (PCNA). After irradiation, in unprotected rats PCNA decreased 6-fold from the basal level. In rats pretreated with either α(2)M or amifostine, PCNA was increased throughout a 4 week follow-up period, indicating that hepatocyte proliferation was unaffected. Since PCNA is an important component of the repair machinery, its increased expression was accompanied by significantly lower DNA damage in α(2)M- and amifostine-treated rats. At 2 weeks after irradiation, the Comet assay revealed a 15-fold increase in DNA damage in unprotected rats, while in α(2)M- and amifostine-treated rats we observed 3- and 4-fold rise in damage, respectively. The improved protection to DNA damage was supported by elevated activity of the antioxidant systems. Compared to untreated rats, pretreatments with α(2)M and amifostine led to similar increases in levels of the inflammatory cytokine IL-6 and the redox-sensitive transcription factor NFκB, promoting upregulation of MnSOD, the major component of the cell's antioxidant axis, and subsequent increases in Mn/CuZnSOD and catalase enzymatic activities. The results show that α(2)M induces protein factors whose interplay underlies radioprotection and support the idea that α(2)M is the central effector of natural radioprotection in the rat.

  5. Effect of retinal laser photocoagulation on the activity of metalloproteinases and the alpha(2)-macroglobulin proteolytic state in the vitreous of eyes with proliferative diabetic retinopathy.

    PubMed

    Sánchez, María C; Luna, Jose D; Barcelona, Pablo F; Gramajo, Ana L; Juarez, Patricio C; Riera, Clelia M; Chiabrando, Gustavo A

    2007-11-01

    Panretinal photocoagulation (PRP) reduces the incidence of severe visual loss in proliferative diabetic retinopathy (PDR). The aim of the study was to determine the effect of PRP on the activity of matrix metalloproteinase-2 (MMP-2) and MMP-9, and also on the alpha(2)-Macroglobulin (alpha(2)M) proteolytic state in the vitreous of eyes with PDR. Vitreous samples were obtained from patients undergoing vitrectomy for the treatment of retinal diseases: 17 with PDR and eight with idiopathic macular hole (MH). Qualitative evaluation of the MMP-2 and MMP-9 activation status was performed by gelatin zymography and quantitative assay was carried out for vitreous total protein content and alpha(2)M. The proteolytic state of alpha(2)M was evaluated by Western blotting. Although all vitreous samples contained proMMP-2, increased proMMP-9 and active MMP-9 were detected in PDR samples without PRP. In addition, after PRP the proMMP-9 activity was significantly decreased, whereas the proMMP-2 activity was not affected. Enhanced total protein and alpha(2)M concentrations were observed in all vitreous samples from PDR patients with and without previous PRP compared with samples from patients with MH. However, a differential proteolytic state of alpha(2)M, expressed as 180/85-90kDa ratio, was detected among PDR patients with and without PRP treatment. Whereas a low 180/85-90kDa ratio of alpha(2)M in vitreous of PDR patients without PRP was observed, a high proportion of 180kDa subunit was principally detected in PDR with PRP. These results demonstrate that PDR occurs with an enhanced activity of MMP-9 and activation of alpha(2)M by proteinases, which is reversed after PRP. In addition, we suggest that alpha(2)M plays a key role in the control and regulation of the ocular neovascularization involved in the pathogenesis of ischemic retinal diseases such as PDR.

  6. Urinary IgG and α2-Macroglobulin Are Powerful Predictors of Outcome and Responsiveness to Steroids and Cyclophosphamide in Idiopathic Focal Segmental Glomerulosclerosis with Nephrotic Syndrome

    PubMed Central

    Bazzi, Claudio; Rizza, Virginia; Casellato, Daniela; Stivali, Gilda; Rachele, Gregorio; Napodano, Pietro; D'Amico, Giuseppe

    2013-01-01

    Objective. To assess whether high-molecular-weight proteins excretion predicts outcome and therapy-responsiveness in patients with FSGS and nephrotic syndrome. Research Design and Methods. Thirty-eight patients measured at biopsy fractional excretion of IgG (FEIgG) and urinary α2-macroglobulin/creatinine ratio (α2m/C). Low and high risk groups were defined by cutoffs assessed by ROC analysis. In all patients first-line therapy was with steroids alone or in combination with cyclophosphamide. Results. α2m/C and FEIgG were correlated with segmental sclerosis (r = 0.546; r = 0.522). Twenty-three patients (61%) entered Remission and 9 (24%) progressed to ESRD. Comparing low and high risk groups, by univariate analysis remission was predicted by FEIgG (77% versus 25%, P = 0.016) and α2m/C (81% versus 17%, P = 0.007) and ESRD at best by FEIgG (0% versus 75%, P < 0.0001) and α2m/C (4% versus 67%, P < 0.0001). By multivariate analysis FEIgG was the only independent predictor of remission and α2m/C the most powerful predictor of ESRD. Low and high risk groups of FEIgG and α2m/C in combination had very high predictive value of sustained remission and ESRD in response to therapy. Conclusions. FEIgG and α2m/C are powerful predictors of outcome and responsiveness to steroids and cyclophosphamide; their predictive value, if validated in prospective studies, may be useful in clinical practice suggesting first-line alternative treatments in high risk patients. PMID:24093110

  7. Differential Levels of Alpha-2-Macroglobulin, Haptoglobin and Sero-Transferrin as Adjunct Markers for TB Diagnosis and Disease Progression in the Malnourished Tribal Population of Melghat, India.

    PubMed

    Bapat, Prachi R; Satav, Ashish R; Husain, Aliabbas A; Shekhawat, Seema D; Kawle, Anuja P; Chu, Justin J; Purohit, Hemant J; Daginawala, Hatim F; Taori, Girdhar M; Kashyap, Rajpal S

    2015-01-01

    Lack of diagnostic capacity has been a crucial barrier preventing an effective response to the challenges of malnutrition and tuberculosis (TB). Point-of-care diagnostic tests for TB in immuno-incompetent, malnourished population are thus needed to ensure rapid and accurate detection. The aim of the study was to identify potential biomarkers specific for TB infection and progression to overt disease in the malnourished population of Melghat. A prospective cohort study was conducted in the year 2009 through 2011 in six villages of the Melghat region. 275 participants consisting of malnourished cases with a) active TB (n = 32), b) latent TB infection (n = 90), c) with no clinical or bacteriological signs of active or latent TB (n = 130) and healthy control subjects (n = 23) were recruited for the study. The proteome changes of the host serum in response to Mycobacterium tuberculosis (M.tb) infection were investigated using one dimensional electrophoresis in combination with matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Three most differentially expressed proteins; alpha-2-macroglobulin (A-2-M), sero-transferrin and haptoglobin were identified by MALDI-TOF MS analysis, which were up-regulated in the malnourished patients with active TB and down-regulated in the malnourished patients compared with the healthy controls. Additionally, follow-up studies indicated that the expression of these proteins increased to nearly two folds in patients who developed active disease from latent state. Our preliminary results suggest that A-2-M, sero-transferrin and haptoglobin may be clinically relevant host biomarkers for TB diagnosis and disease progression in the malnourished population. This study provides preliminary framework for an in-depth analysis of the biomarkers in larger well-characterized cohorts. Evaluation of these biomarkers in follow-up cases may further aid in improving TB diagnosis.

  8. Differential Levels of Alpha-2-Macroglobulin, Haptoglobin and Sero-Transferrin as Adjunct Markers for TB Diagnosis and Disease Progression in the Malnourished Tribal Population of Melghat, India

    PubMed Central

    Bapat, Prachi R.; Satav, Ashish R.; Husain, Aliabbas A.; Shekhawat, Seema D.; Kawle, Anuja P.; Chu, Justin J.; Purohit, Hemant J.; Daginawala, Hatim F.; Taori, Girdhar M.; Kashyap, Rajpal S.

    2015-01-01

    Lack of diagnostic capacity has been a crucial barrier preventing an effective response to the challenges of malnutrition and tuberculosis (TB). Point-of-care diagnostic tests for TB in immuno-incompetent, malnourished population are thus needed to ensure rapid and accurate detection. The aim of the study was to identify potential biomarkers specific for TB infection and progression to overt disease in the malnourished population of Melghat. A prospective cohort study was conducted in the year 2009 through 2011 in six villages of the Melghat region. 275 participants consisting of malnourished cases with a) active TB (n = 32), b) latent TB infection (n = 90), c) with no clinical or bacteriological signs of active or latent TB (n = 130) and healthy control subjects (n = 23) were recruited for the study. The proteome changes of the host serum in response to Mycobacterium tuberculosis (M.tb) infection were investigated using one dimensional electrophoresis in combination with matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Three most differentially expressed proteins; alpha-2-macroglobulin (A-2-M), sero-transferrin and haptoglobin were identified by MALDI-TOF MS analysis, which were up-regulated in the malnourished patients with active TB and down-regulated in the malnourished patients compared with the healthy controls. Additionally, follow-up studies indicated that the expression of these proteins increased to nearly two folds in patients who developed active disease from latent state. Our preliminary results suggest that A-2-M, sero-transferrin and haptoglobin may be clinically relevant host biomarkers for TB diagnosis and disease progression in the malnourished population. This study provides preliminary framework for an in-depth analysis of the biomarkers in larger well-characterized cohorts. Evaluation of these biomarkers in follow-up cases may further aid in improving TB diagnosis. PMID:26241963

  9. Oxytocin, vasopressin and estrogen receptor gene expression in relation to social recognition in female mice

    PubMed Central

    Clipperton-Allen, Amy E.; Lee, Anna W.; Reyes, Anny; Devidze, Nino; Phan, Anna; Pfaff, Donald W.; Choleris, Elena

    2012-01-01

    Inter- and intra-species differences in social behavior and recognition-related hormones and receptors suggest that different distribution and/or expression patterns may relate to social recognition. We used qRT-PCR to investigate naturally occurring differences in expression of estrogen receptor-alpha (ERα), ER-beta (ERβ), progesterone receptor (PR), oxytocin (OT) and receptor, and vasopressin (AVP) and receptors in proestrous female mice. Following four 5 min exposures to the same two conspecifics, one was replaced with a novel mouse in the final trial (T5). Gene expression was examined in mice showing high (85–100%) and low (40–60%) social recognition scores (i.e., preferential novel mouse investigation in T5) in eight socially-relevant brain regions. Results supported OT and AVP involvement in social recognition, and suggest that in the medial preoptic area, increased OT and AVP mRNA, together with ERα and ERβ gene activation, relate to improved social recognition. Initial social investigation correlated with ERs, PR and OTR in the dorsolateral septum, suggesting that these receptors may modulate social interest without affecting social recognition. Finally, increased lateral amygdala gene activation in the LR mice may be associated with general learning impairments, while decreased lateral amygdala activity may indicate more efficient cognitive mechanisms in the HR mice. PMID:22079582

  10. Molecular Recognition of Paired Receptors in the Immune System

    PubMed Central

    Kuroki, Kimiko; Furukawa, Atsushi; Maenaka, Katsumi

    2012-01-01

    Cell surface receptors are responsible for regulating cellular function on the front line, the cell membrane. Interestingly, accumulating evidence clearly reveals that the members of cell surface receptor families have very similar extracellular ligand-binding regions but opposite signaling systems, either inhibitory or stimulatory. These receptors are designated as paired receptors. Paired receptors often recognize not only physiological ligands but also non-self ligands, such as viral and bacterial products, to fight infections. In this review, we introduce several representative examples of paired receptors, focusing on two major structural superfamilies, the immunoglobulin-like and the C-type lectin-like receptors, and explain how these receptors distinguish self and non-self ligands to maintain homeostasis in the immune system. We further discuss the evolutionary aspects of these receptors as well as the potential drug targets for regulating diseases. PMID:23293633

  11. A cannabinoid CB(1) receptor antagonist ameliorates impairment of recognition memory on withdrawal from MDMA (Ecstasy).

    PubMed

    Nawata, Yoko; Hiranita, Takato; Yamamoto, Tsuneyuki

    2010-01-01

    (+/-)-3,4-Methylenedioxymethamphetamine (MDMA, 'Ecstasy') abusers have persistent neuropsychiatric deficits including memory impairments after the cessation of abuse. On the other hand, cannabinoid CB(1) receptors have been implicated in learning/memory, and are highly expressed in the hippocampus, a region of the brain believed to have an important function in certain forms of learning and memory. In this study, we clarified the mechanism underlying the cognitive impairment that develops during MDMA withdrawal from the standpoint of the cannabinoid CB(1) receptors. Mice were administered MDMA (10 mg/kg, i.p.) once a day for 7 days. On the 7th day of withdrawal, a novel object recognition task was performed and the amount of cannabinoid CB(1) receptor protein was measured with western blotting. Recognition performance was impaired on the 7th day of withdrawal. This impairment was blocked by AM251, a cannabinoid CB(1) receptor antagonist, administered 30 min before the training trial or co-administered with MDMA. At this time, the level of cannabinoid CB(1) receptor protein increased significantly in the hippocampus but not the prefrontal cortex or striatum. This increase of CB(1) receptor protein in the hippocampus was also blocked by the co-administration of AM251. Furthermore, CB(1) receptor knockout mice showed no impairment of recognition performance on the withdrawal from MDMA. The impairment of recognition memory during withdrawal from MDMA may result from the activation of cannabinoid CB(1) receptors in the hippocampus.

  12. Inhibition of pattern recognition receptor-mediated inflammation by bioactive phytochemicals

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Emerging evidence reveals that pattern-recognition receptors (PRRs), Toll-like receptors (TLRs) and Nucleotide-binding oligomerization domain proteins (NODs) mediate both infection-induced and sterile inflammation by recognizing pathogen-associated molecular patterns (PAMPs) and endogenous molecules...

  13. Molecular recognition of organic ammonium ions in solution using synthetic receptors

    PubMed Central

    Späth, Andreas

    2010-01-01

    Summary Ammonium ions are ubiquitous in chemistry and molecular biology. Considerable efforts have been undertaken to develop synthetic receptors for their selective molecular recognition. The type of host compounds for organic ammonium ion binding span a wide range from crown ethers to calixarenes to metal complexes. Typical intermolecular interactions are hydrogen bonds, electrostatic and cation–π interactions, hydrophobic interactions or reversible covalent bond formation. In this review we discuss the different classes of synthetic receptors for organic ammonium ion recognition and illustrate the scope and limitations of each class with selected examples from the recent literature. The molecular recognition of ammonium ions in amino acids is included and the enantioselective binding of chiral ammonium ions by synthetic receptors is also covered. In our conclusion we compare the strengths and weaknesses of the different types of ammonium ion receptors which may help to select the best approach for specific applications. PMID:20502608

  14. Diverse binding modes, same goal: the receptor recognition mechanism of botulinum neurotoxin

    PubMed Central

    Lam, Kwok-Ho; Yao, Guorui; Jin, Rongsheng

    2015-01-01

    Botulinum neurotoxins (BoNTs) are among the most deadly toxins known. They act rapidly in a highly specific manner to block neurotransmitter release by cleaving the soluble N-ethylmaleimide sensitive factor attachment protein receptor (SNARE) complex at neuromuscular junctions. The extreme toxicity of BoNTs relies predominantly on their neurotropism that is accomplished by recognition of two host receptors, a polysialo-ganglioside and in the majority of cases a synaptic vesicle protein, through their receptor-binding domains. Two proteins, synaptotagmin and synaptic vesicle glycoprotein 2, have been identified as the receptors for various serotypes of BoNTs. Here, we review recent breakthroughs in the structural studies of BoNT–protein receptor recognitions that highlight a range of diverse mechanisms by which BoNTs manipulate host neuronal proteins for highly specific uptake at neuromuscular junctions. PMID:25701633

  15. Variable Lymphocyte Receptor Recognition of the Immunodominant Glycoprotein of Bacillus anthracis Spores

    SciTech Connect

    Kirchdoerfer, Robert N.; Herrin, Brantley R.; Han, Byung Woo; Turnbough, Jr., Charles L.; Cooper, Max D.; Wilson, Ian A.

    2012-07-25

    Variable lymphocyte receptors (VLRs) are the adaptive immune receptors of jawless fish, which evolved adaptive immunity independent of other vertebrates. In lieu of the immunoglobulin fold-based T and B cell receptors, lymphocyte-like cells of jawless fish express VLRs (VLRA, VLRB, or VLRC) composed of leucine-rich repeats and are similar to toll-like receptors (TLRs) in structure, but antibodies (VLRB) and T cell receptors (VLRA and VLRC) in function. Here, we present the structural and biochemical characterization of VLR4, a VLRB, in complex with BclA, the immunodominant glycoprotein of Bacillus anthracis spores. Using a combination of crystallography, mutagenesis, and binding studies, we delineate the mode of antigen recognition and binding between VLR4 and BclA, examine commonalities in VLRB recognition of antigens, and demonstrate the potential of VLR4 as a diagnostic tool for the identification of B. anthracis spores.

  16. The role of histamine receptors in the consolidation of object recognition memory.

    PubMed

    da Silveira, Clarice Krás Borges; Furini, Cristiane R G; Benetti, Fernando; Monteiro, Siomara da Cruz; Izquierdo, Ivan

    2013-07-01

    Findings have shown that histamine receptors in the hippocampus modulate the acquisition and extinction of fear motivated learning. In order to determine the role of hippocampal histaminergic receptors on recognition memory, adult male Wistar rats with indwelling infusion cannulae stereotaxically placed in the CA1 region of dorsal hippocampus were trained in an object recognition learning task involving exposure to two different stimulus objects in an enclosed environment. In the test session, one of the objects presented during training was replaced by a novel one. Recognition memory retention was assessed 24 h after training by comparing the time spent in exploration (sniffing and touching) of the known object with that of the novel one. When infused in the CA1 region immediately, 30, 120 or 360 min posttraining, the H1-receptor antagonist, pyrilamine, the H2-receptor antagonist, ranitidine, and the H3-receptor agonist, imetit, blocked long-term memory retention in a time dependent manner (30-120 min) without affecting general exploratory behavior, anxiety state or hippocampal function. Our data indicate that histaminergic system modulates consolidation of object recognition memory through H1, H2 and H3 receptors.

  17. Biochemical study of multiple drug recognition sites on central benzodiazepine receptors

    SciTech Connect

    Trifiletti, R.R.

    1986-01-01

    The benzodiazepine receptor complex of mammalian brain possesses recognition sites which mediate (at least in part) the pharmacologic actions of the 1,4-benzodiazepines and barbiturates. Evidence is provided suggesting the existence of least seven distinct drug recognition sites on this complex. Interactions between the various recognition sites have been explored using radioligand binding techniques. This information is utilized to provide a comprehensive scheme for characterizing receptor-active drugs on an anxiolytic-anticonvulsant/proconvulsant continuum using radioligand binding techniques, as well as a comprehensive program for identifying potential endogenous receptor-active substances. Further evidence is provided here supporting the notion of benzodiazepine recognition site heterogeneity. Classical 1,4-benzodiazepines do not appear to differentiate two populations of benzodiazepine receptors in an equilibrium sense, but appear to do so in a kinetic sense. An apparent physical separation of the two receptor subtypes can be achieved by differential solubilization. The benzodiazepine binding subunit can be identified by photoaffinity labeling with the benzodiazepine agonist (/sup 3/H)flunitrazepan. Conditions for reproducible partial proteolytic mapping of (/sup 3/H)flunitrazepam photoaffinity labeled receptors are established. From these maps, it is concluded that there are probably no major differences in the primary sequence of the benzodiazepine binding subunit in various regions of the rat central nervous system.

  18. Sugar recognition: designing artificial receptors for applications in biological diagnostics and imaging.

    PubMed

    Miron, Caitlin E; Petitjean, Anne

    2015-02-09

    At the cellular level, numerous processes ranging from protein folding to disease development are mediated by a sugar-based molecular information system that is much less well known than its DNA- or protein-based counterparts. The subtle structural diversity of such sugar tags nevertheless offers an excellent, if challenging, opportunity to design receptors for the selective recognition of biorelevant sugars. Over the past 40 years, growing interest in the field of sugar recognition has led to the development of several promising artificial receptors, which could soon find widespread use in medical diagnostics and cell imaging.

  19. Recognition of herpes simplex viruses: toll-like receptors and beyond.

    PubMed

    Ma, Yijie; He, Bin

    2014-03-20

    Herpes simplex viruses (HSVs) are human pathogens that establish lytic and latent infections. Reactivation from latency occurs intermittently, which represents a lifelong source of recurrent infection. In this complex process, HSV triggers and neutralizes innate immunity. Therefore, a dynamic equilibrium between HSV and the innate immune system determines the outcome of viral infection. Detection of HSV involves pathogen recognition receptors that include Toll-like receptors, retinoic acid-inducible gene I-like receptors, and cytosolic DNA sensors. Moreover, innate components or pathways exist to sense membrane fusion upon viral entry into host cells. Consequently, this surveillance network activates downstream transcription factors, leading to the induction of type I interferon and inflammatory cytokines. Not surprisingly, with the capacity to establish chronic infection HSV has evolved strategies that modulate or evade innate immunity. In this review, we describe recent advances pertinent to the interplay of HSV and the induction of innate immunity mediated by pathogen recognition receptors or pathways.

  20. N-haloacetylimino neonicotinoids: potency and molecular recognition at the insect nicotinic receptor.

    PubMed

    Tomizawa, Motohiro; Durkin, Kathleen A; Ohno, Ikuya; Nagura, Kyoko; Manabe, Mio; Kumazawa, Satoru; Kagabu, Shinzo

    2011-06-15

    This structure-activity relationship study for neonicotinoids with an N-haloacetylimino pharmacophore identifies several candidate compounds showing outstanding insecticidal potency and consequently leads to establishing their molecular recognition at an insect nicotinic receptor structural model, wherein the neonicotinoid halogen atoms (fluorine, chlorine, bromine, and iodine) variously interact with the receptor loops C-D interfacial niche via H-bonding and/or hydrophobic interactions.

  1. The Pathogen Recognition Receptor NOD2 Regulates Human FOXP3+ T Cell Survival

    PubMed Central

    Rahman, Meher K.; Midtling, Emilie H.; Svingen, Phyllis A.; Xiong, Yuning; Bell, Michael P.; Tung, Jeanne; Smyrk, Tom; Egan, Larry J.; Faubion, William A.

    2013-01-01

    The expression of pathogen recognition receptors in human FOXP3+ T regulatory cells is established, yet the function of these receptors is currently obscure. In the process of studying the function of both peripheral and lamina propria FOXP3+ lymphocytes in patients with the human inflammatory bowel disease Crohn’s disease, we observed a clear deficiency in the quantity of FOXP3+ lymphocytes in patients with disease-associated polymorphisms in the pathogen recognition receptor gene NOD2. Subsequently, we determined that the NOD2 ligand, muramyl dipeptide (MDP), activates NF-κB in primary human FOXP3+ T cells. This activation is functionally relevant, as MDP-stimulated human FOXP3+ T cells are protected from death receptor Fas-mediated apoptosis. Importantly, apoptosis protection was not evident in MDP-stimulated FOXP3+ T cells isolated from a patient with the disease-associated polymorphism. Thus, we propose that one function of pathogen recognition receptors in human T regulatory cells is the protection against death receptor-mediated apoptosis in a Fas ligand-rich environment, such as that of the inflamed intestinal subepithelial space. PMID:20483763

  2. Peptide ligand recognition by G protein-coupled receptors

    PubMed Central

    Krumm, Brian E.

    2015-01-01

    The past few years have seen spectacular progress in the structure determination of G protein-coupled receptors (GPCRs). We now have structural representatives from classes A, B, C, and F. Within the rhodopsin-like class A, most structures belong to the α group, whereas fewer GPCR structures are available from the β, γ, and δ groups, which include peptide GPCRs such as the receptors for neurotensin (β group), opioids, chemokines (γ group), and protease-activated receptors (δ group). Structural information on peptide GPCRs is restricted to complexes with non-peptidic drug-like antagonists with the exception of the chemokine receptor CXCR4 that has been crystallized in the presence of a cyclic peptide antagonist. Notably, the neurotensin receptor 1 is to date the only peptide GPCR whose structure has been solved in the presence of a peptide agonist. Although limited in number, the current peptide GPCR structures reveal great diversity in shape and electrostatic properties of the ligand binding pockets, features that play key roles in the discrimination of ligands. Here, we review these aspects of peptide GPCRs in view of possible models for peptide agonist binding. PMID:25852552

  3. Oxoanion Recognition by Benzene-based Tripodal Pyrrolic Receptors

    SciTech Connect

    Bill, Nathan; Kim, Dae-Sik; Kim, Sung Kuk; Park, Jung Su; Lynch, Vincent M.; Young, Neil J; Hay, Benjamin; Yang, Youjun; Anslyn, Eric; Sessler, Jonathan L.

    2012-01-01

    Two new tripodal receptors based on pyrrole- and dipyrromethane-functionalised derivatives of a sterically geared precursor, 1,3,5-tris(aminomethyl)-2,4,6-triethylbenzene, are reported; these systems, compounds 1 and 2, display high affinity and selectivity for tetrahedral anionic guests, in particular dihydrogen phosphate, pyrophosphate and hydrogen sulphate, in acetonitrile as inferred from isothermal titration calorimetry measurements. Support for the anion-binding ability of these systems comes from theoretical calculations and a single-crystal X-ray diffraction structure of the 2:2 (host:guest) dihydrogen phosphate complex is obtained in the case of the pyrrole-based receptor system, 1. Keywords anion receptors, dihydrogen phosphate, hydrogen sulphate, X-ray structure, theoretical calculations.

  4. Myeloid C-Type Lectin Receptors in Viral Recognition and Antiviral Immunity

    PubMed Central

    Monteiro, João T.; Lepenies, Bernd

    2017-01-01

    Recognition of viral glycans by pattern recognition receptors (PRRs) in innate immunity contributes to antiviral immune responses. C-type lectin receptors (CLRs) are PRRs capable of sensing glycans present in viral pathogens to activate antiviral immune responses such as phagocytosis, antigen processing and presentation, and subsequent T cell activation. The ability of CLRs to elicit and shape adaptive immunity plays a critical role in the inhibition of viral spread within the host. However, certain viruses exploit CLRs for viral entry into host cells to avoid immune recognition. To block CLR interactions with viral glycoproteins, antiviral strategies may involve the use of multivalent glycan carrier systems. In this review, we describe the role of CLRs in antiviral immunity and we highlight their dual function in viral clearance and exploitation by viral pathogens. PMID:28327518

  5. Structural basis for collagen recognition by the immune receptor OSCAR

    PubMed Central

    Zhou, Long; Hinerman, Jennifer M.; Blaszczyk, Michal; Miller, Jeanette L. C.; Conrady, Deborah G.; Barrow, Alexander D.; Chirgadze, Dimitri Y.; Bihan, Dominique; Farndale, Richard W.

    2016-01-01

    The osteoclast-associated receptor (OSCAR) is a collagen-binding immune receptor with important roles in dendritic cell maturation and activation of inflammatory monocytes as well as in osteoclastogenesis. The crystal structure of the OSCAR ectodomain is presented, both free and in complex with a consensus triple-helical peptide (THP). The structures revealed a collagen-binding site in each immunoglobulin-like domain (D1 and D2). The THP binds near a predicted collagen-binding groove in D1, but a more extensive interaction with D2 is facilitated by the unusually wide D1-D2 interdomain angle in OSCAR. Direct binding assays, combined with site-directed mutagenesis, confirm that the primary collagen-binding site in OSCAR resides in D2, in marked contrast to the related collagen receptors, glycoprotein VI (GPVI) and leukocyte-associated immunoglobulin-like receptor-1 (LAIR-1). Monomeric OSCAR D1D2 binds to the consensus THP with a KD of 28 µM measured in solution, but shows a higher affinity (KD 1.5 μM) when binding to a solid-phase THP, most likely due to an avidity effect. These data suggest a 2-stage model for the interaction of OSCAR with a collagen fibril, with transient, low-affinity interactions initiated by the membrane-distal D1, followed by firm adhesion to the primary binding site in D2. PMID:26552697

  6. NOD2 and Toll-Like Receptors Are Nonredundant Recognition Systems of Mycobacterium tuberculosis

    PubMed Central

    2005-01-01

    Infection with Mycobacterium tuberculosis is one of the leading causes of death worldwide. Recognition of M. tuberculosis by pattern recognition receptors is crucial for activation of both innate and adaptive immune responses. In the present study, we demonstrate that nucleotide-binding oligomerization domain 2 (NOD2) and Toll-like receptors (TLRs) are two nonredundant recognition mechanisms of M. tuberculosis. CHO cell lines transfected with human TLR2 or TLR4 were responsive to M. tuberculosis. TLR2 knock-out mice displayed more than 50% defective cytokine production after stimulation with mycobacteria, whereas TLR4-defective mice also released 30% less cytokines compared to controls. Similarly, HEK293T cells transfected with NOD2 responded to stimulation with M. tuberculosis. The important role of NOD2 for the recognition of M. tuberculosis was demonstrated in mononuclear cells of individuals homozygous for the 3020insC NOD2 mutation, who showed an 80% defective cytokine response after stimulation with M. tuberculosis. Finally, the mycobacterial TLR2 ligand 19-kDa lipoprotein and the NOD2 ligand muramyl dipeptide synergized for the induction of cytokines, and this synergism was lost in cells defective in either TLR2 or NOD2. Together, these results demonstrate that NOD2 and TLR pathways are nonredundant recognition mechanisms of M. tuberculosis that synergize for the induction of proinflammatory cytokines. PMID:16322770

  7. Labeled Protein Recognition at a Membrane Bilayer Interface by Embedded Synthetic Receptors

    PubMed Central

    2015-01-01

    Self-folding deep cavitands embedded in a supported lipid bilayer are capable of recognizing suitably labeled proteins at the bilayer interface. The addition of a choline derived binding “handle” to a number of different proteins allows their selective noncovalent recognition, with association constants on the order of 105 M–1. The proteins are displayed at the water:bilayer interface, and a single binding handle allows recognition of the large, charged protein by a small molecule synthetic receptor via complementary shape and charge interactions. PMID:25130415

  8. Gibberellin Receptor GID1: Gibberellin Recognition and Molecular Evolution

    NASA Astrophysics Data System (ADS)

    Kato, Hiroaki; Sato, Tomomi; Ueguchi-Tanaka, Miyako

    Gibberellins (GAs) are phytohormones essential for many developmental processes in plants. We analyzed the crystal structure of a nuclear GA receptor, GIBBERELLIN INSENSITIVE DWARF 1 (GID1) from Oryza sativa. As it was proposed from the sequence similarity, the overall structure of GID1 shows an α/β-hydrolase fold similar to that of the hormone-sensitive lipases (HSLs) except for an amino-terminal lid. The GA-binding site corresponds to the substrate-binding site of HSLs. Almost residues assigned for GA binding showed very little or no activity when they were replaced with Ala. The substitution of the residues corresponding to those of the lycophyte GID1s caused an increase in the binding affinity for GA34, a 2β-hydroxylated GA4. These findings indicate that GID1 originated from HSL and was tinkered to have the specificity for bioactive GAs in the course of plant evolution.

  9. Carboxylate-based receptors for the recognition of carbohydrates in organic and aqueous media.

    PubMed

    Mazik, Monika; Cavga, Hüseyin

    2006-04-14

    Acyclic receptors containing neutral and ionic hydrogen-bonding sites, such as amino-pyridine and carboxylate groups, were prepared and their binding properties toward neutral sugar molecules were studied. The binding studies with disodium and bis(tetramethylammonium) salts containing the dianion 11 have revealed that this type of receptor molecule is able to recognize the selected sugars in both organic and aqueous media. The carboxylate/pyridine-based receptor 11 exhibits in chloroform at least a 100-fold higher affinity for glucopyranosides than the previously described triarmed pyridine-based receptor 1, incorporating only neutral hydrogen-bonding sites. A substantial drop in the association constants is expectedly observed for an ester analogue of 11, compound 9. The dicarboxylate 11 is able to form complexes in water with methyl beta-D-glucopyranoside and D-cellobiose, with a preference for the disaccharide. The studies show the importance of charge-reinforced hydrogen bonds in the recognition of carbohydrates.

  10. Mycobacterium tuberculosis Activates Human Macrophage Peroxisome Proliferator-Activated Receptor γ Linking Mannose Receptor Recognition to Regulation of Immune Responses

    PubMed Central

    Rajaram, Murugesan V. S.; Brooks, Michelle N.; Morris, Jessica D.; Torrelles, Jordi B.; Azad, Abul K.; Schlesinger, Larry S.

    2010-01-01

    Mycobacterium tuberculosis enhances its survival in macrophages by suppressing immune responses in part through its complex cell wall structures. Peroxisome proliferator-activated receptor γ (PPARγ), a nuclear receptor superfamily member, is a transcriptional factor that regulates inflammation and has high expression in alternatively activated alveolar macrophages and macrophage-derived foam cells, both cell types relevant to tuberculosis pathogenesis. In this study, we show that virulent M. tuberculosis and its cell wall mannose-capped lipoarabinomannan induce PPARγ expression through a macrophage mannose receptor-dependent pathway. When activated, PPARγ promotes IL-8 and cyclooxygenase 2 expression, a process modulated by a PPARγ agonist or antagonist. Upstream, MAPK-p38 mediates cytosolic phospholipase A2 activation, which is required for PPARγ ligand production. The induced IL-8 response mediated by mannose-capped lipoarabinomannan and the mannose receptor is independent of TLR2 and NF-κB activation. In contrast, the attenuated Mycobacterium bovis bacillus Calmette-Guérin induces less PPARγ and preferentially uses the NF-κB–mediated pathway to induce IL-8 production. Finally, PPARγ knockdown in human macrophages enhances TNF production and controls the intracellular growth of M. tuberculosis. These data identify a new molecular pathway that links engagement of the mannose receptor, an important pattern recognition receptor for M. tuberculosis, with PPARγ activation, which regulates the macrophage inflammatory response, thereby playing a role in tuberculosis pathogenesis. PMID:20554962

  11. Molecular basis for LDL receptor recognition by PCSK9.

    PubMed

    Kwon, Hyock Joo; Lagace, Thomas A; McNutt, Markey C; Horton, Jay D; Deisenhofer, Johann

    2008-02-12

    Proprotein convertase subtilisin/kexin type 9 (PCSK9) posttranslationally regulates hepatic low-density lipoprotein receptors (LDLRs) by binding to LDLRs on the cell surface, leading to their degradation. The binding site of PCSK9 has been localized to the epidermal growth factor-like repeat A (EGF-A) domain of the LDLR. Here, we describe the crystal structure of a complex between PCSK9 and the EGF-A domain of the LDLR. The binding site for the LDLR EGF-A domain resides on the surface of PCSK9's subtilisin-like catalytic domain containing Asp-374, a residue for which a gain-of-function mutation (Asp-374-Tyr) increases the affinity of PCSK9 toward LDLR and increases plasma LDL-cholesterol (LDL-C) levels in humans. The binding surface on PCSK9 is distant from its catalytic site, and the EGF-A domain makes no contact with either the C-terminal domain or the prodomain. Point mutations in PCSK9 that altered key residues contributing to EGF-A binding (Arg-194 and Phe-379) greatly diminished binding to the LDLR's extracellular domain. The structure of PCSK9 in complex with the LDLR EGF-A domain defines potential therapeutic target sites for blocking agents that could interfere with this interaction in vivo, thereby increasing LDLR function and reducing plasma LDL-C levels.

  12. Structural basis of transcobalamin recognition by human CD320 receptor

    PubMed Central

    Alam, Amer; Woo, Jae-Sung; Schmitz, Jennifer; Prinz, Bernadette; Root, Katharina; Chen, Fan; Bloch, Joël S.; Zenobi, Renato; Locher, Kaspar P.

    2016-01-01

    Cellular uptake of vitamin B12 (cobalamin) requires capture of transcobalamin (TC) from the plasma by CD320, a ubiquitous cell surface receptor of the LDLR family. Here we present the crystal structure of human holo-TC in complex with the extracellular domain of CD320, visualizing the structural basis of the TC-CD320 interaction. The observed interaction chemistry can rationalize the high affinity of CD320 for TC and lack of haptocorrin binding. The in vitro affinity and complex stability of TC-CD320 were quantitated using a solid-phase binding assay and thermostability analysis. Stable complexes with TC were also observed for the disease-causing CD320ΔE88 mutant and for the isolated LDLR-A2 domain. We also determined the structure of the TC-CD320ΔE88 complex, which revealed only minor changes compared with the wild-type complex. Finally, we demonstrate significantly reduced in vitro affinity of TC for CD320 at low pH, recapitulating the proposed ligand release during the endocytic pathway. PMID:27411955

  13. Structural basis of transcobalamin recognition by human CD320 receptor

    NASA Astrophysics Data System (ADS)

    Alam, Amer; Woo, Jae-Sung; Schmitz, Jennifer; Prinz, Bernadette; Root, Katharina; Chen, Fan; Bloch, Joël S.; Zenobi, Renato; Locher, Kaspar P.

    2016-07-01

    Cellular uptake of vitamin B12 (cobalamin) requires capture of transcobalamin (TC) from the plasma by CD320, a ubiquitous cell surface receptor of the LDLR family. Here we present the crystal structure of human holo-TC in complex with the extracellular domain of CD320, visualizing the structural basis of the TC-CD320 interaction. The observed interaction chemistry can rationalize the high affinity of CD320 for TC and lack of haptocorrin binding. The in vitro affinity and complex stability of TC-CD320 were quantitated using a solid-phase binding assay and thermostability analysis. Stable complexes with TC were also observed for the disease-causing CD320ΔE88 mutant and for the isolated LDLR-A2 domain. We also determined the structure of the TC-CD320ΔE88 complex, which revealed only minor changes compared with the wild-type complex. Finally, we demonstrate significantly reduced in vitro affinity of TC for CD320 at low pH, recapitulating the proposed ligand release during the endocytic pathway.

  14. International Union of Basic and Clinical Pharmacology. XCVI. Pattern recognition receptors in health and disease.

    PubMed

    Bryant, Clare E; Orr, Selinda; Ferguson, Brian; Symmons, Martyn F; Boyle, Joseph P; Monie, Tom P

    2015-01-01

    Since the discovery of Toll, in the fruit fly Drosophila melanogaster, as the first described pattern recognition receptor (PRR) in 1996, many families of these receptors have been discovered and characterized. PRRs play critically important roles in pathogen recognition to initiate innate immune responses that ultimately link to the generation of adaptive immunity. Activation of PRRs leads to the induction of immune and inflammatory genes, including proinflammatory cytokines and chemokines. It is increasingly clear that many PRRs are linked to a range of inflammatory, infectious, immune, and chronic degenerative diseases. Several drugs to modulate PRR activity are already in clinical trials and many more are likely to appear in the near future. Here, we review the different families of mammalian PRRs, the ligands they recognize, the mechanisms of activation, their role in disease, and the potential of targeting these proteins to develop the anti-inflammatory therapeutics of the future.

  15. Bacterial and fungal pattern recognition receptors in homologous innate signaling pathways of insects and mammals

    PubMed Central

    Stokes, Bethany A.; Yadav, Shruti; Shokal, Upasana; Smith, L. C.; Eleftherianos, Ioannis

    2015-01-01

    In response to bacterial and fungal infections in insects and mammals, distinct families of innate immune pattern recognition receptors (PRRs) initiate highly complex intracellular signaling cascades. Those cascades induce a variety of immune functions that restrain the spread of microbes in the host. Insect and mammalian innate immune receptors include molecules that recognize conserved microbial molecular patterns. Innate immune recognition leads to the recruitment of adaptor molecules forming multi-protein complexes that include kinases, transcription factors, and other regulatory molecules. Innate immune signaling cascades induce the expression of genes encoding antimicrobial peptides and other key factors that mount and regulate the immune response against microbial challenge. In this review, we summarize our current understanding of the bacterial and fungal PRRs for homologous innate signaling pathways of insects and mammals in an effort to provide a framework for future studies. PMID:25674081

  16. International Union of Basic and Clinical Pharmacology. XCVI. Pattern Recognition Receptors in Health and Disease

    PubMed Central

    Orr, Selinda; Ferguson, Brian; Symmons, Martyn F.; Boyle, Joseph P.; Monie, Tom P.

    2015-01-01

    Since the discovery of Toll, in the fruit fly Drosophila melanogaster, as the first described pattern recognition receptor (PRR) in 1996, many families of these receptors have been discovered and characterized. PRRs play critically important roles in pathogen recognition to initiate innate immune responses that ultimately link to the generation of adaptive immunity. Activation of PRRs leads to the induction of immune and inflammatory genes, including proinflammatory cytokines and chemokines. It is increasingly clear that many PRRs are linked to a range of inflammatory, infectious, immune, and chronic degenerative diseases. Several drugs to modulate PRR activity are already in clinical trials and many more are likely to appear in the near future. Here, we review the different families of mammalian PRRs, the ligands they recognize, the mechanisms of activation, their role in disease, and the potential of targeting these proteins to develop the anti-inflammatory therapeutics of the future. PMID:25829385

  17. Non-classical amine recognition evolved in a large clade of olfactory receptors

    PubMed Central

    Li, Qian; Tachie-Baffour, Yaw; Liu, Zhikai; Baldwin, Maude W; Kruse, Andrew C; Liberles, Stephen D

    2015-01-01

    Biogenic amines are important signaling molecules, and the structural basis for their recognition by G Protein-Coupled Receptors (GPCRs) is well understood. Amines are also potent odors, with some activating olfactory trace amine-associated receptors (TAARs). Here, we report that teleost TAARs evolved a new way to recognize amines in a non-classical orientation. Chemical screens de-orphaned eleven zebrafish TAARs, with agonists including serotonin, histamine, tryptamine, 2-phenylethylamine, putrescine, and agmatine. Receptors from different clades contact ligands through aspartates on transmembrane α-helices III (canonical Asp3.32) or V (non-canonical Asp5.42), and diamine receptors contain both aspartates. Non-classical monoamine recognition evolved in two steps: an ancestral TAAR acquired Asp5.42, gaining diamine sensitivity, and subsequently lost Asp3.32. Through this transformation, the fish olfactory system dramatically expanded its capacity to detect amines, ecologically significant aquatic odors. The evolution of a second, alternative solution for amine detection by olfactory receptors highlights the tremendous structural versatility intrinsic to GPCRs. DOI: http://dx.doi.org/10.7554/eLife.10441.001 PMID:26519734

  18. Serotonin 2a Receptor and Serotonin 1a Receptor Interact Within the Medial Prefrontal Cortex During Recognition Memory in Mice

    PubMed Central

    Morici, Juan F.; Ciccia, Lucia; Malleret, Gaël; Gingrich, Jay A.; Bekinschtein, Pedro; Weisstaub, Noelia V.

    2015-01-01

    Episodic memory, can be defined as the memory for unique events. The serotonergic system one of the main neuromodulatory systems in the brain appears to play a role in it. The serotonin 2a receptor (5-HT2aR) one of the principal post-synaptic receptors for 5-HT in the brain, is involved in neuropsychiatric and neurological disorders associated with memory deficits. Recognition memory can be defined as the ability to recognize if a particular event or item was previously encountered and is thus considered, under certain conditions, a form of episodic memory. As human data suggest that a constitutively decrease of 5-HT2A signaling might affect episodic memory performance we decided to compare the performance of mice with disrupted 5-HT2aR signaling (htr2a−/−) with wild type (htr2a+/+) littermates in different recognition memory and working memory tasks that differed in the level of proactive interference. We found that ablation of 5-HT2aR signaling throughout development produces a deficit in tasks that cannot be solved by single item strategy suggesting that 5-HT2aR signaling is involved in interference resolution. We also found that in the absence of 5-HT2aR signaling serotonin has a deleterious effect on recognition memory retrieval through the activation of 5-HT1aR in the medial prefrontal cortex. PMID:26779016

  19. Pattern recognition receptors in zebrafish provide functional and evolutionary insight into innate immune signaling pathways

    PubMed Central

    Li, Yajuan; Li, Yuelong; Cao, Xiaocong; Jin, Xiangyu; Jin, Tengchuan

    2017-01-01

    Pattern recognition receptors (PRRs) and their signaling pathways have essential roles in recognizing various components of pathogens as well as damaged cells and triggering inflammatory responses that eliminate invading microorganisms and damaged cells. The zebrafish relies heavily on these primary defense mechanisms against pathogens. Here, we review the major PRR signaling pathways in the zebrafish innate immune system and compare these signaling pathways in zebrafish and humans to reveal their evolutionary relationship and better understand their innate immune defense mechanisms. PMID:27721456

  20. Improved localization of cellular membrane receptors using combined fluorescence microscopy and simultaneous topography and recognition imaging.

    PubMed

    Duman, M; Pfleger, M; Zhu, R; Rankl, C; Chtcheglova, L A; Neundlinger, I; Bozna, B L; Mayer, B; Salio, M; Shepherd, D; Polzella, P; Moertelmaier, M; Kada, G; Ebner, A; Dieudonne, M; Schütz, G J; Cerundolo, V; Kienberger, F; Hinterdorfer, P

    2010-03-19

    The combination of fluorescence microscopy and atomic force microscopy has a great potential in single-molecule-detection applications, overcoming many of the limitations coming from each individual technique. Here we present a new platform of combined fluorescence and simultaneous topography and recognition imaging (TREC) for improved localization of cellular receptors. Green fluorescent protein (GFP) labeled human sodium-glucose cotransporter (hSGLT1) expressed Chinese Hamster Ovary (CHO) cells and endothelial cells (MyEnd) from mouse myocardium stained with phalloidin-rhodamine were used as cell systems to study AFM topography and fluorescence microscopy on the same surface area. Topographical AFM images revealed membrane features such as lamellipodia, cytoskeleton fibers, F-actin filaments and small globular structures with heights ranging from 20 to 30 nm. Combined fluorescence and TREC imaging was applied to detect density, distribution and localization of YFP-labeled CD1d molecules on alpha-galactosylceramide (alphaGalCer)-loaded THP1 cells. While the expression level, distribution and localization of CD1d molecules on THP1 cells were detected with fluorescence microscopy, the nanoscale distribution of binding sites was investigated with molecular recognition imaging by using a chemically modified AFM tip. Using TREC on the inverted light microscope, the recognition sites of cell receptors were detected in recognition images with domain sizes ranging from approximately 25 to approximately 160 nm, with the smaller domains corresponding to a single CD1d molecule.

  1. Improved localization of cellular membrane receptors using combined fluorescence microscopy and simultaneous topography and recognition imaging

    NASA Astrophysics Data System (ADS)

    Duman, M.; Pfleger, M.; Zhu, R.; Rankl, C.; Chtcheglova, L. A.; Neundlinger, I.; Bozna, B. L.; Mayer, B.; Salio, M.; Shepherd, D.; Polzella, P.; Moertelmaier, M.; Kada, G.; Ebner, A.; Dieudonne, M.; Schütz, G. J.; Cerundolo, V.; Kienberger, F.; Hinterdorfer, P.

    2010-03-01

    The combination of fluorescence microscopy and atomic force microscopy has a great potential in single-molecule-detection applications, overcoming many of the limitations coming from each individual technique. Here we present a new platform of combined fluorescence and simultaneous topography and recognition imaging (TREC) for improved localization of cellular receptors. Green fluorescent protein (GFP) labeled human sodium-glucose cotransporter (hSGLT1) expressed Chinese Hamster Ovary (CHO) cells and endothelial cells (MyEnd) from mouse myocardium stained with phalloidin-rhodamine were used as cell systems to study AFM topography and fluorescence microscopy on the same surface area. Topographical AFM images revealed membrane features such as lamellipodia, cytoskeleton fibers, F-actin filaments and small globular structures with heights ranging from 20 to 30 nm. Combined fluorescence and TREC imaging was applied to detect density, distribution and localization of YFP-labeled CD1d molecules on α-galactosylceramide (αGalCer)-loaded THP1 cells. While the expression level, distribution and localization of CD1d molecules on THP1 cells were detected with fluorescence microscopy, the nanoscale distribution of binding sites was investigated with molecular recognition imaging by using a chemically modified AFM tip. Using TREC on the inverted light microscope, the recognition sites of cell receptors were detected in recognition images with domain sizes ranging from ~ 25 to ~ 160 nm, with the smaller domains corresponding to a single CD1d molecule.

  2. NMDA Receptor Antagonist Ketamine Distorts Object Recognition by Reducing Feedback to Early Visual Cortex.

    PubMed

    van Loon, Anouk M; Fahrenfort, Johannes J; van der Velde, Bauke; Lirk, Philipp B; Vulink, Nienke C C; Hollmann, Markus W; Scholte, H Steven; Lamme, Victor A F

    2016-05-01

    It is a well-established fact that top-down processes influence neural representations in lower-level visual areas. Electrophysiological recordings in monkeys as well as theoretical models suggest that these top-down processes depend on NMDA receptor functioning. However, this underlying neural mechanism has not been tested in humans. We used fMRI multivoxel pattern analysis to compare the neural representations of ambiguous Mooney images before and after they were recognized with their unambiguous grayscale version. Additionally, we administered ketamine, an NMDA receptor antagonist, to interfere with this process. Our results demonstrate that after recognition, the pattern of brain activation elicited by a Mooney image is more similar to that of its easily recognizable grayscale version than to the pattern evoked by the identical Mooney image before recognition. Moreover, recognition of Mooney images decreased mean response; however, neural representations of separate images became more dissimilar. So from the neural perspective, unrecognizable Mooney images all "look the same", whereas recognized Mooneys look different. We observed these effects in posterior fusiform part of lateral occipital cortex and in early visual cortex. Ketamine distorted these effects of recognition, but in early visual cortex only. This suggests that top-down processes from higher- to lower-level visual areas might operate via an NMDA pathway.

  3. Effects of macromolecular crowding on the inhibition of virus assembly and virus-cell receptor recognition.

    PubMed

    Rincón, Verónica; Bocanegra, Rebeca; Rodríguez-Huete, Alicia; Rivas, Germán; Mateu, Mauricio G

    2011-02-02

    Biological fluids contain a very high total concentration of macromolecules that leads to volume exclusion by one molecule to another. Theory and experiment have shown that this condition, termed macromolecular crowding, can have significant effects on molecular recognition. However, the influence of molecular crowding on recognition events involving virus particles, and their inhibition by antiviral compounds, is virtually unexplored. Among these processes, capsid self-assembly during viral morphogenesis and capsid-cell receptor recognition during virus entry into cells are receiving increasing attention as targets for the development of new antiviral drugs. In this study, we have analyzed the effect of macromolecular crowding on the inhibition of these two processes by peptides. Macromolecular crowding led to a significant reduction in the inhibitory activity of: 1), a capsid-binding peptide and a small capsid protein domain that interfere with assembly of the human immunodeficiency virus capsid, and 2), a RGD-containing peptide able to block the interaction between foot-and-mouth disease virus and receptor molecules on the host cell membrane (in this case, the effect was dependent on the conditions used). The results, discussed in the light of macromolecular crowding theory, are relevant for a quantitative understanding of molecular recognition processes during virus infection and its inhibition.

  4. Common polymorphism in the oxytocin receptor gene (OXTR) is associated with human social recognition skills

    PubMed Central

    Skuse, David H.; Lori, Adriana; Cubells, Joseph F.; Lee, Irene; Conneely, Karen N.; Puura, Kaija; Lehtimäki, Terho; Binder, Elisabeth B.; Young, Larry J.

    2014-01-01

    The neuropeptides oxytocin and vasopressin are evolutionarily conserved regulators of social perception and behavior. Evidence is building that they are critically involved in the development of social recognition skills within rodent species, primates, and humans. We investigated whether common polymorphisms in the genes encoding the oxytocin and vasopressin 1a receptors influence social memory for faces. Our sample comprised 198 families, from the United Kingdom and Finland, in whom a single child had been diagnosed with high-functioning autism. Previous research has shown that impaired social perception, characteristic of autism, extends to the first-degree relatives of autistic individuals, implying heritable risk. Assessments of face recognition memory, discrimination of facial emotions, and direction of gaze detection were standardized for age (7–60 y) and sex. A common SNP in the oxytocin receptor (rs237887) was strongly associated with recognition memory in combined probands, parents, and siblings after correction for multiple comparisons. Homozygotes for the ancestral A allele had impairments in the range −0.6 to −1.15 SD scores, irrespective of their diagnostic status. Our findings imply that a critical role for the oxytocin system in social recognition has been conserved across perceptual boundaries through evolution, from olfaction in rodents to visual memory in humans. PMID:24367110

  5. An Overview of Pathogen Recognition Receptors for Innate Immunity in Dental Pulp

    PubMed Central

    Jang, Ji-Hyun; Shin, Hee Woong; Lee, Jung Min; Lee, Hyeon-Woo; Kim, Eun-Cheol; Park, Sang Hyuk

    2015-01-01

    Pathogen recognition receptors (PRRs) are a class of germ line-encoded receptors that recognize pathogen-associated molecular patterns (PAMPs). The activation of PRRs is crucial for the initiation of innate immunity, which plays a key role in first-line defense until more specific adaptive immunity is developed. PRRs differ in the signaling cascades and host responses activated by their engagement and in their tissue distribution. Currently identified PRR families are the Toll-like receptors (TLRs), the C-type lectin receptors (CLRs), the nucleotide-binding oligomerization domain-like receptors (NLRs), the retinoic acid-inducible gene-I-like receptors (RLRs), and the AIM2-like receptor (ALR). The environment of the dental pulp is substantially different from that of other tissues of the body. Dental pulp resides in a low compliance root canal system that limits the expansion of pulpal tissues during inflammatory processes. An understanding of the PRRs in dental pulp is important for immunomodulation and hence for developing therapeutic targets in the field of endodontics. Here we comprehensively review recent finding on the PRRs and the mechanisms by which innate immunity is activated. We focus on the PRRs expressed on dental pulp and periapical tissues and their role in dental pulp inflammation. PMID:26576076

  6. Structural Insights into Cargo Recognition by the Yeast PTS1 Receptor*

    PubMed Central

    Hagen, Stefanie; Drepper, Friedel; Fischer, Sven; Fodor, Krisztian; Passon, Daniel; Platta, Harald W.; Zenn, Michael; Schliebs, Wolfgang; Girzalsky, Wolfgang; Wilmanns, Matthias; Warscheid, Bettina; Erdmann, Ralf

    2015-01-01

    The peroxisomal matrix protein import is facilitated by cycling import receptors that shuttle between the cytosol and the peroxisomal membrane. The import receptor Pex5p mediates the import of proteins harboring a peroxisomal targeting signal of type I (PTS1). Purified recombinant Pex5p forms a dimeric complex with the PTS1-protein Pcs60p in vitro with a KD of 0.19 μm. To analyze the structural basis for receptor-cargo recognition, the PTS1 and adjacent amino acids of Pcs60p were systematically scanned for Pex5p binding by an in vitro site-directed photo-cross-linking approach. The cross-linked binding regions of the receptor were subsequently identified by high resolution mass spectrometry. Most cross-links were found with TPR6, TPR7, as well as the 7C-loop of Pex5p. Surface plasmon resonance analysis revealed a bivalent interaction mode for Pex5p and Pcs60p. Interestingly, Pcs60p lacking its C-terminal tripeptide sequence was efficiently cross-linked to the same regions of Pex5p. The KD value of the interaction of truncated Pcs60p and Pex5p was in the range of 7.7 μm. Isothermal titration calorimetry and surface plasmon resonance measurements revealed a monovalent binding mode for the interaction of Pex5p and Pcs60p lacking the PTS1. Our data indicate that Pcs60p contains a second contact site for its receptor Pex5p, beyond the C-terminal tripeptide. The physiological relevance of the ancillary binding region was supported by in vivo import studies. The bivalent binding mode might be explained by a two-step concept as follows: first, cargo recognition and initial tethering by the PTS1-receptor Pex5p; second, lock-in of receptor and cargo. PMID:26359497

  7. Dopamine D1 receptor stimulation modulates the formation and retrieval of novel object recognition memory: Role of the prelimbic cortex

    PubMed Central

    Pezze, Marie A.; Marshall, Hayley J.; Fone, Kevin C.F.; Cassaday, Helen J.

    2015-01-01

    Previous studies have shown that dopamine D1 receptor antagonists impair novel object recognition memory but the effects of dopamine D1 receptor stimulation remain to be determined. This study investigated the effects of the selective dopamine D1 receptor agonist SKF81297 on acquisition and retrieval in the novel object recognition task in male Wistar rats. SKF81297 (0.4 and 0.8 mg/kg s.c.) given 15 min before the sampling phase impaired novel object recognition evaluated 10 min or 24 h later. The same treatments also reduced novel object recognition memory tested 24 h after the sampling phase and when given 15 min before the choice session. These data indicate that D1 receptor stimulation modulates both the encoding and retrieval of object recognition memory. Microinfusion of SKF81297 (0.025 or 0.05 μg/side) into the prelimbic sub-region of the medial prefrontal cortex (mPFC) in this case 10 min before the sampling phase also impaired novel object recognition memory, suggesting that the mPFC is one important site mediating the effects of D1 receptor stimulation on visual recognition memory. PMID:26277743

  8. Dopamine D1 receptor stimulation modulates the formation and retrieval of novel object recognition memory: Role of the prelimbic cortex.

    PubMed

    Pezze, Marie A; Marshall, Hayley J; Fone, Kevin C F; Cassaday, Helen J

    2015-11-01

    Previous studies have shown that dopamine D1 receptor antagonists impair novel object recognition memory but the effects of dopamine D1 receptor stimulation remain to be determined. This study investigated the effects of the selective dopamine D1 receptor agonist SKF81297 on acquisition and retrieval in the novel object recognition task in male Wistar rats. SKF81297 (0.4 and 0.8 mg/kg s.c.) given 15 min before the sampling phase impaired novel object recognition evaluated 10 min or 24 h later. The same treatments also reduced novel object recognition memory tested 24 h after the sampling phase and when given 15 min before the choice session. These data indicate that D1 receptor stimulation modulates both the encoding and retrieval of object recognition memory. Microinfusion of SKF81297 (0.025 or 0.05 μg/side) into the prelimbic sub-region of the medial prefrontal cortex (mPFC) in this case 10 min before the sampling phase also impaired novel object recognition memory, suggesting that the mPFC is one important site mediating the effects of D1 receptor stimulation on visual recognition memory.

  9. Preferential recognition of avian-like receptors in human influenza A H7N9 viruses.

    PubMed

    Xu, Rui; de Vries, Robert P; Zhu, Xueyong; Nycholat, Corwin M; McBride, Ryan; Yu, Wenli; Paulson, James C; Wilson, Ian A

    2013-12-06

    The 2013 outbreak of avian-origin H7N9 influenza in eastern China has raised concerns about its ability to transmit in the human population. The hemagglutinin glycoprotein of most human H7N9 viruses carries Leu(226), a residue linked to adaptation of H2N2 and H3N2 pandemic viruses to human receptors. However, glycan array analysis of the H7 hemagglutinin reveals negligible binding to humanlike α2-6-linked receptors and strong preference for a subset of avian-like α2-3-linked glycans recognized by all avian H7 viruses. Crystal structures of H7N9 hemagglutinin and six hemagglutinin-glycan complexes have elucidated the structural basis for preferential recognition of avian-like receptors. These findings suggest that the current human H7N9 viruses are poorly adapted for efficient human-to-human transmission.

  10. An Update on PYRIN Domain-Containing Pattern Recognition Receptors: From Immunity to Pathology

    PubMed Central

    Ratsimandresy, Rojo A.; Dorfleutner, Andrea; Stehlik, Christian

    2013-01-01

    Cytosolic pattern recognition receptors (PRRs) sense a wide range of endogenous danger-associated molecular patterns as well as exogenous pathogen-associated molecular patterns. In particular, Nod-like receptors containing a pyrin domain (PYD), called NLRPs, and AIM2-like receptors (ALRs) have been shown to play a critical role in host defense by facilitating clearance of pathogens and maintaining a healthy gut microflora. NLRPs and ALRs both encode a PYD, which is crucial for relaying signals that result in an efficient innate immune response through activation of several key innate immune signaling pathways. However, mutations in these PRRs have been linked to the development of auto-inflammatory and autoimmune diseases. In addition, they have been implicated in metabolic diseases. In this review, we summarize the function of PYD-containing NLRPs and ALRs and address their contribution to innate immunity, host defense, and immune-linked diseases. PMID:24367371

  11. The targeted recognition of Lactococcus lactis phages to their polysaccharide receptors.

    PubMed

    McCabe, Orla; Spinelli, Silvia; Farenc, Carine; Labbé, Myriam; Tremblay, Denise; Blangy, Stéphanie; Oscarson, Stefan; Moineau, Sylvain; Cambillau, Christian

    2015-05-01

    Each phage infects a limited number of bacterial strains through highly specific interactions of the receptor-binding protein (RBP) at the tip of phage tail and the receptor at the bacterial surface. Lactococcus lactis is covered with a thin polysaccharide pellicle (hexasaccharide repeating units), which is used by a subgroup of phages as a receptor. Using L. lactis and phage 1358 as a model, we investigated the interaction between the phage RBP and the pellicle hexasaccharide of the host strain. A core trisaccharide (TriS), derived from the pellicle hexasaccharide repeating unit, was chemically synthesised, and the crystal structure of the RBP/TriS complex was determined. This provided unprecedented structural details of RBP/receptor site-specific binding. The complete hexasaccharide repeating unit was modelled and found to aptly fit the extended binding site. The specificity observed in in vivo phage adhesion assays could be interpreted in view of the reported structure. Therefore, by combining synthetic carbohydrate chemistry, X-ray crystallography and phage plaquing assays, we suggest that phage adsorption results from distinct recognition of the RBP towards the core TriS or the remaining residues of the hexasacchride receptor. This study provides a novel insight into the adsorption process of phages targeting saccharides as their receptors.

  12. Hippocampal NMDA receptors are involved in rats' spontaneous object recognition only under high memory load condition.

    PubMed

    Sugita, Manami; Yamada, Kazuo; Iguchi, Natsumi; Ichitani, Yukio

    2015-10-22

    The possible involvement of hippocampal N-methyl-D-aspartate (NMDA) receptors in spontaneous object recognition was investigated in rats under different memory load conditions. We first estimated rats' object memory span using 3-5 objects in "Different Objects Task (DOT)" in order to confirm the highest memory load condition in object recognition memory. Rats were allowed to explore a field in which 3 (3-DOT), 4 (4-DOT), or 5 (5-DOT) different objects were presented. After a delay period, they were placed again in the same field in which one of the sample objects was replaced by another object, and their object exploration behavior was analyzed. Rats could differentiate the novel object from the familiar ones in 3-DOT and 4-DOT but not in 5-DOT, suggesting that rats' object memory span was about 4. Then, we examined the effects of hippocampal AP5 infusion on performance in both 2-DOT (2 different objects were used) and 4-DOT. The drug treatment before the sample phase impaired performance only in 4-DOT. These results suggest that hippocampal NMDA receptors play a critical role in spontaneous object recognition only when the memory load is high.

  13. Expanding the universe of cytokines and pattern recognition receptors: galectins and glycans in innate immunity.

    PubMed

    Cerliani, Juan P; Stowell, Sean R; Mascanfroni, Iván D; Arthur, Connie M; Cummings, Richard D; Rabinovich, Gabriel A

    2011-02-01

    Effective immunity relies on the recognition of pathogens and tumors by innate immune cells through diverse pattern recognition receptors (PRRs) that lead to initiation of signaling processes and secretion of pro- and anti-inflammatory cytokines. Galectins, a family of endogenous lectins widely expressed in infected and neoplastic tissues have emerged as part of the portfolio of soluble mediators and pattern recognition receptors responsible for eliciting and controlling innate immunity. These highly conserved glycan-binding proteins can control immune cell processes through binding to specific glycan structures on pathogens and tumors or by acting intracellularly via modulation of selective signaling pathways. Recent findings demonstrate that various galectin family members influence the fate and physiology of different innate immune cells including polymorphonuclear neutrophils, mast cells, macrophages, and dendritic cells. Moreover, several pathogens may actually utilize galectins as a mechanism of host invasion. In this review, we aim to highlight and integrate recent discoveries that have led to our current understanding of the role of galectins in host-pathogen interactions and innate immunity. Challenges for the future will embrace the rational manipulation of galectin-glycan interactions to instruct and shape innate immunity during microbial infections, inflammation, and cancer.

  14. A Temporally Distinct Role for Group I and Group II Metabotropic Glutamate Receptors in Object Recognition Memory

    ERIC Educational Resources Information Center

    Brown, Malcolm Watson; Warburton, Elizabeth Clea; Barker, Gareth Robert Isaac; Bashir, Zafar Iqbal

    2006-01-01

    Recognition memory, involving the ability to discriminate between a novel and familiar object, depends on the integrity of the perirhinal cortex (PRH). Glutamate, the main excitatory neurotransmitter in the cortex, is essential for many types of memory processes. Of the subtypes of glutamate receptor, metabotropic receptors (mGluRs) have received…

  15. ɣδ T cell receptor ligands and modes of antigen recognition

    PubMed Central

    Champagne, Eric

    2011-01-01

    T lymphocytes expressing the γδ-type of T cell receptors for antigens contribute to all aspects of immune responses, including defenses against viruses, bacteria, parasites and tumors, allergy and autoimmunity. Multiple subsets have been individualized in humans as well as in mice and they appear to recognize in a TCR-dependent manner antigens as diverse as small non-peptidic molecules, soluble or membrane-anchored polypeptides and molecules related to MHC antigens on cell surfaces, implying diverse modes of antigen recognition. We review here the γδ TCR ligands which have been identified along the years and their characteristics, with emphasis on a few systems which have been extensively studied such as human γδ T cells responding to phosphoantigens or murine γδ T cells activated by allogeneic MHC antigens. We discuss a speculative model of antigen recognition involving simultaneous TCR recognition of MHC-like and non-MHC ligands which could fit with most available data and shares many similarities with the classical model of MHC-restricted antigen recognition for peptides or lipids by T cells subsets with αβ-type TCRs. PMID:21298486

  16. Evidence for halogen bond covalency in acyclic and interlocked halogen-bonding receptor anion recognition

    DOE PAGES

    Robinson, Sean W.; Mustoe, Chantal L.; White, Nicholas G.; ...

    2014-12-05

    The synthesis and anion binding properties of novel halogen-bonding (XB) bis-iodotriazole-pyridinium-containing acyclic and [2]catenane anion host systems are described. The XB acyclic receptor displays selectivity for acetate over halides with enhanced anion recognition properties compared to the analogous hydrogen-bonding (HB) acyclic receptor. A reversal in halide selectivity is observed in the XB [2]catenane, in comparison to the acyclic XB receptor, due to the interlocked host’s unique three-dimensional binding cavity, and no binding is observed for oxoanions. Notable halide anion association constant values determined for the [2]catenane in competitive organic–aqueous solvent mixtures demonstrate considerable enhancement of anion recognition as compared tomore » the HB catenane analogue. X-ray crystallographic analysis of a series of halide catenane complexes reveal strong XB interactions in the solid state. These interactions were studied using Cl and Br K-edge X-ray Absorption Spectroscopy (XAS) indicating intense pre-edge features characteristic of charge transfer from the halide to its bonding partner (σAX←X–* ← X1s), and providing a direct measure of the degree of covalency in the halogen bond(s). Lastly, the data reveal that the degree of covalency is similar to that which is observed in transition metal coordinate covalent bonds. These results are supported by DFT results, which correlate well with the experimental data.« less

  17. Structural basis of pathogen recognition by an integrated HMA domain in a plant NLR immune receptor

    PubMed Central

    Maqbool, A; Saitoh, H; Franceschetti, M; Stevenson, CEM; Uemura, A; Kanzaki, H; Kamoun, S; Terauchi, R; Banfield, MJ

    2015-01-01

    Plants have evolved intracellular immune receptors to detect pathogen proteins known as effectors. How these immune receptors detect effectors remains poorly understood. Here we describe the structural basis for direct recognition of AVR-Pik, an effector from the rice blast pathogen, by the rice intracellular NLR immune receptor Pik. AVR-PikD binds a dimer of the Pikp-1 HMA integrated domain with nanomolar affinity. The crystal structure of the Pikp-HMA/AVR-PikD complex enabled design of mutations to alter protein interaction in yeast and in vitro, and perturb effector-mediated response both in a rice cultivar containing Pikp and upon expression of AVR-PikD and Pikp in the model plant Nicotiana benthamiana. These data reveal the molecular details of a recognition event, mediated by a novel integrated domain in an NLR, which initiates a plant immune response and resistance to rice blast disease. Such studies underpin novel opportunities for engineering disease resistance to plant pathogens in staple food crops. DOI: http://dx.doi.org/10.7554/eLife.08709.001 PMID:26304198

  18. Evidence for halogen bond covalency in acyclic and interlocked halogen-bonding receptor anion recognition

    SciTech Connect

    Robinson, Sean W.; Mustoe, Chantal L.; White, Nicholas G.; Brown, Asha; Thompson, Amber L.; Kennepohl, Pierre; Beer, Paul D.

    2014-12-05

    The synthesis and anion binding properties of novel halogen-bonding (XB) bis-iodotriazole-pyridinium-containing acyclic and [2]catenane anion host systems are described. The XB acyclic receptor displays selectivity for acetate over halides with enhanced anion recognition properties compared to the analogous hydrogen-bonding (HB) acyclic receptor. A reversal in halide selectivity is observed in the XB [2]catenane, in comparison to the acyclic XB receptor, due to the interlocked host’s unique three-dimensional binding cavity, and no binding is observed for oxoanions. Notable halide anion association constant values determined for the [2]catenane in competitive organic–aqueous solvent mixtures demonstrate considerable enhancement of anion recognition as compared to the HB catenane analogue. X-ray crystallographic analysis of a series of halide catenane complexes reveal strong XB interactions in the solid state. These interactions were studied using Cl and Br K-edge X-ray Absorption Spectroscopy (XAS) indicating intense pre-edge features characteristic of charge transfer from the halide to its bonding partner (σAX←X–* ← X1s), and providing a direct measure of the degree of covalency in the halogen bond(s). Lastly, the data reveal that the degree of covalency is similar to that which is observed in transition metal coordinate covalent bonds. These results are supported by DFT results, which correlate well with the experimental data.

  19. Structural basis of pathogen recognition by an integrated HMA domain in a plant NLR immune receptor.

    PubMed

    Maqbool, A; Saitoh, H; Franceschetti, M; Stevenson, C E M; Uemura, A; Kanzaki, H; Kamoun, S; Terauchi, R; Banfield, M J

    2015-08-25

    Plants have evolved intracellular immune receptors to detect pathogen proteins known as effectors. How these immune receptors detect effectors remains poorly understood. Here we describe the structural basis for direct recognition of AVR-Pik, an effector from the rice blast pathogen, by the rice intracellular NLR immune receptor Pik. AVR-PikD binds a dimer of the Pikp-1 HMA integrated domain with nanomolar affinity. The crystal structure of the Pikp-HMA/AVR-PikD complex enabled design of mutations to alter protein interaction in yeast and in vitro, and perturb effector-mediated response both in a rice cultivar containing Pikp and upon expression of AVR-PikD and Pikp in the model plant Nicotiana benthamiana. These data reveal the molecular details of a recognition event, mediated by a novel integrated domain in an NLR, which initiates a plant immune response and resistance to rice blast disease. Such studies underpin novel opportunities for engineering disease resistance to plant pathogens in staple food crops.

  20. Interfamily transfer of a plant pattern-recognition receptor confers broad-spectrum bacterial resistance.

    PubMed

    Lacombe, Séverine; Rougon-Cardoso, Alejandra; Sherwood, Emma; Peeters, Nemo; Dahlbeck, Douglas; van Esse, H Peter; Smoker, Matthew; Rallapalli, Ghanasyam; Thomma, Bart P H J; Staskawicz, Brian; Jones, Jonathan D G; Zipfel, Cyril

    2010-04-01

    Plant diseases cause massive losses in agriculture. Increasing the natural defenses of plants may reduce the impact of phytopathogens on agricultural productivity. Pattern-recognition receptors (PRRs) detect microbes by recognizing conserved pathogen-associated molecular patterns (PAMPs). Although the overall importance of PAMP-triggered immunity for plant defense is established, it has not been used to confer disease resistance in crops. We report that activity of a PRR is retained after its transfer between two plant families. Expression of EFR (ref. 4), a PRR from the cruciferous plant Arabidopsis thaliana, confers responsiveness to bacterial elongation factor Tu in the solanaceous plants Nicotiana benthamiana and tomato (Solanum lycopersicum), making them more resistant to a range of phytopathogenic bacteria from different genera. Our results in controlled laboratory conditions suggest that heterologous expression of PAMP recognition systems could be used to engineer broad-spectrum disease resistance to important bacterial pathogens, potentially enabling more durable and sustainable resistance in the field.

  1. Perception of pathogenic or beneficial bacteria and their evasion of host immunity: pattern recognition receptors in the frontline

    PubMed Central

    Trdá, Lucie; Boutrot, Freddy; Claverie, Justine; Brulé, Daphnée; Dorey, Stephan; Poinssot, Benoit

    2015-01-01

    Plants are continuously monitoring the presence of microorganisms to establish an adapted response. Plants commonly use pattern recognition receptors (PRRs) to perceive microbe- or pathogen-associated molecular patterns (MAMPs/PAMPs) which are microorganism molecular signatures. Located at the plant plasma membrane, the PRRs are generally receptor-like kinases (RLKs) or receptor-like proteins (RLPs). MAMP detection will lead to the establishment of a plant defense program called MAMP-triggered immunity (MTI). In this review, we overview the RLKs and RLPs that assure early recognition and control of pathogenic or beneficial bacteria. We also highlight the crucial function of PRRs during plant-microbe interactions, with a special emphasis on the receptors of the bacterial flagellin and peptidoglycan. In addition, we discuss the multiple strategies used by bacteria to evade PRR-mediated recognition. PMID:25904927

  2. Molecular recognition of parathyroid hormone by its G protein-coupled receptor

    SciTech Connect

    Pioszak, Augen A.; Xu, H. Eric

    2008-08-07

    Parathyroid hormone (PTH) is central to calcium homeostasis and bone maintenance in vertebrates, and as such it has been used for treating osteoporosis. It acts primarily by binding to its receptor, PTH1R, a member of the class B G protein-coupled receptor (GPCR) family that also includes receptors for glucagon, calcitonin, and other therapeutically important peptide hormones. Despite considerable interest and much research, determining the structure of the receptor-hormone complex has been hindered by difficulties in purifying the receptor and obtaining diffraction-quality crystals. Here, we present a method for expression and purification of the extracellular domain (ECD) of human PTH1R engineered as a maltose-binding protein (MBP) fusion that readily crystallizes. The 1.95-{angstrom} structure of PTH bound to the MBP-PTH1R-ECD fusion reveals that PTH docks as an amphipathic helix into a central hydrophobic groove formed by a three-layer {alpha}-{beta}-{beta}{alpha} fold of the PTH1R ECD, resembling a hot dog in a bun. Conservation in the ECD scaffold and the helical structure of peptide hormones emphasizes this hot dog model as a general mechanism of hormone recognition common to class B GPCRs. Our findings reveal critical insights into PTH actions and provide a rational template for drug design that targets this hormone signaling pathway.

  3. [Role of antimicrobial peptides (AMP) and pattern recognition receptors (PRR) in the intestinal mucosa homeostasis].

    PubMed

    Lapis, Károly

    2009-11-22

    Homeostasis and integrity of bowel mucosa is assured by well controlled mechanical, biochemical and immunological mechanisms. First line of defense is presented by the antimicrobial peptides (AMP), which form a continuous layer on the bowel surface, produced by intestinal specific (Paneth) and non-specific epithelial cells. AMPs have a significant antimicrobial, antifungal and antiviral, as well as immunomodulatory effects. Next line of defense is the pattern recognition receptors (PRR), which allows identifying conservative molecular patterns of different pathogens, and starts antimicrobial and inflammatory mechanisms through gene-expression induction. We review the most recent knowledge and studies concerning these mechanisms.

  4. Blueprints of signaling interactions between pattern recognition receptors: implications for the design of vaccine adjuvants.

    PubMed

    Timmermans, Kim; Plantinga, Theo S; Kox, Matthijs; Vaneker, Michiel; Scheffer, Gert Jan; Adema, Gosse J; Joosten, Leo A B; Netea, Mihai G

    2013-03-01

    Innate immunity activation largely depends on recognition of microorganism structures by Pattern Recognition Receptors (PRRs). PRR downstream signaling results in production of pro- and anti-inflammatory cytokines and other mediators. Moreover, PRR engagement in antigen-presenting cells initiates the activation of adaptive immunity. Recent reports suggest that for the activation of innate immune responses and initiation of adaptive immunity, synergistic effects between two or more PRRs are necessary. No systematic analysis of the interaction between the major PRR pathways were performed to date. In this study, a systematical analysis of the interactions between PRR signaling pathways was performed. PBMCs derived from 10 healthy volunteers were stimulated with either a single PRR ligand or a combination of two PRR ligands. Known ligands for the major PRR families were used: Toll-like receptors (TLRs), C-type lectin receptors (CLRs), NOD-like receptors (NLRs), and RigI-helicases. After 24 h of incubation, production of tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1β), IL-6, and IL-10 was measured in supernatants by enzyme-linked immunosorbent assay (ELISA). The consistency of the PRR interactions (both inhibitory and synergistic) between the various individuals was assessed. A number of PRR-dependent signaling interactions were found to be consistent, both between individuals and with regard to multiple cytokines. The combinations of TLR2 and NOD2, TLR5 and NOD2, TLR5 and TLR3, and TLR5 and TLR9 acted as synergistic combinations. Surprisingly, inhibitory interactions between TLR4 and TLR2, TLR4 and Dectin-1, and TLR2 and TLR9 as well as TLR3 and TLR2 were observed. These consistent signaling interactions between PRR combinations may represent promising targets for immunomodulation and vaccine adjuvant development.

  5. Molecular Recognition of Corticotropin releasing Factor by Its G protein-coupled Receptor CRFR1

    SciTech Connect

    Pioszak, Augen A.; Parker, Naomi R.; Suino-Powell, Kelly; Xu, H. Eric

    2009-01-15

    The bimolecular interaction between corticotropin-releasing factor (CRF), a neuropeptide, and its type 1 receptor (CRFR1), a class B G-protein-coupled receptor (GPCR), is crucial for activation of the hypothalamic-pituitary-adrenal axis in response to stress, and has been a target of intense drug design for the treatment of anxiety, depression, and related disorders. As a class B GPCR, CRFR1 contains an N-terminal extracellular domain (ECD) that provides the primary ligand binding determinants. Here we present three crystal structures of the human CRFR1 ECD, one in a ligand-free form and two in distinct CRF-bound states. The CRFR1 ECD adopts the alpha-beta-betaalpha fold observed for other class B GPCR ECDs, but the N-terminal alpha-helix is significantly shorter and does not contact CRF. CRF adopts a continuous alpha-helix that docks in a hydrophobic surface of the ECD that is distinct from the peptide-binding site of other class B GPCRs, thereby providing a basis for the specificity of ligand recognition between CRFR1 and other class B GPCRs. The binding of CRF is accompanied by clamp-like conformational changes of two loops of the receptor that anchor the CRF C terminus, including the C-terminal amide group. These structural studies provide a molecular framework for understanding peptide binding and specificity by the CRF receptors as well as a template for designing potent and selective CRFR1 antagonists for therapeutic applications.

  6. Reptile Toll-like receptor 5 unveils adaptive evolution of bacterial flagellin recognition

    PubMed Central

    Voogdt, Carlos G. P.; Bouwman, Lieneke I.; Kik, Marja J. L.; Wagenaar, Jaap A.; van Putten, Jos P. M.

    2016-01-01

    Toll-like receptors (TLR) are ancient innate immune receptors crucial for immune homeostasis and protection against infection. TLRs are present in mammals, birds, amphibians and fish but have not been functionally characterized in reptiles despite the central position of this animal class in vertebrate evolution. Here we report the cloning, characterization, and function of TLR5 of the reptile Anolis carolinensis (Green Anole lizard). The receptor (acTLR5) displays the typical TLR protein architecture with 22 extracellular leucine rich repeats flanked by a N- and C-terminal leucine rich repeat domain, a membrane-spanning region, and an intracellular TIR domain. The receptor is phylogenetically most similar to TLR5 of birds and most distant to fish TLR5. Transcript analysis revealed acTLR5 expression in multiple lizard tissues. Stimulation of acTLR5 with TLR ligands demonstrated unique responsiveness towards bacterial flagellin in both reptile and human cells. Comparison of acTLR5 and human TLR5 using purified flagellins revealed differential sensitivity to Pseudomonas but not Salmonella flagellin, indicating development of species-specific flagellin recognition during the divergent evolution of mammals and reptiles. Our discovery of reptile TLR5 fills the evolutionary gap regarding TLR conservation across vertebrates and provides novel insights in functional evolution of host-microbe interactions. PMID:26738735

  7. Reptile Toll-like receptor 5 unveils adaptive evolution of bacterial flagellin recognition.

    PubMed

    Voogdt, Carlos G P; Bouwman, Lieneke I; Kik, Marja J L; Wagenaar, Jaap A; van Putten, Jos P M

    2016-01-07

    Toll-like receptors (TLR) are ancient innate immune receptors crucial for immune homeostasis and protection against infection. TLRs are present in mammals, birds, amphibians and fish but have not been functionally characterized in reptiles despite the central position of this animal class in vertebrate evolution. Here we report the cloning, characterization, and function of TLR5 of the reptile Anolis carolinensis (Green Anole lizard). The receptor (acTLR5) displays the typical TLR protein architecture with 22 extracellular leucine rich repeats flanked by a N- and C-terminal leucine rich repeat domain, a membrane-spanning region, and an intracellular TIR domain. The receptor is phylogenetically most similar to TLR5 of birds and most distant to fish TLR5. Transcript analysis revealed acTLR5 expression in multiple lizard tissues. Stimulation of acTLR5 with TLR ligands demonstrated unique responsiveness towards bacterial flagellin in both reptile and human cells. Comparison of acTLR5 and human TLR5 using purified flagellins revealed differential sensitivity to Pseudomonas but not Salmonella flagellin, indicating development of species-specific flagellin recognition during the divergent evolution of mammals and reptiles. Our discovery of reptile TLR5 fills the evolutionary gap regarding TLR conservation across vertebrates and provides novel insights in functional evolution of host-microbe interactions.

  8. Allosteric modulation of peroxisomal membrane protein recognition by farnesylation of the peroxisomal import receptor PEX19

    PubMed Central

    Emmanouilidis, Leonidas; Schütz, Ulrike; Tripsianes, Konstantinos; Madl, Tobias; Radke, Juliane; Rucktäschel, Robert; Wilmanns, Matthias; Schliebs, Wolfgang; Erdmann, Ralf; Sattler, Michael

    2017-01-01

    The transport of peroxisomal membrane proteins (PMPs) requires the soluble PEX19 protein as chaperone and import receptor. Recognition of cargo PMPs by the C-terminal domain (CTD) of PEX19 is required for peroxisome biogenesis in vivo. Farnesylation at a C-terminal CaaX motif in PEX19 enhances the PMP interaction, but the underlying molecular mechanisms are unknown. Here, we report the NMR-derived structure of the farnesylated human PEX19 CTD, which reveals that the farnesyl moiety is buried in an internal hydrophobic cavity. This induces substantial conformational changes that allosterically reshape the PEX19 surface to form two hydrophobic pockets for the recognition of conserved aromatic/aliphatic side chains in PMPs. Mutations of PEX19 residues that either mediate farnesyl contacts or are directly involved in PMP recognition abolish cargo binding and cannot complement a ΔPEX19 phenotype in human Zellweger patient fibroblasts. Our results demonstrate an allosteric mechanism for the modulation of protein function by farnesylation. PMID:28281558

  9. Molecular Recognition by a Polymorphic Cell Surface Receptor Governs Cooperative Behaviors in Bacteria

    PubMed Central

    Dey, Arup; Wall, Daniel

    2013-01-01

    Cell-cell recognition is a fundamental process that allows cells to coordinate multicellular behaviors. Some microbes, such as myxobacteria, build multicellular fruiting bodies from free-living cells. However, how bacterial cells recognize each other by contact is poorly understood. Here we show that myxobacteria engage in recognition through interactions between TraA cell surface receptors, which leads to the fusion and exchange of outer membrane (OM) components. OM exchange is shown to be selective among 17 environmental isolates, as exchange partners parsed into five major recognition groups. TraA is the determinant of molecular specificity because: (i) exchange partners correlated with sequence conservation within its polymorphic PA14-like domain and (ii) traA allele replacements predictably changed partner specificity. Swapping traA alleles also reprogrammed social interactions among strains, including the regulation of motility and conferred immunity from inter-strain killing. We suggest that TraA helps guide the transition of single cells into a coherent bacterial community, by a proposed mechanism that is analogous to mitochondrial fusion and fission cycling that mixes contents to establish a homogenous population. In evolutionary terms, traA functions as a rare greenbeard gene that recognizes others that bear the same allele to confer beneficial treatment. PMID:24244178

  10. Structural basis of collagen recognition by human osteoclast-associated receptor and design of osteoclastogenesis inhibitors.

    PubMed

    Haywood, Joel; Qi, Jianxun; Chen, Chun-Chi; Lu, Guangwen; Liu, Yingxia; Yan, Jinghua; Shi, Yi; Gao, George F

    2016-01-26

    Human osteoclast-associated receptor (OSCAR) is an immunoglobulin (Ig)-like collagen receptor that is up-regulated on osteoclasts during osteoclastogenesis and is expressed in a range of myeloid cells. As a member of the leukocyte receptor complex family of proteins, OSCAR shares a high degree of sequence and structural homology with other collagen receptors of this family, including glycoprotein VI, leukocyte-associated Ig-like receptor-1, and leukocyte Ig-like receptor B4, but recognizes a unique collagen sequence. Here, we present the crystal structures of OSCAR in its free form and in complex with a triple-helical collagen-like peptide (CLP). These structures reveal that the CLP peptide binds only one of the two Ig-like domains, the membrane-proximal domain (domain 2) of OSCAR, with the middle and trailing chain burying a total of 661 Å(2) of solvent-accessible collagen surface. This binding mode is facilitated by the unusual topography of the OSCAR protein, which displays an obtuse interdomain angle and a rotation of domain 2 relative to the membrane-distal domain 1. Moreover, the binding of the CLP to OSCAR appears to be mediated largely by tyrosine residues and conformational changes at a shallow Phe pocket. Furthermore, we investigated CLP peptides as inhibitors of osteoclastogenesis and found that a peptide length of 40 amino acids is required to ensure adequate inhibition of osteoclastogenesis in vitro. These findings provide valuable structural insights into the mode of collagen recognition by OSCAR and into the use of synthetic peptide matrikines for osteoclastogenesis inhibition.

  11. Toward a structure-based model of salvinorin A recognition of the kappa-opioid receptor.

    PubMed

    Kane, Brian E; McCurdy, Christopher R; Ferguson, David M

    2008-03-27

    The structural basis to salvinorin A recognition of the kappa-opioid receptor is evaluated using a combination of site-directed mutagenesis and molecular-modeling techniques. The results show that salvinorin A recognizes a collection of residues in transmembrane II and VII, including Q115, Y119, Y313, I316, and Y320. The mutation of one hydrophobic residue in particular, I316, was found to completely abolish salvinorin A binding. As expected, none of the residues in transmembrane III or VI commonly associated with opiate recognition (such as D138 or E297) appear to be required for ligand binding. On the basis of the results presented here and elsewhere, a binding site model is proposed that aligns salvinorin A vertically within a pocket spanning transmembrane II and VII, with the 2' substituent directed toward the extracellular domains. The model explains the role that hydrophobic contacts play in binding this lipophilic ligand and gives insight into the structural basis to the mu-opioid receptor selectivity of 2'-benzoyl salvinorin (herkinorin).

  12. Bioconjugation of Small Molecules to RNA Impedes Its Recognition by Toll-Like Receptor 7

    PubMed Central

    Hellmuth, Isabell; Freund, Isabel; Schlöder, Janine; Seidu-Larry, Salifu; Thüring, Kathrin; Slama, Kaouthar; Langhanki, Jens; Kaloyanova, Stefka; Eigenbrod, Tatjana; Krumb, Matthias; Röhm, Sandra; Peneva, Kalina; Opatz, Till; Jonuleit, Helmut; Dalpke, Alexander H.; Helm, Mark

    2017-01-01

    A fundamental mechanism of the innate immune system is the recognition, via extra- and intracellular pattern-recognition receptors, of pathogen-associated molecular patterns. A prominent example is represented by foreign nucleic acids, triggering the activation of several signaling pathways. Among these, the endosomal toll-like receptor 7 (TLR7) is known to be activated by single-stranded RNA (ssRNA), which can be specifically influenced through elements of sequence structure and posttranscriptional modifications. Furthermore, small molecules TLR7 agonists (smTLRa) are applied as boosting adjuvants in vaccination processes. In this context, covalent conjugations between adjuvant and vaccines have been reported to exhibit synergistic effects. Here, we describe a concept to chemically combine three therapeutic functions in one RNA bioconjugate. This consists in the simultaneous TLR7 stimulation by ssRNA and smTLRa as well as the therapeutic function of the RNA itself, e.g., as a vaccinating or knockdown agent. We have hence synthesized bioconjugates of mRNA and siRNA containing covalently attached smTLRa and tested their function in TLR7 stimulation. Strikingly, the bioconjugates displayed decreased rather than synergistically increased stimulation. The decrease was distinct from the antagonistic action of an siRNA bearing a Gm motive, as observed by direct comparison of the effects in the presence of otherwise stimulatory RNA. In summary, these investigations showed that TRL7 activation can be impeded by bioconjugation of small molecules to RNA. PMID:28392787

  13. Estradiol enhances object recognition memory in Swiss female mice by activating hippocampal estrogen receptor α.

    PubMed

    Pereira, Luciana M; Bastos, Cristiane P; de Souza, Jéssica M; Ribeiro, Fabíola M; Pereira, Grace S

    2014-10-01

    In rodents, 17β-estradiol (E2) enhances hippocampal function and improves performance in several memory tasks. Regarding the object recognition paradigm, E2 commonly act as a cognitive enhancer. However, the types of estrogen receptor (ER) involved, as well as the underlying molecular mechanisms are still under investigation. In the present study, we asked whether E2 enhances object recognition memory by activating ERα and/or ERβ in the hippocampus of Swiss female mice. First, we showed that immediately post-training intraperitoneal (i.p.) injection of E2 (0.2 mg/kg) allowed object recognition memory to persist 48 h in ovariectomized (OVX) Swiss female mice. This result indicates that Swiss female mice are sensitive to the promnesic effects of E2 and is in accordance with other studies, which used C57/BL6 female mice. To verify if the activation of hippocampal ERα or ERβ would be sufficient to improve object memory, we used PPT and DPN, which are selective ERα and ERβ agonists, respectively. We found that PPT, but not DPN, improved object memory in Swiss female mice. However, DPN was able to improve memory in C57/BL6 female mice, which is in accordance with other studies. Next, we tested if the E2 effect on improving object memory depends on ER activation in the hippocampus. Thus, we tested if the infusion of intra-hippocampal TPBM and PHTPP, selective antagonists of ERα and ERβ, respectively, would block the memory enhancement effect of E2. Our results showed that TPBM, but not PHTPP, blunted the promnesic effect of E2, strongly suggesting that in Swiss female mice, the ERα and not the ERβ is the receptor involved in the promnesic effect of E2. It was already demonstrated that E2, as well as PPT and DPN, increase the phospho-ERK2 level in the dorsal hippocampus of C57/BL6 mice. Here we observed that PPT increased phospho-ERK1, while DPN decreased phospho-ERK2 in the dorsal hippocampus of Swiss female mice subjected to the object recognition sample phase

  14. Different roles for M1 and M2 receptors within perirhinal cortex in object recognition and discrimination.

    PubMed

    Bartko, Susan J; Winters, Boyer D; Saksida, Lisa M; Bussey, Timothy J

    2014-04-01

    Recognition and discrimination of objects and individuals are critical cognitive faculties in both humans and non-human animals, and cholinergic transmission has been shown to be essential for both of these functions. In the present study we focused on the role of M1 and M2 muscarinic receptors in perirhinal cortex (PRh)-dependent object recognition and discrimination. The selective M1 antagonists pirenzepine and the snake toxin MT-7, and a selective M2 antagonist, AF-DX 116, were infused directly into PRh. Pre-sample infusions of both pirenzepine and AF-DX 116 significantly impaired object recognition memory in a delay-dependent manner. However, pirenzepine and MT-7, but not AF-DX 116, impaired oddity discrimination performance in a perceptual difficulty-dependent manner. The findings indicate distinct functions for M1 and M2 receptors in object recognition and discrimination.

  15. Molecular mechanism for differential recognition of membrane phosphatidylserine by the immune regulatory receptor Tim4.

    PubMed

    Tietjen, Gregory T; Gong, Zhiliang; Chen, Chiu-Hao; Vargas, Ernesto; Crooks, James E; Cao, Kathleen D; Heffern, Charles T R; Henderson, J Michael; Meron, Mati; Lin, Binhua; Roux, Benot; Schlossman, Mark L; Steck, Theodore L; Lee, Ka Yee C; Adams, Erin J

    2014-04-15

    Recognition of phosphatidylserine (PS) lipids exposed on the extracellular leaflet of plasma membranes is implicated in both apoptotic cell removal and immune regulation. The PS receptor T cell immunoglobulin and mucin-domain-containing molecule 4 (Tim4) regulates T-cell immunity via phagocytosis of both apoptotic (high PS exposure) and nonapoptotic (intermediate PS exposure) activated T cells. The latter population must be removed at lower efficiency to sensitively control immune tolerance and memory cell population size, but the molecular basis for how Tim4 achieves this sensitivity is unknown. Using a combination of interfacial X-ray scattering, molecular dynamics simulations, and membrane binding assays, we demonstrate how Tim4 recognizes PS in the context of a lipid bilayer. Our data reveal that in addition to the known Ca(2+)-coordinated, single-PS binding pocket, Tim4 has four weaker sites of potential ionic interactions with PS lipids. This organization makes Tim4 sensitive to PS surface concentration in a manner capable of supporting differential recognition on the basis of PS exposure level. The structurally homologous, but functionally distinct, Tim1 and Tim3 are significantly less sensitive to PS surface density, likely reflecting the differences in immunological function between the Tim proteins. These results establish the potential for lipid membrane parameters, such as PS surface density, to play a critical role in facilitating selective recognition of PS-exposing cells. Furthermore, our multidisciplinary approach overcomes the difficulties associated with characterizing dynamic protein/membrane systems to reveal the molecular mechanisms underlying Tim4's recognition properties, and thereby provides an approach capable of providing atomic-level detail to uncover the nuances of protein/membrane interactions.

  16. Staphylococcal protein Ecb impairs complement receptor-1 mediated recognition of opsonized bacteria

    PubMed Central

    Amdahl, Hanne; Tan, Lydia; Meri, Taru; Kuusela, Pentti I.; van Strijp, Jos A.

    2017-01-01

    Staphyloccus aureus is a major human pathogen leading frequently to sepsis and soft tissue infections with abscesses. Multiple virulence factors including several immune modulating molecules contribute to its survival in the host. When S. aureus invades the human body, one of the first line defenses is the complement system, which opsonizes the bacteria with C3b and attract neutrophils by release of chemotactic peptides. Neutrophils express Complement receptor-1 [CR1, CD35) that interacts with the C3b-opsonized particles and thereby plays an important role in pathogen recognition by phagocytic cells. In this study we observed that a fraction of S. aureus culture supernatant prevented binding of C3b to neutrophils. This fraction consisted of S. aureus leukocidins and Efb. The C-terminus of Efb is known to bind C3b and shares significant sequence homology to the extracellular complement binding protein [Ecb). Here we show that S. aureus Ecb displays various mechanisms to block bacterial recognition by neutrophils. The presence of Ecb blocked direct interaction between soluble CR1 and C3b and reduced the cofactor activity of CR1 in proteolytic inactivation of C3b. Furthermore, Ecb could dose-dependently prevent recognition of C3b by cell-bound CR1 that lead to impaired phagocytosis of NHS-opsonized S. aureus. Phagocytosis was furthermore reduced in the presence of soluble CR1 [sCR1). These data indicate that the staphylococcal protein Ecb prevents recognition of C3b opsonized bacteria by neutrophil CR1 leading to impaired killing by phagocytosis and thereby contribute to immune evasion of S. aureus. PMID:28273167

  17. Selection and identification of ligand peptides targeting a model of castrate-resistant osteogenic prostate cancer and their receptors.

    PubMed

    Mandelin, Jami; Cardó-Vila, Marina; Driessen, Wouter H P; Mathew, Paul; Navone, Nora M; Lin, Sue-Hwa; Logothetis, Christopher J; Rietz, Anna Cecilia; Dobroff, Andrey S; Proneth, Bettina; Sidman, Richard L; Pasqualini, Renata; Arap, Wadih

    2015-03-24

    We performed combinatorial peptide library screening in vivo on a novel human prostate cancer xenograft that is androgen-independent and induces a robust osteoblastic reaction in bonelike matrix and soft tissue. We found two peptides, PKRGFQD and SNTRVAP, which were enriched in the tumors, targeted the cell surface of androgen-independent prostate cancer cells in vitro, and homed to androgen receptor-null prostate cancer in vivo. Purification of tumor homogenates by affinity chromatography on these peptides and subsequent mass spectrometry revealed a receptor for the peptide PKRGFQD, α-2-macroglobulin, and for SNTRVAP, 78-kDa glucose-regulated protein (GRP78). These results indicate that GRP78 and α-2-macroglobulin are highly active in osteoblastic, androgen-independent prostate cancer in vivo. These previously unidentified ligand-receptor systems should be considered for targeted drug development against human metastatic androgen-independent prostate cancer.

  18. Closely Related Antibody Receptors Exploit Fundamentally Different Strategies for Steroid Recognition

    SciTech Connect

    Verdino, P.; Aldag, C.; Hilvert, D.; Wilson, I.A.

    2009-05-26

    Molecular recognition by the adaptive immune system relies on specific high-affinity antibody receptors that are generated from a restricted set of starting sequences through homologous recombination and somatic mutation. The steroid binding antibody DB3 and the catalytic Diels-Alderase antibody 1E9 derive from the same germ line sequences but exhibit very distinct specificities and functions. However, mutation of only two of the 36 sequence differences in the variable domains, Leu{sup H47}Trp and Arg{sup H100}Trp, converts 1E9 into a high-affinity steroid receptor with a ligand recognition profile similar to DB3. To understand how these changes switch binding specificity and function, we determined the crystal structures of the 1E9 Leu{sup H47}Trp/Arg{sup H100}Trp double mutant (1E9dm) as an unliganded Fab at 2.05 {angstrom} resolution and in complex with two configurationally distinct steroids at 2.40 and 2.85 {angstrom}. Surprisingly, despite the functional mimicry of DB3, 1E9dm employs a distinct steroid binding mechanism. Extensive structural rearrangements occur in the combining site, where residue H47 acts as a specificity switch and H100 adapts to different ligands. Unlike DB3, 1E9dm does not use alternative binding pockets or different sets of hydrogen-bonding interactions to bind configurationally distinct steroids. Rather, the different steroids are inserted more deeply into the 1E9dm combining site, creating more hydrophobic contacts that energetically compensate for the lack of hydrogen bonds. These findings demonstrate how subtle mutations within an existing molecular scaffold can dramatically modulate the function of immune receptors by inducing unanticipated, but compensating, mechanisms of ligand interaction.

  19. Macrophage recognition of toxic advanced glycosylation end products through the macrophage surface-receptor nucleolin.

    PubMed

    Miki, Yuichi; Dambara, Hikaru; Tachibana, Yoshihiro; Hirano, Kazuya; Konishi, Mio; Beppu, Masatoshi

    2014-01-01

    Advanced glycosylation end-products (AGEs) are non-enzymatically glycosylated proteins that play an important role in several diseases and aging processes, including angiopathy, renal failure, diabetic complications, and some neurodegenerative diseases. In particular, glyceraldehyde (GCA)- and glycolaldehyde (GOA)-derived AGEs are deemed toxic AGEs, due to their cytotoxicity. Recently, the shuttling-protein nucleolin has been shown to possess scavenger receptor-activity. Here, we investigated whether or not macrophages recognize toxic AGEs through nucleolin receptors expressed on their surface. Free amino acid groups and arginine residues found in bovine serum albumin (BSA) were time-dependently modified by incubation with GCA and GOA. In addition, average molecular size was increased by incubation with GCA and GOA. While GCA-treated BSA (GCA-BSA) and GOA-treated BSA (GOA-BSA) were recognized by thioglycollate-elicited mouse peritoneal macrophages in proportion to their respective aldehyde-modification ratios, aldehyde-untreated control-BSA was not. Surface plasmon-resonance analysis revealed that nucleolin strongly associated with GCA-BSA and GOA-BSA, but not with control-BSA. Further, pretreating macrophages with anti-nucleolin antibody, but not control-Immunoglobulin G, inhibited recognition of GCA-BSA and GOA-BSA by macrophages. Additionally, AGRO, a nucleolin-specific oligonucleotide aptamer, inhibited recognition of GCA-BSA and GOA-BSA. Moreover, nucleolin-transfected HEK293 cells recognized more GCA-BSA and GOA-BSA than control HEK cells did. Binding of nucleolin and GCA-BSA/GOA-BSA was also blocked by anti-nucleolin antibody at molecular level. These results indicate that nucleolin is a receptor that allows macrophages to recognize toxic AGEs.

  20. Differential effects of m1 and m2 receptor antagonists in perirhinal cortex on visual recognition memory in monkeys.

    PubMed

    Wu, Wei; Saunders, Richard C; Mishkin, Mortimer; Turchi, Janita

    2012-07-01

    Microinfusions of the nonselective muscarinic antagonist scopolamine into perirhinal cortex impairs performance on visual recognition tasks, indicating that muscarinic receptors in this region play a pivotal role in recognition memory. To assess the mnemonic effects of selective blockade in perirhinal cortex of muscarinic receptor subtypes, we locally infused either the m1-selective antagonist pirenzepine or the m2-selective antagonist methoctramine in animals performing one-trial visual recognition, and compared these scores with those following infusions of equivalent volumes of saline. Compared to these control infusions, injections of pirenzepine, but not of methoctramine, significantly impaired recognition accuracy. Further, similar doses of scopolamine and pirenzepine yielded similar deficits, suggesting that the deficits obtained earlier with scopolamine were due mainly, if not exclusively, to blockade of m1 receptors. The present findings indicate that m1 and m2 receptors have functionally dissociable roles, and that the formation of new visual memories is critically dependent on the cholinergic activation of m1 receptors located on perirhinal cells.

  1. Aldehyde Recognition and Discrimination by Mammalian Odorant Receptors via Functional Group-Specific Hydration Chemistry

    PubMed Central

    2015-01-01

    The mammalian odorant receptors (ORs) form a chemical-detecting interface between the atmosphere and the nervous system. This large gene family is composed of hundreds of membrane proteins predicted to form as many unique small molecule binding niches within their G-protein coupled receptor (GPCR) framework, but very little is known about the molecular recognition strategies they use to bind and discriminate between small molecule odorants. Using rationally designed synthetic analogs of a typical aliphatic aldehyde, we report evidence that among the ORs showing specificity for the aldehyde functional group, a significant percentage detect the aldehyde through its ability to react with water to form a 1,1-geminal (gem)-diol. Evidence is presented indicating that the rat OR-I7, an often-studied and modeled OR known to require the aldehyde function of octanal for activation, is likely one of the gem-diol activated receptors. A homology model based on an activated GPCR X-ray structure provides a structural hypothesis for activation of OR-I7 by the gem-diol of octanal. PMID:25181321

  2. Kappa Opioid Receptor-Mediated Disruption of Novel Object Recognition: Relevance for Psychostimulant Treatment

    PubMed Central

    Paris, Jason J.; Reilley, Kate J.; McLaughlin, Jay P.

    2012-01-01

    Kappa opioid receptor (KOR) agonists are potentially valuable as therapeutics for the treatment of psychostimulant reward as they suppress dopamine signaling in reward circuitry to repress drug seeking behavior. However, KOR agonists are also associated with sedation and cognitive dysfunction. The extent to which learning and memory disruption or hypolocomotion underlie KOR agonists’ role in counteracting the rewarding effects of psychostimulants is of interest. C57BL/6J mice were pretreated with vehicle (saline, 0.9%), the KOR agonist (trans)-3,4-dichloro-N-methyl-N-[2-(1- pyrrolidinyl)-cyclohexyl] benzeneacetamide (U50,488), or the peripherally-restricted agonist D-Phe-D-Phe-D-lle-D-Arg- NH2 (ffir-NH2), through central (i.c.v.) or peripheral (i.p.) routes of administration. Locomotor activity was assessed via activity monitoring chambers and rotorod. Cognitive performance was assessed in a novel object recognition task. Prolonged hypolocomotion was observed following administration of 1.0 and 10.0, but not 0.3 mg/kg U50,488. Central, but not peripheral, administration of ffir-NH2 (a KOR agonist that does not cross the blood-brain barrier) also reduced motor behavior. Systemic pretreatment with the low dose of U50,488 (0.3 mg/kg, i.p.) significantly impaired performance in the novel object recognition task. Likewise, ffir-NH2 significantly reduced novel object recognition after central (i.c.v.), but not peripheral (i.p.), administration. U50,488- and ffir-NH2-mediated deficits in novel object recognition were prevented by pretreatment with KOR antagonists. Cocaine-induced conditioned place preference was subsequently assessed and was reduced by pretreatment with U50,488 (0.3 mg/kg, i.p.). Together, these results suggest that the activation of centrally-located kappa opioid receptors may induce cognitive and mnemonic disruption independent of hypolocomotor effects which may contribute to the KOR-mediated suppression of psychostimulant reward. PMID:22900234

  3. Recognition of lipopeptide patterns by Toll-like receptor 2-Toll-like receptor 6 heterodimer.

    PubMed

    Kang, Jin Young; Nan, Xuehua; Jin, Mi Sun; Youn, Suk-Jun; Ryu, Young Hee; Mah, Shinjee; Han, Seung Hyun; Lee, Hayyoung; Paik, Sang-Gi; Lee, Jie-Oh

    2009-12-18

    Toll-like receptor 2 (TLR2) initiates potent immune responses by recognizing diacylated and triacylated lipopeptides. Its ligand specificity is controlled by whether it heterodimerizes with TLR1 or TLR6. We have determined the crystal structures of TLR2-TLR6-diacylated lipopeptide, TLR2-lipoteichoic acid, and TLR2-PE-DTPA complexes. PE-DTPA, 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine-N-diethylenetriaminepentaacetic acid, is a synthetic phospholipid derivative. Two major factors contribute to the ligand specificity of TLR2-TLR1 or TLR2-TLR6 heterodimers. First, the lipid channel of TLR6 is blocked by two phenylalanines. Simultaneous mutation of these phenylalanines made TLR2-TLR6 fully responsive not only to diacylated but also to triacylated lipopeptides. Second, the hydrophobic dimerization interface of TLR2-TLR6 is increased by 80%, which compensates for the lack of amide lipid interaction between the lipopeptide and TLR2-TLR6. The structures of the TLR2-lipoteichoic acid and the TLR2-PE-DTPA complexes demonstrate that a precise interaction pattern of the head group is essential for a robust immune response by TLR2 heterodimers.

  4. T cell receptor recognition of CD1b presenting a mycobacterial glycolipid

    PubMed Central

    Gras, Stephanie; Van Rhijn, Ildiko; Shahine, Adam; Cheng, Tan-Yun; Bhati, Mugdha; Tan, Li Lynn; Halim, Hanim; Tuttle, Kathryn D.; Gapin, Laurent; Le Nours, Jérôme; Moody, D. Branch; Rossjohn, Jamie

    2016-01-01

    CD1 proteins present microbial lipids to T cells. Germline-encoded mycolyl lipid-reactive (GEM) T cells with conserved αβ T cell receptors (TCRs) recognize CD1b presenting mycobacterial mycolates. As the molecular basis underpinning TCR recognition of CD1b remains unknown, here we determine the structure of a GEM TCR bound to CD1b presenting glucose-6-O-monomycolate (GMM). The GEM TCR docks centrally above CD1b, whereby the conserved TCR α-chain extensively contacts CD1b and GMM. Through mutagenesis and study of T cells from tuberculosis patients, we identify a consensus CD1b footprint of TCRs present among GEM T cells. Using both the TCR α- and β-chains as tweezers to surround and grip the glucose moiety of GMM, GEM TCRs create a highly specific mechanism for recognizing this mycobacterial glycolipid. PMID:27807341

  5. Structural Basis for Hormone Recognition by the Human CRFR2[alpha] G Protein-coupled Receptor

    SciTech Connect

    Pal, Kuntal; Swaminathan, Kunchithapadam; Xu, H. Eric; Pioszak, Augen A.

    2012-05-09

    The mammalian corticotropin releasing factor (CRF)/urocortin (Ucn) peptide hormones include four structurally similar peptides, CRF, Ucn1, Ucn2, and Ucn3, that regulate stress responses, metabolism, and cardiovascular function by activating either of two related class B G protein-coupled receptors, CRFR1 and CRFR2. CRF and Ucn1 activate both receptors, whereas Ucn2 and Ucn3 are CRFR2-selective. The molecular basis for selectivity is unclear. Here, we show that the purified N-terminal extracellular domains (ECDs) of human CRFR1 and the CRFR2{alpha} isoform are sufficient to discriminate the peptides, and we present three crystal structures of the CRFR2{alpha} ECD bound to each of the Ucn peptides. The CRFR2{alpha} ECD forms the same fold observed for the CRFR1 and mouse CRFR2{beta} ECDs but contains a unique N-terminal {alpha}-helix formed by its pseudo signal peptide. The CRFR2{alpha} ECD peptide-binding site architecture is similar to that of CRFR1, and binding of the {alpha}-helical Ucn peptides closely resembles CRF binding to CRFR1. Comparing the electrostatic surface potentials of the ECDs suggests a charge compatibility mechanism for ligand discrimination involving a single amino acid difference in the receptors (CRFR1 Glu104/CRFR2{alpha} Pro-100) at a site proximate to peptide residue 35 (Arg in CRF/Ucn1, Ala in Ucn2/3). CRFR1 Glu-104 acts as a selectivity filter preventing Ucn2/3 binding because the nonpolar Ala-35 is incompatible with the negatively charged Glu-104. The structures explain the mechanisms of ligand recognition and discrimination and provide a molecular template for the rational design of therapeutic agents selectively targeting these receptors.

  6. Neural Androgen Receptors Modulate Gene Expression and Social Recognition But Not Social Investigation

    PubMed Central

    Karlsson, Sara A.; Studer, Erik; Kettunen, Petronella; Westberg, Lars

    2016-01-01

    The role of sex and androgen receptors (ARs) for social preference and social memory is rather unknown. In this study of mice we compared males, females and males lacking ARs specifically in the nervous system, ARNesDel, with respect to social preference, assessed with the three-chambered apparatus test, and social recognition, assessed with the social discrimination procedure. In the social discrimination test we also evaluated the tentative importance of the sex of the stimulus animal. Novel object recognition and olfaction were investigated to complement the results from the social tests. Gene expression analysis was performed to reveal molecules involved in the effects of sex and androgens on social behaviors. All three test groups showed social preference in the three-chambered apparatus test. In both social tests an AR-independent sexual dimorphism was seen in the persistence of social investigation of female conspecifics, whereas the social interest toward male stimuli mice was similar in all groups. Male and female controls recognized conspecifics independent of their sex, whereas ARNesDel males recognized female but not male stimuli mice. Moreover, the non-social behaviors were not affected by AR deficiency. The gene expression analyses of hypothalamus and amygdala indicated that Oxtr, Cd38, Esr1, Cyp19a1, Ucn3, Crh, and Gtf2i were differentially expressed between the three groups. In conclusion, our results suggest that ARs are required for recognition of male but not female conspecifics, while being dispensable for social investigation toward both sexes. In addition, the AR seems to regulate genes related to oxytocin, estrogen and William’s syndrome. PMID:27014003

  7. Atypical natural killer T-cell receptor recognition of CD1d–lipid antigens

    PubMed Central

    Le Nours, Jérôme; Praveena, T.; Pellicci, Daniel G.; Gherardin, Nicholas A.; Ross, Fiona J.; Lim, Ricky T.; Besra, Gurdyal S.; Keshipeddy, Santosh; Richardson, Stewart K.; Howell, Amy R.; Gras, Stephanie; Godfrey, Dale I.; Rossjohn, Jamie; Uldrich, Adam P.

    2016-01-01

    Crucial to Natural Killer T (NKT) cell function is the interaction between their T-cell receptor (TCR) and CD1d-antigen complex. However, the diversity of the NKT cell repertoire and the ensuing interactions with CD1d-antigen remain unclear. We describe an atypical population of CD1d–α-galactosylceramide (α-GalCer)-reactive human NKT cells that differ markedly from the prototypical TRAV10-TRAJ18-TRBV25-1+ type I NKT cell repertoire. These cells express a range of TCR α- and β-chains that show differential recognition of glycolipid antigens. Two atypical NKT TCRs (TRAV21-TRAJ8-TRBV7–8 and TRAV12-3-TRAJ27-TRBV6-5) bind orthogonally over the A′-pocket of CD1d, adopting distinct docking modes that contrast with the docking mode of all type I NKT TCR-CD1d-antigen complexes. Moreover, the interactions with α-GalCer differ between the type I and these atypical NKT TCRs. Accordingly, diverse NKT TCR repertoire usage manifests in varied docking strategies and specificities towards CD1d–α-GalCer and related antigens, thus providing far greater scope for diverse glycolipid antigen recognition. PMID:26875526

  8. Inducible Prophage Mutant of Escherichia coli Can Lyse New Host and the Key Sites of Receptor Recognition Identification

    PubMed Central

    Chen, Mianmian; Zhang, Lei; Xin, Sipei; Yao, Huochun; Lu, Chengping; Zhang, Wei

    2017-01-01

    The use of bacteriophages as therapeutic agents is hindered by their narrow and specific host range, and by a lack of the knowledge concerning the molecular mechanism of receptor recognition. Two P2-like coliphages, named P88 and pro147, were induced from Escherichia coli strains K88 and DE147, respectively. A comparison of the genomes of these two and other P2-like coliphages obtained from GenBank showed that the tail fiber protein genes, which are the key genes for receptor recognition in other myoviridae phages, showed more diversity than the conserved lysin, replicase, and terminase genes. Firstly, replacing hypervariable region 2 (HR2: amino acids 716–746) of the tail fiber protein of P88 with that of pro147 changed the host range of P88. Then, replacing six amino acids in HR2 with the corresponding residues from pro147 altered the host range only in these mutants with changes at position 730 (leucine) and 744 (glutamic acid). Thus, we predicted that these amino acids are vital to establish the host range of P88. This study provided a vector of lysogenic bacteria that could be used to change or expand the phage host range of P88. These results illustrated that, in P2-like phage P88, the tail fiber protein determined the receptor recognition. Amino acids 716–746 and the amino acids at positions 730 and 744 were important for receptor recognition. PMID:28203234

  9. Molecular recognition of human ephrinB2 cell surface receptor by an emergent African henipavirus.

    PubMed

    Lee, Benhur; Pernet, Olivier; Ahmed, Asim A; Zeltina, Antra; Beaty, Shannon M; Bowden, Thomas A

    2015-04-28

    The discovery of African henipaviruses (HNVs) related to pathogenic Hendra virus (HeV) and Nipah virus (NiV) from Southeast Asia and Australia presents an open-ended health risk. Cell receptor use by emerging African HNVs at the stage of host-cell entry is a key parameter when considering the potential for spillover and infection of human populations. The attachment glycoprotein from a Ghanaian bat isolate (GhV-G) exhibits <30% sequence identity with Asiatic NiV-G/HeV-G. Here, through functional and structural analysis of GhV-G, we show how this African HNV targets the same human cell-surface receptor (ephrinB2) as the Asiatic HNVs. We first characterized this virus-receptor interaction crystallographically. Compared with extant HNV-G-ephrinB2 structures, there was significant structural variation in the six-bladed β-propeller scaffold of the GhV-G receptor-binding domain, but not the Greek key fold of the bound ephrinB2. Analysis revealed a surprisingly conserved mode of ephrinB2 interaction that reflects an ongoing evolutionary constraint among geographically distal and phylogenetically divergent HNVs to maintain the functionality of ephrinB2 recognition during virus-host entry. Interestingly, unlike NiV-G/HeV-G, we could not detect binding of GhV-G to ephrinB3. Comparative structure-function analysis further revealed several distinguishing features of HNV-G function: a secondary ephrinB2 interaction site that contributes to more efficient ephrinB2-mediated entry in NiV-G relative to GhV-G and cognate residues at the very C terminus of GhV-G (absent in Asiatic HNV-Gs) that are vital for efficient receptor-induced fusion, but not receptor binding per se. These data provide molecular-level details for evaluating the likelihood of African HNVs to spill over into human populations.

  10. Molecular recognition of human ephrinB2 cell surface receptor by an emergent African henipavirus

    PubMed Central

    Lee, Benhur; Pernet, Olivier; Ahmed, Asim A.; Zeltina, Antra; Beaty, Shannon M.; Bowden, Thomas A.

    2015-01-01

    The discovery of African henipaviruses (HNVs) related to pathogenic Hendra virus (HeV) and Nipah virus (NiV) from Southeast Asia and Australia presents an open-ended health risk. Cell receptor use by emerging African HNVs at the stage of host-cell entry is a key parameter when considering the potential for spillover and infection of human populations. The attachment glycoprotein from a Ghanaian bat isolate (GhV-G) exhibits <30% sequence identity with Asiatic NiV-G/HeV-G. Here, through functional and structural analysis of GhV-G, we show how this African HNV targets the same human cell-surface receptor (ephrinB2) as the Asiatic HNVs. We first characterized this virus−receptor interaction crystallographically. Compared with extant HNV-G–ephrinB2 structures, there was significant structural variation in the six-bladed β-propeller scaffold of the GhV-G receptor-binding domain, but not the Greek key fold of the bound ephrinB2. Analysis revealed a surprisingly conserved mode of ephrinB2 interaction that reflects an ongoing evolutionary constraint among geographically distal and phylogenetically divergent HNVs to maintain the functionality of ephrinB2 recognition during virus–host entry. Interestingly, unlike NiV-G/HeV-G, we could not detect binding of GhV-G to ephrinB3. Comparative structure–function analysis further revealed several distinguishing features of HNV-G function: a secondary ephrinB2 interaction site that contributes to more efficient ephrinB2-mediated entry in NiV-G relative to GhV-G and cognate residues at the very C terminus of GhV-G (absent in Asiatic HNV-Gs) that are vital for efficient receptor-induced fusion, but not receptor binding per se. These data provide molecular-level details for evaluating the likelihood of African HNVs to spill over into human populations. PMID:25825759

  11. Crystal structure of pentapeptide-independent chemotaxis receptor methyltransferase (CheR) reveals idiosyncratic structural determinants for receptor recognition.

    PubMed

    Batra, Monu; Sharma, Rajesh; Malik, Anjali; Dhindwal, Sonali; Kumar, Pravindra; Tomar, Shailly

    2016-12-01

    Chemotactic methyltransferase, CheR catalyse methylation of specific glutamate residues in the cytoplasmic domain of methyl-accepting chemotactic protein receptors (MCPRs). The methylation of MCPRs is essential for the chemical sensing and chemotactic bacterial mobility towards favorable chemicals or away from unfavorable ones. In this study, crystal structure of B. subtilis CheR (BsCheR) in complex with S-adenosyl-l-homocysteine (SAH) has been determined to 1.8Å resolution. This is the first report of crystal structure belonging to the pentapeptide-independent CheR (PICheR) class. Till date, only one crystal structure of CheR from S. typhimurium (StCheR) belonging to pentapeptide-dependent CheR (PDCheR) class is available. Structural analysis of BsCheR reveals a helix-X-helix motif (HXH) with Asp53 as the linker residue in the N-terminal domain. The key structural features of the PDCheR β-subdomain involved in the formation of a tight complex with the pentapeptide binding motif in MCPRs were found to be absent in the structure of BsCheR. Additionally, isothermal titration calorimetry (ITC) experiments were performed to investigate S-adenosyl-(l)-methionine (SAM) binding affinity and KD was determined to be 0.32mM. The structure of BsCheR reveals that mostly residues of the large C-terminal domain contribute to SAH binding, with contributions of few residues from the linker region and the N-terminal domain. Structural investigations and sequence analysis carried out in this study provide critical insights into the distinct receptor recognition mechanism of the PDCheR and PICheR methyltransferase classes.

  12. Capturing intercellular sugar-mediated ligand-receptor recognitions via a simple yet highly biospecific interfacial system.

    PubMed

    Li, Zhen; Deng, Si-Si; Zang, Yi; Gu, Zhen; He, Xiao-Peng; Chen, Guo-Rong; Chen, Kaixian; James, Tony D; Li, Jia; Long, Yi-Tao

    2013-01-01

    Intercellular ligand-receptor recognitions are crucial natural interactions that initiate a number of biological and pathological events. We present here the simple construction of a unique class of biomimetic interfaces based on a graphene-mediated self-assembly of glycosyl anthraquinones to a screen-printed electrode for the detection of transmembrane glycoprotein receptors expressed on a hepatoma cell line. We show that an electroactive interface confined with densely clustered galactosyl ligands is able to ingeniously recognize the asialoglycoprotein receptors on live Hep-G2 cells employing simple electrochemical techniques. The only facility used is a personal laptop in connection with a cheap and portable electrochemical workstation.

  13. Enantioselective Recognition for Many Different Kinds of Chiral Guests by One Chiral Receptor Based on Tetraphenylethylene Cyclohexylbisurea.

    PubMed

    Xiong, Jia-Bin; Xie, Wen-Zhao; Sun, Jian-Ping; Wang, Jin-Hua; Zhu, Zhi-Hua; Feng, Hai-Tao; Guo, Dong; Zhang, Hui; Zheng, Yan-Song

    2016-05-06

    A neutral chiral receptor based on TPE cyclohexylbisurea was synthesized and could discriminate the enantiomers of many different kinds of chiral reagents, including chiral acidic compounds, basic compounds, amino acids, and even neutral alcohols. The (1)H NMR spectra disclosed that the ability of chiral recognition could be ascribed to the multiple hydrogen bonds and CH-π interactions between the TPE urea receptor and the enantiomer of the chiral guest, which led to the selective aggregation of the receptor with one of the two enantiomers. This result exhibited a great potential in enantiomer discernment and high-throughput analysis of enantiomer composition of these chiral analytes by one chiral AIE molecule.

  14. Capturing intercellular sugar-mediated ligand-receptor recognitions via a simple yet highly biospecific interfacial system

    NASA Astrophysics Data System (ADS)

    Li, Zhen; Deng, Si-Si; Zang, Yi; Gu, Zhen; He, Xiao-Peng; Chen, Guo-Rong; Chen, Kaixian; James, Tony D.; Li, Jia; Long, Yi-Tao

    2013-07-01

    Intercellular ligand-receptor recognitions are crucial natural interactions that initiate a number of biological and pathological events. We present here the simple construction of a unique class of biomimetic interfaces based on a graphene-mediated self-assembly of glycosyl anthraquinones to a screen-printed electrode for the detection of transmembrane glycoprotein receptors expressed on a hepatoma cell line. We show that an electroactive interface confined with densely clustered galactosyl ligands is able to ingeniously recognize the asialoglycoprotein receptors on live Hep-G2 cells employing simple electrochemical techniques. The only facility used is a personal laptop in connection with a cheap and portable electrochemical workstation.

  15. Anion recognition by simple chromogenic and chromo-fluorogenic salicylidene Schiff base or reduced-Schiff base receptors

    NASA Astrophysics Data System (ADS)

    Dalapati, Sasanka; Jana, Sankar; Guchhait, Nikhil

    2014-08-01

    This review contains extensive application of anion sensing ability of salicylidene type Schiff bases and their reduced forms having various substituents with respect to phenolic sbnd OH group. Some of these molecular systems behave as receptor for recognition or sensing of various anions in organic or aqueous-organic binary solvent mixture as well as in the solid supported test kits. Development of Schiff base or reduced Schiff base receptors for anion recognition event is commonly based on the theory of hydrogen bonding interaction or deprotonation of phenolic -OH group. The process of charge transfer (CT) or inhibition of excited proton transfer (ESIPT) or followed by photo-induced electron transfer (PET) lead to naked-eye color change, UV-vis spectral change, chemical shift in the NMR spectra and fluorescence spectral modifications. In this review we have tried to discuss about the anion sensing properties of Schiff base or reduced Schiff base receptors.

  16. Human gestation-associated tissues express functional cytosolic nucleic acid sensing pattern recognition receptors.

    PubMed

    Bryant, Aled H; Menzies, Georgina E; Scott, Louis M; Spencer-Harty, Samantha; Davies, Lleucu B; Smith, Rachel A; Jones, Ruth H; Thornton, Catherine A

    2017-03-13

    The role of viral infections in adverse pregnancy outcomes has gained interest in recent years. Innate immune pattern recognition receptors (PRRs) and their signalling pathways that yield a cytokine output in response to pathogenic stimuli have been postulated to link infection at the maternal-fetal interface and adverse pregnancy outcomes. The objective of this study was to investigate the expression and functional response of nucleic acid ligand responsive Toll-like receptors (TLR3, 7, 8 and 9), and retinoic acid-inducible gene 1 (RIG-I)-like receptors (RIG-I, MDA5 and LGP2) in human term gestation-associated tissues (placenta, choriodecidua and amnion) using an explant model. Immunohistochemistry revealed that these PRRs were expressed by the term placenta, choriodecidua and amnion. A statistically significant increase in interleukin (IL)-6 and/or IL-8 production in response to specific agonists for TLR3 (Poly(I:C); low and high molecular weight), TLR7 (Imiquimod), TLR8 (ssRNA40) and RIG-I/MDA5 (Poly(I:C)LyoVec) was observed; there was no response to a TLR9 (ODN21798) agonist. A hierarchical clustering approach was used to compare the response of each tissue type to the ligands studied and revealed that the placenta and choriodecidua generate a more similar IL-8 response, while the choriodecidua and amnion generate a more similar IL-6 response to nucleic acid ligands. These findings demonstrate that responsiveness via TLR3, TLR7, TLR8 and RIG-1/MDA5 is a broad feature of human term gestation-associated tissues with differential responses by tissue that might underpin adverse obstetric outcomes. This article is protected by copyright. All rights reserved.

  17. Toll-like receptor recognition of bacteria in fish: ligand specificity and signal pathways.

    PubMed

    Zhang, Jie; Kong, Xianghui; Zhou, Chuanjiang; Li, Li; Nie, Guoxing; Li, Xuejun

    2014-12-01

    Pattern recognition receptors (PRRs) recognize the conserved molecular structure of pathogens and trigger the signaling pathways that activate immune cells in response to pathogen infection. Toll-like receptors (TLRs) are the first and best characterized innate immune receptors. To date, at least 20 TLR types (TLR1, 2, 3, 4, 5M, 5S, 7, 8, 9, 13, 14, 18, 19, 20, 21, 22, 23, 24, 25, and 26) have been found in more than a dozen of fish species. However, of the TLRs identified in fish, direct evidence of ligand specificity has only been shown for TLR2, TLR3, TLR5M, TLR5S, TLR9, TLR21, and TLR22. Some studies have suggested that TLR2, TLR5M, TLR5S, TLR9, and TLR21 could specifically recognize PAMPs from bacteria. In addition, other TLRs including TLR1, TLR4, TLR14, TLR18, and TLR25 may also be sensors of bacteria. TLR signaling pathways in fish exhibit some particular features different from that in mammals. In this review, the ligand specificity and signal pathways of TLRs that recognize bacteria in fish are summarized. References for further studies on the specificity for recognizing bacteria using TLRs and the following reactions triggered are discussed. In-depth studies should be continuously performed to identify the ligand specificity of all TLRs in fish, particularly non-mammalian TLRs, and their signaling pathways. The discovery of TLRs and their functions will contribute to the understanding of disease resistance mechanisms in fish and provide new insights for drug intervention to manipulate immune responses.

  18. Two distinct receptors account for recognition of maleyl-albumin in human monocytes during differentiation in vitro.

    PubMed Central

    Haberland, M E; Rasmussen, R R; Olch, C L; Fogelman, A M

    1986-01-01

    A comparison of the receptor-mediated interaction of malondialdehyde-low density lipoprotein and maleyl-albumin has been examined in human monocytes during differentiation in vitro. The recognition of both ligands by the scavenger receptor of these cells has been confirmed. We now report that human monocytes express a second cellular surface receptor for maleyl-albumin that is distinct from the scavenger receptor. The activity of the maleyl-albumin receptor, determined by both binding and lysosomal hydrolytic assays, substantially exceeds that of the scavenger receptor in freshly isolated monocytes. A dramatic and rapid decline in the activity of the maleyl-albumin receptor occurs within 72 to 96 h during differentiation in vitro. At day 7, while only 5-10% of the original activity of the maleyl-albumin receptor remains, it is similar to that of the maximally expressed scavenger receptor. Both the binding and hydrolysis of ligand mediated by the maleyl-albumin receptor are specifically inhibited by alpha-casein and alkaline-treated albumin; neither of these proteins is recognized by the scavenger receptor. The occurrence of the exceptionally active maleyl-albumin receptor on freshly isolated human monocytes suggests that it participates in processes necessary to the function of the cells that diminish in importance after differentiation of the monocytes into macrophages in vitro. Furthermore, while maleyl-albumin is a useful adjunct to studies of cellular events mediated by the scavenger receptor, the presence of a second receptor for maleyl-albumin must be taken into account as a potential contributing and complicating event. Images PMID:3949974

  19. Gene polymorphisms in pattern recognition receptors and susceptibility to idiopathic recurrent vulvovaginal candidiasis

    PubMed Central

    Rosentul, Diana C.; Delsing, Corine E.; Jaeger, Martin; Plantinga, Theo S.; Oosting, Marije; Costantini, Irene; Venselaar, Hanka; Joosten, Leo A. B.; van der Meer, Jos W. M.; Dupont, Bertrand; Kullberg, Bart-Jan; Sobel, Jack D.; Netea, Mihai G.

    2014-01-01

    Objective: Approximately 5% of women suffer from recurrent vulvovaginal candidiasis (RVVC). It has been hypothesized that genetic factors play an important role in the susceptibility to RVVC. The aim of this study was to assess the effect of genetic variants of genes encoding for pattern recognition receptors (PRRs) on susceptibility to RVVC. Study design: For the study, 119 RVVC patients and 263 healthy controls were recruited. Prevalence of polymorphisms in five PRRs involved in recognition of Candida were investigated in patients and controls. In silico and functional studies were performed to assess their functional effects. Results: Single nucleotide polymorphisms (SNPs) in TLR1, TLR4, CLEC7A, and CARD9 did not affect the susceptibility to RVVC. In contrast, a non-synonymous polymorphism in TLR2 (rs5743704, Pro631His) increased the susceptibility to RVVC almost 3-fold. Furthermore, the TLR2 rs5743704 SNP had deleterious effects on protein function as assessed by in silico analysis, and in vitro functional assays suggested that it reduces production of IL-17 and IFNγ upon stimulation of peripheral blood mononuclear cells with Candida albicans. No effects were observed on serum mannose-binding lectin concentrations. Condensation: This study demonstrates the association of susceptibility to RVVC with genetic variation in TLR2, most likely caused by decreased induction of mucosal antifungal host defense. Conclusion: Genetic variation in TLR2 may significantly enhance susceptibility to RVVC by modulating host defense mechanisms against Candida. Additional studies are warranted to assess systematically the role of host genetic variation for susceptibility to RVVC. PMID:25295030

  20. FERM-dependent E3 ligase recognition is a conserved mechanism for targeted degradation of lipoprotein receptors

    PubMed Central

    Calkin, Anna C.; Goult, Benjamin T.; Zhang, Li; Fairall, Louise; Hong, Cynthia; Schwabe, John W. R.; Tontonoz, Peter

    2011-01-01

    The E3 ubiquitin ligase IDOL (inducible degrader of the LDL receptor) regulates LDL receptor (LDLR)-dependent cholesterol uptake, but its mechanism of action, including the molecular basis for its stringent specificity, is poorly understood. Here we show that IDOL uses a singular strategy among E3 ligases for target recognition. The IDOL FERM domain binds directly to a recognition sequence in the cytoplasmic tails of lipoprotein receptors. This physical interaction is independent of IDOL's really interesting new gene (RING) domain E3 ligase activity and its capacity for autoubiquitination. Furthermore, IDOL controls its own stability through autoubiquitination of a unique FERM subdomain fold not present in other FERM proteins. Key residues defining the IDOL–LDLR interaction and IDOL autoubiquitination are functionally conserved in their insect homologs. Finally, we demonstrate that target recognition by IDOL involves a tripartite interaction between the FERM domain, membrane phospholipids, and the lipoprotein receptor tail. Our data identify the IDOL–LDLR interaction as an evolutionarily conserved mechanism for the regulation of lipid uptake and suggest that this interaction could potentially be exploited for the pharmacologic modulation of lipid metabolism. PMID:22109552

  1. FERM-dependent E3 ligase recognition is a conserved mechanism for targeted degradation of lipoprotein receptors.

    PubMed

    Calkin, Anna C; Goult, Benjamin T; Zhang, Li; Fairall, Louise; Hong, Cynthia; Schwabe, John W R; Tontonoz, Peter

    2011-12-13

    The E3 ubiquitin ligase IDOL (inducible degrader of the LDL receptor) regulates LDL receptor (LDLR)-dependent cholesterol uptake, but its mechanism of action, including the molecular basis for its stringent specificity, is poorly understood. Here we show that IDOL uses a singular strategy among E3 ligases for target recognition. The IDOL FERM domain binds directly to a recognition sequence in the cytoplasmic tails of lipoprotein receptors. This physical interaction is independent of IDOL's really interesting new gene (RING) domain E3 ligase activity and its capacity for autoubiquitination. Furthermore, IDOL controls its own stability through autoubiquitination of a unique FERM subdomain fold not present in other FERM proteins. Key residues defining the IDOL-LDLR interaction and IDOL autoubiquitination are functionally conserved in their insect homologs. Finally, we demonstrate that target recognition by IDOL involves a tripartite interaction between the FERM domain, membrane phospholipids, and the lipoprotein receptor tail. Our data identify the IDOL-LDLR interaction as an evolutionarily conserved mechanism for the regulation of lipid uptake and suggest that this interaction could potentially be exploited for the pharmacologic modulation of lipid metabolism.

  2. Coiled coil miniprotein randomization on phage leads to charge pattern mimicry of the receptor recognition determinant of interleukin 5.

    PubMed

    Li, Chuanzhao; Plugariu, Carmela G; Bajgier, Joanna; White, John R; Liefer, Kristin M; Wu, Sheng-Jiun; Chaiken, Irwin

    2002-01-01

    Phage display was used to identify sequences that mimic structural determinants in interleukin5 (IL5) for IL5 receptor recognition. A coiled coil stem loop (CCSL) miniprotein scaffold library was constructed with its turn region randomized and panned for binding variants against human IL5 receptor alpha chain (IL5Ralpha). Competition enzyme-linked immunosorbent assays identified CCSL-phage selectants for which binding to IL5Ralpha was competed by IL5. The most frequently selected and IL5-competed CCSL-phage contain charged residues Arg and Glu in their turn sequences, in this regard resembling a beta strand sequence in the 'CD turn' region, of IL5, that has been proposed to present a key determinant for IL5 receptor alpha chain recognition. The most dominant CCSL-phage selectant sequence, PVEGRV, contains a negative/positive charge pattern similar to that seen in the original CD turn. To test the relatedness of CCSL-phage selectant sequences to the IL5 receptor recognition epitope, PVEGRV was grafted into the sequence 87--92 of a monomeric IL5. The resulting IL5 variant, [(87)PVEGRV(92)]GM1, was able to bind to IL5Ralpha in biosensor assays, to elicit TF-1 cell proliferation and to induce STAT5 phosphorylation in TF-1 cells. The results help discern sequence patterns in the IL5 CD turn region which are key in driving receptor recognition and demonstrate the utility of CCSL miniprotein scaffold phage display to identify local IL5 mimetic sequence arrangements that may ultimately lead to IL5 antagonists.

  3. Antibody recognition of the pandemic H1N1 Influenza virus hemagglutinin receptor binding site.

    PubMed

    Hong, Minsun; Lee, Peter S; Hoffman, Ryan M B; Zhu, Xueyong; Krause, Jens C; Laursen, Nick S; Yoon, Sung-Il; Song, Langzhou; Tussey, Lynda; Crowe, James E; Ward, Andrew B; Wilson, Ian A

    2013-11-01

    Influenza virus is a global health concern due to its unpredictable pandemic potential. This potential threat was realized in 2009 when an H1N1 virus emerged that resembled the 1918 virus in antigenicity but fortunately was not nearly as deadly. 5J8 is a human antibody that potently neutralizes a broad spectrum of H1N1 viruses, including the 1918 and 2009 pandemic viruses. Here, we present the crystal structure of 5J8 Fab in complex with a bacterially expressed and refolded globular head domain from the hemagglutinin (HA) of the A/California/07/2009 (H1N1) pandemic virus. 5J8 recognizes a conserved epitope in and around the receptor binding site (RBS), and its HCDR3 closely mimics interactions of the sialic acid receptor. Electron microscopy (EM) reconstructions of 5J8 Fab in complex with an HA trimer from a 1986 H1 strain and with an engineered stabilized HA trimer from the 2009 H1 pandemic virus showed a similar mode of binding. As for other characterized RBS-targeted antibodies, 5J8 uses avidity to extend its breadth and affinity against divergent H1 strains. 5J8 selectively interacts with HA insertion residue 133a, which is conserved in pandemic H1 strains and has precluded binding of other RBS-targeted antibodies. Thus, the RBS of divergent HAs is targeted by 5J8 and adds to the growing arsenal of common recognition motifs for design of therapeutics and vaccines. Moreover, consistent with previous studies, the bacterially expressed H1 HA properly refolds, retaining its antigenic structure, and presents a low-cost and rapid alternative for engineering and manufacturing candidate flu vaccines.

  4. CRF₂ receptor-deficiency reduces recognition memory deficits and vulnerability to stress induced by cocaine withdrawal.

    PubMed

    Morisot, Nadège; Le Moine, Catherine; Millan, Mark J; Contarino, Angelo

    2014-12-01

    Psychostimulant drug abuse, dependence and withdrawal are associated with cognitive dysfunction and impact stress-sensitive systems. The corticotropin-releasing factor (CRF) system orchestrates stress responses via CRF1 and CRF2 receptors and is implicated in substance use disorders. However, CRF2 role in psychostimulant drug-induced cognitive dysfunction remains to be elucidated. In the present study, wild-type and CRF2-/- mice are injected with cocaine and memory assessed by the novel object recognition (NOR) task throughout relatively long periods of drug withdrawal. Following recovery from the drug-induced memory deficits, the mice are stressed prior to the NOR task and brain gene expression evaluated by in situ hybridization. Cocaine impairs NOR memory in wild-type and CRF2-/- mice. However, following cocaine withdrawal NOR memory deficits last less time in CRF2-/- than in wild-type mice. Furthermore, a relatively mild stressor induces the re-emergence of NOR deficits in long-term cocaine-withdrawn wild-type but not CRF2-/- mice. Cocaine-withdrawn mice show a genotype-independent higher c-fos expression in the NOR memory-relevant perirhinal cortex than drug-naïve mice. However neither genotype nor drug withdrawal affect the expression of tyrosine hydroxylase in the ventral tegmental area or the locus coeruleus and CRF in the central nucleus of the amygdala or the paraventricular nucleus of the hypothalamus, brain regions implicated in stress and drug responses. These data indicate a new role for the CRF2 receptor in cognitive deficits induced by cocaine withdrawal, both as regards to their duration and their re-induction by stress. Interestingly, prototypical brain stress systems other than CRF do not appear to be involved.

  5. Two Independent Histidines One in Human Prolactin and One in Its Receptor Are Critical for pH-dependent Receptor Recognition and Activation

    SciTech Connect

    M Kulkarni; M Tettamanzi; J Murphy; C Keeler; D Myszka; N Chayen; E Lolis; M Hodsdon

    2011-12-31

    Human prolactin (hPRL), a member of the family of hematopoietic cytokines, functions as both an endocrine hormone and autocrine/paracrine growth factor. We have previously demonstrated that recognition of the hPRL-receptor depends strongly on solution acidity over the physiologic range from pH 6 to pH 8. The hPRL-receptor binding interface contains four histidines whose protonation is hypothesized to regulate pH-dependent receptor recognition. Here, we systematically dissect its molecular origin by characterizing the consequences of His to Ala mutations on pH-dependent receptor binding kinetics, site-specific histidine protonation, and high resolution structures of the intermolecular interface. Thermodynamic modeling of the pH dependence to receptor binding affinity reveals large changes in site-specific protonation constants for a majority of interface histidines upon complexation. Removal of individual His imidazoles reduces these perturbations in protonation constants, which is most likely explained by the introduction of solvent-filled, buried cavities in the crystallographic structures without inducing significant conformational rearrangements.

  6. Differential Responses of Pattern Recognition Receptors to Outer Membrane Vesicles of Three Periodontal Pathogens.

    PubMed

    Cecil, Jessica D; O'Brien-Simpson, Neil M; Lenzo, Jason C; Holden, James A; Chen, Yu-Yen; Singleton, William; Gause, Katelyn T; Yan, Yan; Caruso, Frank; Reynolds, Eric C

    2016-01-01

    Highly purified outer membrane vesicles (OMVs) of the periodontal pathogens, Porphyromonas gingivalis, Treponema denticola and Tannerella forsythia were produced using tangential flow ultrafiltration, ultracentrifugation and Optiprep density gradient separation. Cryo-TEM and light scattering showed OMVs to be single lipid-bilayers with modal diameters of 75 to 158 nm. Enumeration of OMVs by nanoparticle flow-cytometry at the same stage of late exponential culture indicated that P. gingivalis was the most prolific OMV producer. P. gingivalis OMVs induced strong TLR2 and TLR4-specific responses and moderate responses in TLR7, TLR8, TLR9, NOD1 and NOD2 expressing-HEK-Blue cells. Responses to T. forsythia OMVs were less than those of P. gingivalis and T. denticola OMVs induced only weak responses. Compositional analyses of OMVs from the three pathogens demonstrated differences in protein, fatty acids, lipopolysaccharide, peptidoglycan fragments and nucleic acids. Periodontal pathogen OMVs induced differential pattern recognition receptor responses that have implications for their role in chronic periodontitis.

  7. A Novel Loop Domain in Superantigens Extends Their T Cell Receptor Recognition Site

    SciTech Connect

    Gunther,S.; Varma, A.; Moza, B.; Kasper, K.; Wyatt, A.; Zhu, P.; Nur-ur Rahman, A.; Li, Y.; Mariuzza, R.; et al.

    2007-01-01

    Superantigens (SAGs) interact with host immune receptors to induce a massive release of inflammatory cytokines that can lead to toxic shock syndrome and death. Bacterial SAGs can be classified into five distinct evolutionary groups. Group V SAGs are characterized by the {alpha}3-{beta}8 loop, a unique {approx}15 amino acid residue extension that is required for optimal T cell activation. Here, we report the X-ray crystal structures of the group V SAG staphylococcal enterotoxin K (SEK) alone and in complex with the TCR hV{beta}5.1 domain. SEK adopts a unique TCR binding orientation relative to other SAG-TCR complexes, which results in the {alpha}3-{beta}8 loop contacting the apical loop of framework region 4, thereby extending the known TCR recognition site of SAGs. These interactions are absolutely required for TCR binding and T cell activation by SEK, and dictate the TCR V{beta} domain specificity of SEK and other group V SAGs.

  8. Differential Responses of Pattern Recognition Receptors to Outer Membrane Vesicles of Three Periodontal Pathogens

    PubMed Central

    Lenzo, Jason C.; Holden, James A.; Chen, Yu-Yen; Singleton, William; Gause, Katelyn T.; Yan, Yan; Caruso, Frank; Reynolds, Eric C.

    2016-01-01

    Highly purified outer membrane vesicles (OMVs) of the periodontal pathogens, Porphyromonas gingivalis, Treponema denticola and Tannerella forsythia were produced using tangential flow ultrafiltration, ultracentrifugation and Optiprep density gradient separation. Cryo-TEM and light scattering showed OMVs to be single lipid-bilayers with modal diameters of 75 to 158 nm. Enumeration of OMVs by nanoparticle flow-cytometry at the same stage of late exponential culture indicated that P. gingivalis was the most prolific OMV producer. P. gingivalis OMVs induced strong TLR2 and TLR4-specific responses and moderate responses in TLR7, TLR8, TLR9, NOD1 and NOD2 expressing-HEK-Blue cells. Responses to T. forsythia OMVs were less than those of P. gingivalis and T. denticola OMVs induced only weak responses. Compositional analyses of OMVs from the three pathogens demonstrated differences in protein, fatty acids, lipopolysaccharide, peptidoglycan fragments and nucleic acids. Periodontal pathogen OMVs induced differential pattern recognition receptor responses that have implications for their role in chronic periodontitis. PMID:27035339

  9. Recognition of the Thomsen-Friedenreich Pancarcinoma Carbohydrate Antigen by a Lamprey Variable Lymphocyte Receptor*

    PubMed Central

    Luo, Ming; Velikovsky, C. Alejandro; Yang, Xinbo; Siddiqui, Maqbool A.; Hong, Xia; Barchi, Joseph J.; Gildersleeve, Jeffrey C.; Pancer, Zeev; Mariuzza, Roy A.

    2013-01-01

    Variable lymphocyte receptors (VLRs) are leucine-rich repeat proteins that mediate adaptive immunity in jawless vertebrates. VLRs were recently shown to recognize glycans, such as the tumor-associated Thomsen-Friedenreich antigen (TFα; Galβ1–3GalNAcα), with a selectivity rivaling or exceeding that of lectins and antibodies. To understand the basis for TFα recognition by one such VLR (VLRB.aGPA.23), we measured thermodynamic parameters for the binding interaction and determined the structure of the VLRB.aGPA.23-TFα complex to 2.2 Å resolution. In the structure, four tryptophan residues form a tight hydrophobic cage encasing the TFα disaccharide that completely excludes buried water molecules. This cage together with hydrogen bonding of sugar hydroxyls to polar side chains explains the exquisite selectivity of VLRB.aGPA.23. The topology of the glycan-binding site of VLRB.aGPA.23 differs markedly from those of lectins or antibodies, which typically consist of long, convex grooves for accommodating the oligosaccharide. Instead, the TFα disaccharide is sandwiched between a variable loop and the concave surface of the VLR formed by the β-strands of the leucine-rich repeat modules. Longer oligosaccharides are predicted to extend perpendicularly across the β-strands, requiring them to bend to match the concavity of the VLR solenoid. PMID:23782692

  10. Enter at your own risk: how enteroviruses navigate the dangerous world of pattern recognition receptor signaling.

    PubMed

    Harris, Katharine G; Coyne, Carolyn B

    2013-09-01

    Enteroviruses are the most common human viral pathogens worldwide. This genus of small, non-enveloped, single stranded RNA viruses includes coxsackievirus, rhinovirus, echovirus, and poliovirus species. Infection with these viruses can induce mild symptoms that resemble the common cold, but can also be associated with more severe syndromes such as poliomyelitis, neurological diseases including aseptic meningitis and encephalitis, myocarditis, and the onset of type I diabetes. In humans, polarized epithelial cells lining the respiratory and/or digestive tracts represent the initial sites of infection by enteroviruses. Control of infection in the host is initiated through the engagement of a variety of pattern recognition receptors (PRRs). PRRs act as the sentinels of the innate immune system and serve to alert the host to the presence of a viral invader. This review assembles the available data annotating the role of PRRs in the response to enteroviral infection as well as the myriad ways by which enteroviruses both interrupt and manipulate PRR signaling to enhance their own replication, thereby inducing human disease.

  11. Structural Basis of Natural Promoter Recognition by a Unique Nuclear Receptor, HNF4[alpha

    SciTech Connect

    Lu, Peng; Rha, Geun Bae; Melikishvili, Manana; Wu, Guangteng; Adkins, Brandon C.; Fried, Michael G.; Chi, Young-In

    2010-11-09

    HNF4{alpha} (hepatocyte nuclear factor 4{alpha}) plays an essential role in the development and function of vertebrate organs, including hepatocytes and pancreatic {beta}-cells by regulating expression of multiple genes involved in organ development, nutrient transport, and diverse metabolic pathways. As such, HNF4{alpha} is a culprit gene product for a monogenic and dominantly inherited form of diabetes, known as maturity onset diabetes of the young (MODY). As a unique member of the nuclear receptor superfamily, HNF4{alpha} recognizes target genes containing two hexanucleotide direct repeat DNA-response elements separated by one base pair (DR1) by exclusively forming a cooperative homodimer. We describe here the 2.0 {angstrom} crystal structure of human HNF4{alpha} DNA binding domain in complex with a high affinity promoter element of another MODY gene, HNF1{alpha}, which reveals the molecular basis of unique target gene selection/recognition, DNA binding cooperativity, and dysfunction caused by diabetes-causing mutations. The predicted effects of MODY mutations have been tested by a set of biochemical and functional studies, which show that, in contrast to other MODY gene products, the subtle disruption of HNF4{alpha} molecular function can cause significant effects in afflicted MODY patients.

  12. Exploring new molecular architectures for anion recognition: synthesis and ATP binding properties of new cyclam-based ditopic polyammonium receptors.

    PubMed

    Pouessel, Jacky; Bazzicalupi, Carla; Bencini, Andrea; Bernard, Hélène; Giorgi, Claudia; Handel, Henri; Matera, Irene; Le Bris, Nathalie; Tripier, Raphaël; Valtancoli, Barbara

    2011-06-06

    Synthesis and characterization of three new polyamine receptors, composed of a cyclam unit (cyclam=1,4,8,11-tetraazacyclotetradecane) linked by a 2,6-dimethylpyridinyl spacer to the linear polyamines 1,4,8,11-tetraazaundecane (L1py), 1,4,7-triazaheptane (L2py), and to a quaternary ammonium group (L3py(+)), are reported. All receptors form highly charged polyammonium cations at neutral pH, suitable for anion recognition studies. ATP recognition was analyzed by using potentiometric, calorimetric, (1)H and (31)P NMR measurements in aqueous solution. All receptors form 1:1 adducts with ATP in aqueous solution, stabilized by charge-charge and hydrogen-bonding interactions between their ammonium groups and the anionic triphosphate chain of ATP. The binding ability of the three receptors for ATP increases in the order of L3py(+)receptors to wrap around the phosphate chain of ATP.

  13. Highly effective recognition of carbohydrates by phenanthroline-based receptors: alpha- versus beta-anomer binding preference.

    PubMed

    Mazik, Monika; Hartmann, Andrè; Jones, Peter G

    2009-09-14

    (1)H NMR spectroscopic titrations in competitive and non-competitive media, as well as binding studies in two-phase systems, such as phase transfer of sugars from aqueous into organic solvents and dissolution of solid carbohydrates in apolar media revealed both highly effective recognition of neutral carbohydrates and interesting binding preferences of an acyclic phenanthroline-based receptor 1. Compared to the previously described acyclic receptors, compound 1 displays significantly higher binding affinities, the rare capability to extract sugars from water into non-polar organic solutions and alpha- versus beta-anomer binding preference in the recognition of glycosides, which differs from those observed for other receptor systems. X-ray crystallographic investigations revealed the presence of water molecules in the binding pocket of 1 that are engaged in the formation of hydrogen-bonding motifs similar to those suggested by molecular modelling for the sugar OH groups in the receptor-sugar complexes. The molecular modelling calculations, synthesis, crystal structure and binding properties of 1 are described and compared with those of the previously described receptors.

  14. NLRP7 and related inflammasome activating pattern recognition receptors and their function in host defense and disease.

    PubMed

    Radian, Alexander D; de Almeida, Lucia; Dorfleutner, Andrea; Stehlik, Christian

    2013-01-01

    Host defense requires the maturation and release of the pro-inflammatory cytokines interleukin (IL)-1β and IL-18 and the induction of pyroptotic cell death, which depends on the activation of inflammatory Caspases within inflammasomes by innate immune cells. Several cytosolic pattern recognition receptors (PRRs) have been implicated in this process in response to infectious and sterile agonists. Here we summarize the current knowledge on inflammasome-organizing PRRs, emphasizing the recently described NLRP7, and their implications in human disease.

  15. The broad pattern recognition spectrum of the Toll-like receptor in mollusk Zhikong scallop Chlamys farreri.

    PubMed

    Wang, Mengqiang; Wang, Lingling; Guo, Ying; Sun, Rui; Yue, Feng; Yi, Qilin; Song, Linsheng

    2015-10-01

    Toll-like receptors (TLRs) are among the most studied pattern recognition receptors (PRRs) playing essential roles in innate immune defenses. In the present study, the basic features of CfTLR in mollusk Zhikong scallop Chlamys farreri, including sequence homology, tissue distribution, subcellular localization and ligands spectrum, were investigated to elucidate its pattern recognition. The elements of extracellular domains (ECD) in CfTLR displayed high homology to the corresponding parts of the ECDs in TLRs from Homo sapiens. CfTLR protein was detected in hemocytes, mantle, gills, hepatopancreas, kidney and gonad of the scallops, and it was localized in both the plasma membranes and the lysosomes in HEK293T cells. CfTLR could activate NFκB in response to multiple HsTLR ligands including Pam3CSK4, glucan (GLU), peptidoglycan (PGN), polyriboinosinic:polyribocytidylic acid (poly I:C), Imiquimod and three types of CpG. Additionally, the scallop serum could enhance the induction of NFκB in the CfTLR expressing cells elicited by most PAMPs, including GLU, PGN, Imiquimod and four types of CpG. It could be concluded that this primitive mollusk TLR shared a hybrid function in pattern recognition and could recognize broader ligands than mammalian TLRs, and its mosaic capability of pathogen associated molecular pattern (PAMP) recognition might be based on the basic features of its structure, ligand properties and the assistance of some components in scallop serum.

  16. Dopamine D1 receptor activity modulates object recognition memory consolidation in the perirhinal cortex but not in the hippocampus.

    PubMed

    Balderas, Israela; Moreno-Castilla, Perla; Bermudez-Rattoni, Federico

    2013-10-01

    It has been proposed that distributed neuronal networks in the medial temporal lobe process different characteristics of a recognition event; the hippocampus has been associated with contextual recollection while the perirhinal cortex has been linked with familiarity. Here we show that D1 dopamine receptor activity in these two structures participates differentially in object recognition memory consolidation. The D1 receptor antagonist SCH23390 was infused bilaterally 15 min before a 5 min sample phase in either rats' perirhinal cortex or dorsal hippocampus, and they were tested 90 min for short-term memory or 24 h later for long-term memory. SCH23390 impaired long-term memory when infused in the perirhinal cortex but not when infused in the hippocampus. Conversely, when the D1 receptor agonist SKF38393 was infused 10 min before a 3 min sample phase in the perirhinal cortex, long-term memory was enhanced, however, this was not observed when the D1 agonist was infused in the hippocampus. Short-term memory was spared when SCH23390 or SKF38393 were infused in the perirhinal cortex or the dorsal hippocampus suggesting that acquisition was unaffected. These results suggest that dopaminergic transmission in these medial temporal lobe structures have a differential involvement in object recognition memory consolidation.

  17. Design and Function of Supramolecular Recognition Systems Based on Guest-Targeting Probe-Modified Cyclodextrin Receptors for ATP.

    PubMed

    Fujita, Kyohhei; Fujiwara, Shoji; Yamada, Tatsuru; Tsuchido, Yuji; Hashimoto, Takeshi; Hayashita, Takashi

    2017-01-20

    In this study, we have developed a rational design strategy to obtain highly selective supramolecular recognition systems of cyclodextrins (CyDs) on the basis of the lock and key principle. We designed and synthesized dipicolylamine (dpa)-modified γ-CyD-Cu(2+) complexes possessing an azobenzene unit (Cu·1-γ-CyD) and examined how they recognized phosphoric acid derivatives in water. The results revealed that Cu·1-γ-CyD recognized ATP with high selectivity over other phosphoric acid derivatives. The significant blue shift in the UV-vis spectra and (1)H NMR analysis suggested that the selective ATP recognition was based on the multipoint interactions between the adenine moiety of ATP and both the CyD cavity and the azobenzene unit in addition to the recognition of phosphoric moieties by the Cu-dpa complex site. Our unique receptor made it capable of distinguishing ATP from AMP and ADP, revealing the discrimination of even a length of one phosphoric group. This study demonstrates that, compared to conventional recognition systems of CyDs, this multipoint recognition system confers a higher degree of selectivity for certain organic molecules, such as ATP, over their similar derivatives.

  18. Structural basis of RNA recognition and activation by innate immune receptor RIG-I

    SciTech Connect

    Jiang, Fuguo; Ramanathan, Anand; Miller, Matthew T.; Tang, Guo-Qing; Gale, Jr., Michael; Patel, Smita S.; Marcotrigiano, Joseph

    2012-05-29

    Retinoic-acid-inducible gene-I (RIG-I; also known as DDX58) is a cytoplasmic pathogen recognition receptor that recognizes pathogen-associated molecular pattern (PAMP) motifs to differentiate between viral and cellular RNAs. RIG-I is activated by blunt-ended double-stranded (ds)RNA with or without a 5'-triphosphate (ppp), by single-stranded RNA marked by a 5'-ppp and by polyuridine sequences. Upon binding to such PAMP motifs, RIG-I initiates a signalling cascade that induces innate immune defences and inflammatory cytokines to establish an antiviral state. The RIG-I pathway is highly regulated and aberrant signalling leads to apoptosis, altered cell differentiation, inflammation, autoimmune diseases and cancer. The helicase and repressor domains (RD) of RIG-I recognize dsRNA and 5'-ppp RNA to activate the two amino-terminal caspase recruitment domains (CARDs) for signalling. Here, to understand the synergy between the helicase and the RD for RNA binding, and the contribution of ATP hydrolysis to RIG-I activation, we determined the structure of human RIG-I helicase-RD in complex with dsRNA and an ATP analogue. The helicase-RD organizes into a ring around dsRNA, capping one end, while contacting both strands using previously uncharacterized motifs to recognize dsRNA. Small-angle X-ray scattering, limited proteolysis and differential scanning fluorimetry indicate that RIG-I is in an extended and flexible conformation that compacts upon binding RNA. These results provide a detailed view of the role of helicase in dsRNA recognition, the synergy between the RD and the helicase for RNA binding and the organization of full-length RIG-I bound to dsRNA, and provide evidence of a conformational change upon RNA binding. The RIG-I helicase-RD structure is consistent with dsRNA translocation without unwinding and cooperative binding to RNA. The structure yields unprecedented insight into innate immunity and has a broader impact on other areas of biology, including RNA

  19. Human-specific evolution of killer cell immunoglobulin-like receptor recognition of major histocompatibility complex class I molecules.

    PubMed

    Parham, Peter; Norman, Paul J; Abi-Rached, Laurent; Guethlein, Lisbeth A

    2012-03-19

    In placental mammals, natural killer (NK) cells are a population of lymphocytes that make unique contributions to immune defence and reproduction, functions essential for survival of individuals, populations and species. Modulating these functions are conserved and variable NK-cell receptors that recognize epitopes of major histocompatibility complex (MHC) class I molecules. In humans, for example, recognition of human leucocyte antigen (HLA)-E by the CD94:NKG2A receptor is conserved, whereas recognition of HLA-A, B and C by the killer cell immunoglobulin-like receptors (KIRs) is diversified. Competing demands of the immune and reproductive systems, and of T-cell and NK-cell immunity-combined with the segregation on different chromosomes of variable NK-cell receptors and their MHC class I ligands-drive an unusually rapid evolution that has resulted in unprecedented levels of species specificity, as first appreciated from comparison of mice and humans. Counterparts to human KIR are present only in simian primates. Observed in these species is the coevolution of KIR and the four MHC class I epitopes to which human KIR recognition is restricted. Unique to hominids is the emergence of the MHC-C locus as a supplier of specialized and superior ligands for KIR. This evolutionary trend is most highly elaborated in the chimpanzee. Unique to the human KIR locus are two groups of KIR haplotypes that are present in all human populations and subject to balancing selection. Group A KIR haplotypes resemble chimpanzee KIR haplotypes and are enriched for genes encoding KIR that bind HLA class I, whereas group B KIR haplotypes are enriched for genes encoding receptors with diminished capacity to bind HLA class I. Correlating with their balance in human populations, B haplotypes favour reproductive success, whereas A haplotypes favour successful immune defence. Evolution of the B KIR haplotypes is thus unique to the human species.

  20. Receptor architecture of visual areas in the face and word-form recognition region of the posterior fusiform gyrus.

    PubMed

    Caspers, Julian; Palomero-Gallagher, Nicola; Caspers, Svenja; Schleicher, Axel; Amunts, Katrin; Zilles, Karl

    2015-01-01

    Recently, two extrastriate visual areas on the posterior fusiform gyrus, areas FG1 and FG2, were identified based on cytoarchitectonical criteria (Caspers et al. in Brain Struct Funct 218:511-526, 2013a). They are located within the object-related ventral visual stream at the transition between early and higher-order (category-specific) visual areas. FG2 has a topographical position which is best comparable to the face or visual word-form recognition area. However, the precise function of FG2 is presently unknown. Since transmitter receptors are key molecules of neurotransmission, we analysed the regional and laminar distribution of 15 different receptor binding sites by means of quantitative in vitro receptor autoradiography. Significant differences between receptor densities of both areas were found for NMDA, GABAB, M3, nicotinic α4/β2 and 5-HT1A receptors as well as for GABAA associated benzodiazepine binding sites. These results support the cytoarchitectonic segregation of FG1 and FG2 into two distinct cortical areas. In addition, principal component and hierarchical cluster analyses of the multireceptor data of both fusiform areas and 24 visual, auditory, somatosensory and multimodal association areas not only revealed the typical receptor architectonic characteristics of visual areas for FG1 and FG2, but also suggest their putative function as object recognition regions due to the similarity of their receptor fingerprints with those of areas of the ventral visual stream. Furthermore, FG1 and FG2 build a cluster with the multimodal association areas of the inferior parietal lobule. This underlines their hierarchically high position in the visual system of the human cerebral cortex.

  1. Substrate Recognition by the Cdh1 Destruction Box Receptor Is a General Requirement for APC/CCdh1-mediated Proteolysis.

    PubMed

    Qin, Liang; Guimarães, Dimitrius Santiago P S F; Melesse, Michael; Hall, Mark C

    2016-07-22

    The anaphase-promoting complex, or cyclosome (APC/C), is a ubiquitin ligase that selectively targets proteins for degradation in mitosis and the G1 phase and is an important component of the eukaryotic cell cycle control system. How the APC/C specifically recognizes its substrates is not fully understood. Although well characterized degron motifs such as the destruction box (D-box) and KEN-box are commonly found in APC/C substrates, many substrates apparently lack these motifs. A variety of alternative APC/C degrons have been reported, suggesting either that multiple modes of substrate recognition are possible or that our definitions of degron structure are incomplete. We used an in vivo yeast assay to compare the G1 degradation rate of 15 known substrates of the APC/C co-activator Cdh1 under normal conditions and conditions that impair binding of D-box, KEN-box, and the recently identified ABBA motif degrons to Cdh1. The D-box receptor was required for efficient proteolysis of all Cdh1 substrates, despite the absence of canonical D-boxes in many. In contrast, the KEN-box receptor was only required for normal proteolysis of a subset of substrates and the ABBA motif receptor for a single substrate in our system. Our results suggest that binding to the D-box receptor may be a shared requirement for recognition and processing of all Cdh1 substrates.

  2. Matricryptic sites control tissue injury responses in the cardiovascular system: relationships to pattern recognition receptor regulated events.

    PubMed

    Davis, George E

    2010-03-01

    This review addresses new concepts related to the importance of how cells within the cardiovascular system respond to matricryptic sites generated from the extracellular matrix (ECM) following tissue injury. A model is presented whereby matricryptic sites exposed from the ECM result in activation of multiple cell surface receptors including integrins, scavenger receptors, and toll-like receptors which together are hypothesized to coactivate downstream signaling pathways which alter cell behaviors following tissue injury. Of great interest are the relationships between matricryptic fragments of ECM called matricryptins and other stimuli that activate cells during injury states such as released components from cells (DNA, RNA, cytoskeletal components such as actin) or products from infectious agents in innate immunity responses. These types of cell activating molecules, which are composed of repeating molecular elements, are known to interact with pattern recognition receptors that (i) are expressed from cell surfaces, (ii) are released from cells following tissue injury, or (iii) circulate as components of plasma. Thus, cell recognition of matricryptic sites from the ECM appears to be an important component of a broad cell and tissue sensory system to detect and respond to environmental cues generated following varied types of tissue injury.

  3. Type I interferon and pattern recognition receptor signaling following particulate matter inhalation

    PubMed Central

    2012-01-01

    Background Welding, a process that generates an aerosol containing gases and metal-rich particulates, induces adverse physiological effects including inflammation, immunosuppression and cardiovascular dysfunction. This study utilized microarray technology and subsequent pathway analysis as an exploratory search for markers/mechanisms of in vivo systemic effects following inhalation. Mice were exposed by inhalation to gas metal arc – stainless steel (GMA-SS) welding fume at 40 mg/m3 for 3 hr/d for 10 d and sacrificed 4 hr, 14 d and 28 d post-exposure. Whole blood cells, aorta and lung were harvested for global gene expression analysis with subsequent Ingenuity Pathway Analysis and confirmatory qRT-PCR. Serum was collected for protein profiling. Results The novel finding was a dominant type I interferon signaling network with the transcription factor Irf7 as a central component maintained through 28 d. Remarkably, these effects showed consistency across all tissues indicating a systemic type I interferon response that was complemented by changes in serum proteins (decreased MMP-9, CRP and increased VCAM1, oncostatin M, IP-10). In addition, pulmonary expression of interferon α and β and Irf7 specific pattern recognition receptors (PRR) and signaling molecules (Ddx58, Ifih1, Dhx58, ISGF3) were induced, an effect that showed specificity when compared to other inflammatory exposures. Also, a canonical pathway indicated a coordinated response of multiple PRR and associated signaling molecules (Tlr7, Tlr2, Clec7a, Nlrp3, Myd88) to inhalation of GMA-SS. Conclusion This methodological approach has the potential to identify consistent, prominent and/or novel pathways and provides insight into mechanisms that contribute to pulmonary and systemic effects following toxicant exposure. PMID:22776377

  4. Sequence variability of the pattern recognition receptor Mermaid mediates specificity of marine nematode symbioses.

    PubMed

    Bulgheresi, Silvia; Gruber-Vodicka, Harald R; Heindl, Niels R; Dirks, Ulrich; Kostadinova, Maria; Breiteneder, Heimo; Ott, Joerg A

    2011-06-01

    Selection of a specific microbial partner by the host is an all-important process. It guarantees the persistence of highly specific symbioses throughout host generations. The cuticle of the marine nematode Laxus oneistus is covered by a single phylotype of sulfur-oxidizing bacteria. They are embedded in a layer of host-secreted mucus containing the mannose-binding protein Mermaid. This Ca(2+)-dependent lectin mediates symbiont aggregation and attachment to the nematode. Here, we show that Stilbonema majum-a symbiotic nematode co-occurring with L. oneistus in shallow water sediment-is covered by bacteria phylogenetically distinct to those covering L. oneistus. Mermaid cDNA analysis revealed extensive protein sequence variability in both the nematode species. We expressed three recombinant Mermaid isoforms, which based on the structural predictions display the most different carbohydrate recognition domains (CRDs). We show that the three CRDs (DNT, DDA and GDA types) possess different affinities for L. oneistus and S. majum symbionts. In particular, the GDA type, exclusively expressed by S. majum, displays highest agglutination activity towards its symbionts and lowest towards its L. oneistus symbionts. Moreover, incubation of L. oneistus in the GDA type does not result in complete symbiont detachment, whereas incubation in the other types does. This indicates that the presence of particular Mermaid isoforms on the nematode surface has a role in the attachment of specific symbionts. This is the first report of the functional role of sequence variability in a microbe-associated molecular patterns receptor in a beneficial association.

  5. Sequence variability of the pattern recognition receptor Mermaid mediates specificity of marine nematode symbioses

    PubMed Central

    Bulgheresi, Silvia; Gruber-Vodicka, Harald R; Heindl, Niels R; Dirks, Ulrich; Kostadinova, Maria; Breiteneder, Heimo; Ott, Joerg A

    2011-01-01

    Selection of a specific microbial partner by the host is an all-important process. It guarantees the persistence of highly specific symbioses throughout host generations. The cuticle of the marine nematode Laxus oneistus is covered by a single phylotype of sulfur-oxidizing bacteria. They are embedded in a layer of host-secreted mucus containing the mannose-binding protein Mermaid. This Ca2+-dependent lectin mediates symbiont aggregation and attachment to the nematode. Here, we show that Stilbonema majum—a symbiotic nematode co-occurring with L. oneistus in shallow water sediment—is covered by bacteria phylogenetically distinct to those covering L. oneistus. Mermaid cDNA analysis revealed extensive protein sequence variability in both the nematode species. We expressed three recombinant Mermaid isoforms, which based on the structural predictions display the most different carbohydrate recognition domains (CRDs). We show that the three CRDs (DNT, DDA and GDA types) possess different affinities for L. oneistus and S. majum symbionts. In particular, the GDA type, exclusively expressed by S. majum, displays highest agglutination activity towards its symbionts and lowest towards its L. oneistus symbionts. Moreover, incubation of L. oneistus in the GDA type does not result in complete symbiont detachment, whereas incubation in the other types does. This indicates that the presence of particular Mermaid isoforms on the nematode surface has a role in the attachment of specific symbionts. This is the first report of the functional role of sequence variability in a microbe-associated molecular patterns receptor in a beneficial association. PMID:21228893

  6. A fibrinogen-related protein identified from hepatopancreas of crayfish is a potential pattern recognition receptor.

    PubMed

    Chen, Qiming; Bai, Suhua; Dong, Chaohua

    2016-09-01

    Fibrinogen-related protein (FREP) family is a large group of proteins containing fibrinogen-like (FBG) domain and plays multiple physiological roles in animals. However, their immune functions in crayfish are not fully explored. In the present study, a novel fibrinogen-like protein (designated as PcFBN1) was identified and characterized from hepatopancreas of red swamp crayfish Procambarus clarkii. The cDNA sequence of PcFBN1 contains an open reading frame (ORF) of 1353 bp encoding a protein of 450 amino acids. Sequence and structural analysis indicated that PcFBN1 contains an FBG domain in C-terminal and a putative signal peptide of 19 amino acids in N-terminal. Semi-quantitative PCR revealed that the main expression of PcFBN1 was observed in hepatopancreas and hemocyte. Temporal expression analysis exhibited that PcFBN1 expression could be significantly induced by heat-killed Aeromonas hydrophila. Tissue distribution and temporal change of PcFBN1 suggested that PcFBN1 may be involved in immune responses of red swamp crayfish. Recombinant PcFBN1 protein binds and agglutinates both gram-negative bacteria Escherichia coli and gram-positive bacteria Micrococcus lysodeikticus. Moreover, binding and agglutination is Ca(2+) dependent. Further analysis indicated that PcFBN1 recognizes some acetyl group-containing substance LPS and PGN. RNAi experiment revealed that PcFBN1 is required for bacterial clearance and survival from A. hydrophila infection. Reduction of PcFBN1 expression significantly decreased the survival and enhanced the number of A. hydrophila in the hemolymph. These results indicated that PcFBN1 plays an important role in the innate immunity of red swamp crayfish as a potential pattern recognition receptor.

  7. The molecular mechanism of species-specific recognition of lipopolysaccharides by the MD-2/TLR4 receptor complex.

    PubMed

    Oblak, Alja; Jerala, Roman

    2015-02-01

    Lipid A, a component of bacterial lipopolysaccharide, is a conserved microbe-associated molecular pattern that activates the MD-2/TLR4 receptor complex. Nevertheless, bacteria produce lipid A molecules of considerable structural diversity. The human MD-2/TLR4 receptor most efficiently recognizes hexaacylated bisphosphorylated lipid A produced by enterobacteria, but in some animal species the immune response can be elicited also by alternative lipid A varieties, such as tetraacylated lipid IVa or pentaacylated lipid A of Rhodobacter spheroides. Several crystal structures revealed that hexaacylated lipid A and tetraacylated lipid IVa activate the murine MD-2/TLR4 in a similar manner, but failed to explain the antagonistic vs. agonistic activity of lipid IVa in the human vs. equine receptor, respectively. Targeted mutagenesis studies of the receptor complex revealed intricate combination of electrostatic and hydrophobic interactions primarily within the MD-2 co-receptor, but with a contribution of TLR4 as well, that contribute to species-specific recognition of lipid A. We will review current knowledge regarding lipid A diversity and species-specific activation of the MD-2/TLR4 receptor complex in different species (e.g. human, mouse or equine) by lipid A varieties.

  8. Distinct roles of the hippocampus and perirhinal cortex in GABAA receptor blockade-induced enhancement of object recognition memory.

    PubMed

    Kim, Jong Min; Kim, Dong Hyun; Lee, Younghwan; Park, Se Jin; Ryu, Jong Hoon

    2014-03-13

    It is well known that the hippocampus plays a role in spatial and contextual memory, and that spatial information is tightly regulated by the hippocampus. However, it is still highly controversial whether the hippocampus plays a role in object recognition memory. In a pilot study, the administration of bicuculline, a GABAA receptor antagonist, enhanced memory in the passive avoidance task, but not in the novel object recognition task. In the present study, we hypothesized that these different results are related to the characteristics of each task and the different roles of hippocampus and perirhinal cortex. A region-specific drug-treatment model was employed to clarify the role of the hippocampus and perirhinal cortex in object recognition memory. After a single habituation in the novel object recognition task, intra-perirhinal cortical injection of bicuculline increased and intra-hippocampal injection decreased the exploration time ratio to novel object. In addition, when animals were repeatedly habituated to the context, intra-perirhinal cortical administration of bicuculline still increased exploration time ratio to novel object, but the effect of intra-hippocampal administration disappeared. Concurrent increases of c-Fos expression and ERK phosphorylation were observed in the perirhinal cortex of the object with context-exposed group either after single or repeated habituation to the context, but no changes were noted in the hippocampus. Altogether, these results suggest that object recognition memory formation requires the perirhinal cortex but not the hippocampus, and that hippocampal activation interferes with object recognition memory by the information encoding of unfamiliar environment.

  9. Differential Involvement of Dopamine D1 Receptor and MEK Signaling Pathway in the Ventromedial Prefrontal Cortex in Consolidation and Reconsolidation of Recognition Memory

    ERIC Educational Resources Information Center

    Maroun, Mouna; Akirav, Irit

    2009-01-01

    We investigated MEK and D1 receptors in the ventromedial prefrontal cortex (vmPFC) in consolidation and reconsolidation of recognition memory in rats nonhabituated to the experimental context (NH) or with reduced arousal due to extensive prior habituation (H). The D1 receptor antagonist enhanced consolidation and impaired reconsolidation in NH but…

  10. Solution NMR studies provide structural basis for endotoxin pattern recognition by the innate immune receptor CD14

    SciTech Connect

    Albright, Seth; Chen Bin; Holbrook, Kristen; Jain, Nitin U.

    2008-04-04

    CD14 functions as a key pattern recognition receptor for a diverse array of Gram-negative and Gram-positive cell-wall components in the host innate immune response by binding to pathogen-associated molecular patterns (PAMPs) at partially overlapping binding site(s). To determine the potential contribution of CD14 residues in this pattern recognition, we have examined using solution NMR spectroscopy, the binding of three different endotoxin ligands, lipopolysaccharide, lipoteichoic acid, and a PGN-derived compound, muramyl dipeptide to a {sup 15}N isotopically labeled 152-residue N-terminal fragment of sCD14 expressed in Pichia pastoris. Mapping of NMR spectral changes upon addition of ligands revealed that the pattern of residues affected by binding of each ligand is partially similar and partially different. This first direct structural observation of the ability of specific residue combinations of CD14 to differentially affect endotoxin binding may help explain the broad specificity of CD14 in ligand recognition and provide a structural basis for pattern recognition. Another interesting finding from the observed spectral changes is that the mode of binding may be dynamically modulated and could provide a mechanism for binding endotoxins with structural diversity through a common binding site.

  11. Isopropylamino and isobutylamino groups as recognition sites for carbohydrates: acyclic receptors with enhanced binding affinity toward β-galactosides.

    PubMed

    Mazik, Monika; Sonnenberg, Claudia

    2010-10-01

    Binding motifs observed in the crystal structures of protein-carbohydrate complexes, in particular the participation of the isopropyl/isobutyl side chain of valine/leucine in the formation of van der Waals contacts, have inspired the design of new artificial carbohydrate receptors. The new compounds, containing a trisubstituted triethylbenzene core, were expected to recognize sugar molecules through a combination of NH···O and OH···N hydrogen bonds, CH···π interactions, and numerous van der Waals contacts. (1)H NMR spectroscopic titrations in competitive and noncompetitive media, as well as binding studies in two-phase systems, such as dissolution of solid carbohydrates in apolar media and phase transfer of sugars from aqueous into organic solvents, revealed effective recognition of neutral carbohydrates and β- vs α-anomer binding preferences in the recognition of glycosides as well as significantly increased binding affinity of the receptors toward β-galactoside in comparison with the previously described receptors.

  12. Fluorescent charge-assisted halogen-bonding macrocyclic halo-imidazolium receptors for anion recognition and sensing in aqueous media.

    PubMed

    Zapata, Fabiola; Caballero, Antonio; White, Nicholas G; Claridge, Tim D W; Costa, Paulo J; Félix, Vítor; Beer, Paul D

    2012-07-18

    The synthesis and anion binding properties of a new family of fluorescent halogen bonding (XB) macrocyclic halo-imidazolium receptors are described. The receptors contain chloro-, bromo-, and iodo-imidazolium motifs incorporated into a cyclic structure using naphthalene spacer groups. The large size of the iodine atom substituents resulted in the isolation of anti and syn conformers of the iodo-imidazoliophane, whereas the chloro- and bromo-imidazoliophane analogues exhibit solution dynamic conformational behavior. The syn iodo-imidazoliophane isomer forms novel dimeric isostructural XB complexes of 2:2 stoichiometry with bromide and iodide anions in the solid state. Solution phase DOSY NMR experiments indicate iodide recognition takes place via cooperative convergent XB-iodide 1:1 stoichiometric binding in aqueous solvent mixtures. (1)H NMR and fluorescence spectroscopic titration experiments with a variety of anions in the competitive CD(3)OD/D(2)O (9:1) aqueous solvent mixture demonstrated the bromo- and syn iodo-imidazoliophane XB receptors to bind selectively iodide and bromide respectively, and sense these halide anions exclusively via a fluorescence response. The protic-, chloro-, and anti iodo-imidazoliophane receptors proved to be ineffectual anion complexants in this aqueous methanolic solvent mixture. Computational DFT and molecular dynamics simulations corroborate the experimental observations that bromo- and syn iodo-imidazoliophane XB receptors form stable cooperative convergent XB associations with bromide and iodide.

  13. HLA class I, NKG2D, and natural cytotoxicity receptors regulate multiple myeloma cell recognition by natural killer cells.

    PubMed

    Carbone, Ennio; Neri, Paola; Mesuraca, Maria; Fulciniti, Mariateresa T; Otsuki, Takemi; Pende, Daniela; Groh, Veronika; Spies, Thomas; Pollio, Giuditta; Cosman, David; Catalano, Lucio; Tassone, Pierfrancesco; Rotoli, Bruno; Venuta, Salvatore

    2005-01-01

    The role of natural killer (NK) cells in multiple myeloma is not fully understood. Here, NK susceptibility of myeloma cells derived from distinct disease stages was evaluated in relation to major histocompatibility complex (MHC) class I, MHC class I chain-related protein A (MICA), MHC class I chain-related protein B (MICB), and UL16 binding protein (ULBP) expression. MHC class I molecules were hardly detectable on bone marrow cells of early-stage myeloma, while late-stage pleural effusion-derived cell lines showed a strong MHC class I expression. Conversely, a high MICA level was found on bone marrow myeloma cells, while it was low or not measurable on pleural effusion myeloma cells. The reciprocal surface expression of these molecules on bone marrow- and pleural effusion-derived cell was confirmed at mRNA levels. While bone marrow-derived myeloma cells were readily recognized by NK cells, pleural effusion-derived lines were resistant. NK protection of pleural effusion cells was MHC class I dependent. Receptor blocking experiments demonstrated that natural cytotoxicity receptor (NCR) and NK receptor member D of the lectin-like receptor family (NKG2D) were the key NK activating receptors for bone marrow-derived myeloma cell recognition. In ex vivo experiments patient's autologous fresh NK cells recognized bone marrow-derived myeloma cells. Our data support the hypothesis that NK cell cytotoxicity could sculpture myeloma and represents an important immune effector mechanism in controlling its intramedullary stages.

  14. The changing of the guard: the Pto/Prf receptor complex of tomato and pathogen recognition.

    PubMed

    Ntoukakis, Vardis; Saur, Isabel M L; Conlan, Brendon; Rathjen, John P

    2014-08-01

    One important model for disease resistance is the Prf recognition complex of tomato, which responds to different bacterial effectors. Prf incorporates a protein kinase called Pto as its recognition domain that mimics effector virulence targets, and activates resistance after interaction with specific effectors. Recent findings show that this complex is oligomeric, and reveal how this impacts mechanism. Oligomerisation brings two or more kinases into proximity, where they can phosphorylate each other after effector perception. Effector attack on one kinase activates another in trans, constituting a molecular trap for the effector. Oligomerisation of plant resistance proteins may be a general concept that broadens pathogen recognition and restricts the ability of pathogens to evolve virulence.

  15. Atomic force microscopy-based molecular studies on the recognition of immunogenic chlorinated ovalbumin by macrophage receptors.

    PubMed

    Zapotoczny, Szczepan; Biedroń, Rafał; Marcinkiewicz, Janusz; Nowakowska, Maria

    2012-02-01

    This report presents simple and reliable approach developed to study the specific recognition events between chlorinated ovalbumin (OVA) and macrophages using atomic force microscopy (AFM). Thanks to the elimination of nonspecific adhesion, the interactions of the native and chlorinated OVA with a membrane of macrophages could be quantified using exclusively the so-called adhesion frequency (AF). The proposed system not only enabled the application of AFM-based force measurements for such poorly defined ligand-receptor pairs but also significantly improved both the acquisition and the processing of the data. The proteins were immobilized on the gold-coated AFM tips from the aqueous solutions containing charged thiol adsorbates. Such surface dilution of the proteins ensured the presence of single or just a few macromolecules at the tip-surface contact. The formation of negatively charged monolayer on the tip dramatically limited its nonspecific interactions with the macrophage surface. In such systems, AF was used as a measure of the recognition events even if the interaction forces varied significantly for sets of measurements. The system with the native OVA, a weak immunogen, showed only negligible AF compared with 85% measured for the immunogenic chlorinated OVA. The AF values varied with the tip-macrophage contact time and loading velocity. Blocking of the receptors by the chlorinated OVA was also confirmed. The developed approach can be also used to study other ligand-receptor interactions in poorly defined biological systems with intrinsically broad distribution of the rupture forces, thus opening new fields for AFM-based recognition on molecular level.

  16. Posttraining activation of CB1 cannabinoid receptors in the CA1 region of the dorsal hippocampus impairs object recognition long-term memory.

    PubMed

    Clarke, Julia R; Rossato, Janine I; Monteiro, Siomara; Bevilaqua, Lia R M; Izquierdo, Iván; Cammarota, Martín

    2008-09-01

    Evidence indicates that brain endocannabinoids are involved in memory processing. However, the participation of CB1 and CB2 cannabinoid receptors in recognition memory has not been yet conclusively determined. Therefore, we evaluated the effect of the posttraining activation of hippocampal cannabinoid receptors on the consolidation of object recognition memory. Rats with infusion cannulae stereotaxically aimed to the CA1 region of the dorsal hippocampus were trained in an object recognition learning task involving exposure to two different stimulus objects. Memory retention was assessed at different times after training. In the test sessions, one of the objects presented during training was replaced by a novel one. When infused in the CA1 region immediately after training, the non-selective cannabinoid receptor agonist WIN-55,212-2 and the endocannabinoid membrane transporter inhibitor VDM-11 blocked long-term memory retention in a dose-dependent manner without affecting short-term memory, exploratory behavior, anxiety state or the functionality of the hippocampus. The amnesic effect of WIN-55,212-2 and VDM-11 was not due to state-dependency and was completely reversed by co-infusion of the CB1 receptor antagonist AM-251 and mimicked by the CB1 receptor agonist ACEA but not by the CB2 receptor agonists JWH-015 and palmitoylethanolamide. Our data indicate that activation of hippocampal CB1 receptors early after training hampers consolidation of object recognition memory.

  17. CB1 receptor antagonism in the granular insular cortex or somatosensory area facilitates consolidation of object recognition memory.

    PubMed

    O'Brien, Lesley D; Sticht, Martin A; Mitchnick, Krista A; Limebeer, Cheryl L; Parker, Linda A; Winters, Boyer D

    2014-08-22

    Cannabinoid agonists typically impair memory, whereas CB1 receptor antagonists enhance memory performance under specific conditions. The insular cortex has been implicated in object memory consolidation. Here we show that infusions of the CB1 receptor antagonist SR141716 enhances long-term object recognition memory in rats in a dose-dependent manner (facilitation with 1.5, but not 0.75 or 3 μg/μL) when administered into the granular insular cortex; the SR141716 facilitation was seen with a memory delay of 72 h, but not when the delay was shorter (1 h), consistent with enhancement of memory consolidation. Moreover, a sub-group of rats with cannulas placed in the somatosensory area were also facilitated. These results highlight the robust potential of cannabinoid antagonists to facilitate object memory consolidation, as well as the capacity for insular and somatosensory cortices to contribute to object processing, perhaps through enhancement of tactile representation.

  18. Targeting of plant pattern recognition receptor-triggered immunity by bacterial type-III secretion system effectors.

    PubMed

    Macho, Alberto P; Zipfel, Cyril

    2015-02-01

    During infection, microbes are detected by surface-localized pattern recognition receptors (PRRs), leading to an innate immune response that prevents microbial ingress. Therefore, successful pathogens must evade or inhibit PRR-triggered immunity to cause disease. In the past decade, a number of type-III secretion system effector (T3Es) proteins from plant pathogenic bacteria have been shown to suppress this layer of innate immunity. More recently, the detailed mechanisms of action have been defined for several of these effectors. Interestingly, effectors display a wide array of virulence targets, being able to prevent activation of immune receptors and to hijack immune signaling pathways. Besides being a fascinating example of pathogen-host co-evolution, effectors have also emerged as valuable tools to dissect important biological processes in host cells.

  19. Structural basis of receptor sulfotyrosine recognition by a CC chemokine: the N-terminal region of CCR3 bound to CCL11/eotaxin-1.

    PubMed

    Millard, Christopher J; Ludeman, Justin P; Canals, Meritxell; Bridgford, Jessica L; Hinds, Mark G; Clayton, Daniel J; Christopoulos, Arthur; Payne, Richard J; Stone, Martin J

    2014-11-04

    Trafficking of leukocytes in immune surveillance and inflammatory responses is activated by chemokines engaging their receptors. Sulfation of tyrosine residues in peptides derived from the eosinophil chemokine receptor CCR3 dramatically enhances binding to cognate chemokines. We report the structural basis of this recognition and affinity enhancement. We describe the structure of a CC chemokine (CCL11/eotaxin-1) bound to a fragment of a chemokine receptor: residues 8–23 of CCR3, including two sulfotyrosine residues. We also show that intact CCR3 is sulfated and sulfation enhances receptor activity. The CCR3 sulfotyrosine residues form hydrophobic, salt bridge and cation-p interactions with residues that are highly conserved in CC chemokines. However, the orientation of the chemokine relative to the receptor N terminus differs substantially from those observed for two CXC chemokines, suggesting that initial binding of the receptor sulfotyrosine residues guides subsequent steps in receptor activation, thereby influencing the receptor conformational changes and signaling.

  20. Structure of the Neisserial Outer Membrane Protein Opa60: Loop Flexibility Essential to Receptor Recognition and Bacterial Engulfment

    PubMed Central

    2015-01-01

    The structure and dynamics of Opa proteins, which we report herein, are responsible for the receptor-mediated engulfment of Neisseria gonorrheae or Neisseria meningitidis by human cells and can offer deep understanding into the molecular recognition of pathogen–host receptor interactions. Such interactions are vital to understanding bacterial pathogenesis as well as the mechanism of foreign body entry to a human cell, which may provide insights for the development of targeted pharmaceutical delivery systems. The size and dynamics of the extracellular loops of Opa60 required a hybrid refinement approach wherein membrane and distance restraints were used to generate an initial NMR structural ensemble, which was then further refined using molecular dynamics in a DMPC bilayer. The resulting ensemble revealed that the extracellular loops, which bind host receptors, occupy compact conformations, interact with each other weakly, and are dynamic on the nanosecond time scale. We predict that this conformational sampling is critical for enabling diverse Opa loop sequences to engage a common set of receptors. PMID:24813921

  1. Exchanging ligand-binding specificity between a pair of mouse olfactory receptor paralogs reveals odorant recognition principles

    PubMed Central

    Baud, Olivia; Yuan, Shuguang; Veya, Luc; Filipek, Slawomir; Vogel, Horst; Pick, Horst

    2015-01-01

    A multi-gene family of ~1000 G protein-coupled olfactory receptors (ORs) constitutes the molecular basis of mammalian olfaction. Due to the lack of structural data its remarkable capacity to detect and discriminate thousands of odorants remains poorly understood on the structural level of the receptor. Using site-directed mutagenesis we transferred ligand specificity between two functionally related ORs and thereby revealed amino acid residues of central importance for odorant recognition and discrimination of the two receptors. By exchanging two of three residues, differing at equivalent positions of the putative odorant binding site between the mouse OR paralogs Olfr73 (mOR-EG) and Olfr74 (mOR-EV), we selectively changed ligand preference but remarkably also signaling activation strength in both ORs. Computer modeling proposed structural details at atomic resolution how the very same odorant molecule might interact with different contact residues to induce different functional responses in two related receptors. Our findings provide a mechanistic explanation of how the olfactory system distinguishes different molecular aspects of a given odorant molecule, and unravel important molecular details of the combinatorial encoding of odorant identity at the OR level. PMID:26449412

  2. Gene expression values of pattern-recognition receptors in porcine leukocytes and their response to Salmonella enterica serovar Typhimurium infection.

    PubMed

    Osvaldova, Alena; Stepanova, Hana; Faldyna, Martin; Matiasovic, Jan

    2017-03-01

    Pattern-recognition receptors (PRRs) recognize pathogen-associated molecular patterns and play an important role in triggering innate immune responses. PRRs distribution and function is well documented in mice and humans, but studies in pigs are scarce. Salmonella enterica serovar Typhimurium is common pathogen found in pigs and was used as a model for interaction with PRRs. This study investigated expression of PRRs in porcine leukocyte subpopulations at the mRNA level. Eight subpopulations of leukocytes comprising NK cells, Th, Tc, double positive T cells and γδ T cells, B cells, monocytes and neutrophils were sorted, and the expression of 12 PRRs was measured, including selected Toll-like receptors and their co-receptors, NOD-like receptor NOD2, RP-105, CD14, and dectin. The highest expression rates of most PRRs were observed in monocytes and neutrophils. The B cells expressed high levels of TLR1, TLR6, TLR9, TLR10, and RP-105. Only monocytes and γδ T cells were found to respond to Salmonella enterica serovar Typhimurium infection by intensification of PRRs expression. In Th and B cells, PRRs mRNA down-regulation was detected after infection.

  3. Structure of an LDLR-RAP Complex Reveals a General Mode for Ligand Recognition by Lipoprotein Receptors

    SciTech Connect

    Fisher,C.; Beglova, N.; Blacklow, s.

    2006-01-01

    Proteins of the low-density lipoprotein receptor (LDLR) family are remarkable in their ability to bind an extremely diverse range of protein and lipoprotein ligands, yet the basis for ligand recognition is poorly understood. Here, we report the 1.26 Angstroms X-ray structure of a complex between a two-module region of the ligand binding domain of the LDLR and the third domain of RAP, an escort protein for LDLR family members. The RAP domain forms a three-helix bundle with two docking sites, one for each LDLR module. The mode of recognition at each site is virtually identical: three conserved, calcium-coordinating acidic residues from each LDLR module encircle a lysine side chain protruding from the second helix of RAP. This metal-dependent mode of electrostatic recognition, together with avidity effects resulting from the use of multiple sites, represents a general binding strategy likely to apply in the binding of other basic ligands to LDLR family proteins.

  4. Recognition of core and flanking amino acids of MHC class II-bound peptides by the T cell receptor.

    PubMed

    Sant'Angelo, Derek B; Robinson, Eve; Janeway, Charles A; Denzin, Lisa K

    2002-09-01

    CD4 T cells recognize peptides bound to major histocompatibility complex (MHC) class II molecules. Most MHC class II molecules have four binding pockets occupied by amino acids 1, 4, 6, and 9 of the minimal peptide epitope, while the residues at positions 2, 3, 5, 7, and 8 are available to interact with the T cell receptor (TCR). In addition MHC class II bound peptides have flanking residues situated outside of this peptide core. Here we demonstrate that the flanking residues of the conalbumin peptide bound to I-A(k) have no effect on recognition by the D10 TCR. To study the role of peptide flanks for recognition by a second TCR, we determined the MHC and TCR contacting amino acids of the I-A(b) bound Ealpha peptide. The Ealpha peptide is shown to bind I-A(b) using four alanines as anchor residues. TCR recognition of Ealpha peptides with altered flanking residues again suggested that, in general, no specific interactions occurred with the peptide flanks. However, using an HLA-DM-mediated technique to measure peptide binding to MHC class II molecules, we found that the peptide flanking residues contribute substantially to MHC binding.

  5. Structural basis for sulfation-dependent self-glycan recognition by the human immune-inhibitory receptor Siglec-8

    PubMed Central

    Pröpster, Johannes M.; Yang, Fan; Rabbani, Said; Ernst, Beat; Allain, Frédéric H.-T.

    2016-01-01

    Siglec-8 is a human immune-inhibitory receptor that, when engaged by specific self-glycans, triggers eosinophil apoptosis and inhibits mast cell degranulation, providing an endogenous mechanism to down-regulate immune responses of these central inflammatory effector cells. Here we used solution NMR spectroscopy to dissect the fine specificity of Siglec-8 toward different sialylated and sulfated carbohydrate ligands and determined the structure of the Siglec-8 lectin domain in complex with its prime glycan target 6′-sulfo sialyl Lewisx. A canonical motif for sialic acid recognition, extended by a secondary motif formed by unique loop regions, recognizing 6-O–sulfated galactose dictates tight specificity distinct from other Siglec family members and any other endogenous glycan recognition receptors. Structure-guided mutagenesis revealed key contacts of both interfaces to be equally essential for binding. Our work provides critical structural and mechanistic insights into how Siglec-8 selectively recognizes its glycan target, rationalizes the functional impact of site-specific glycan sulfation in modulating this lectin–glycan interaction, and will enable the rational design of Siglec-8–targeted agonists to treat eosinophil- and mast cell-related allergic and inflammatory diseases, such as asthma. PMID:27357658

  6. Pathogen recognition by Toll-like receptor 2 activates Weibel-Palade body exocytosis in human aortic endothelial cells.

    PubMed

    Into, Takeshi; Kanno, Yosuke; Dohkan, Jun-ichi; Nakashima, Misako; Inomata, Megumi; Shibata, Ken-ichiro; Lowenstein, Charles J; Matsushita, Kenji

    2007-03-16

    The endothelial cell-specific granule Weibel-Palade body releases vasoactive substances capable of modulating vascular inflammation. Although innate recognition of pathogens by Toll-like receptors (TLRs) is thought to play a crucial role in promotion of inflammatory responses, the molecular basis for early-phase responses of endothelial cells to bacterial pathogens has not fully been understood. We here report that human aortic endothelial cells respond to bacterial lipoteichoic acid (LTA) and synthetic bacterial lipopeptides, but not lipopolysaccharide or peptidoglycan, to induce Weibel-Palade body exocytosis, accompanied by release or externalization of the storage components von Willebrand factor and P-selectin. LTA could activate rapid Weibel-Palade body exocytosis through a TLR2- and MyD88-dependent mechanism without de novo protein synthesis. This process was at least mediated through MyD88-dependent phosphorylation and activation of phospholipase Cgamma. Moreover, LTA activated interleukin-1 receptor-associated kinase-1-dependent delayed exocytosis with de novo protein synthesis and phospholipase Cgamma-dependent activation of the NF-kappaB pathway. Increased TLR2 expression by transfection or interferon-gamma treatment increased TLR2-mediated Weibel-Palade body exocytosis, whereas reduced TLR2 expression under laminar flow decreased the response. Thus, we propose a novel role for TLR2 in induction of a primary proinflammatory event in aortic endothelial cells through Weibel-Palade body exocytosis, which may be an important step for linking innate recognition of bacterial pathogens to vascular inflammation.

  7. Vγ2Vδ2 T Cell Receptor Recognition of Prenyl Pyrophosphates is Dependent on all Complementarity Determining Regions1

    PubMed Central

    Wang, Hong; Fang, Zhimei; Morita, Craig T.

    2010-01-01

    γδ T cells differ from αβ T cells in the antigens they recognize and their functions in immunity. While most αβ T cell receptors (TCR) recognize peptides presented by MHC class I or II, human γδ T cells expressing Vγ2Vδ2 TCRs recognize nonpeptide prenyl pyrophosphates. To define the molecular basis for this recognition, the effect of mutations in the TCR complementarity-determining regions (CDR) was assessed. Mutations in all CDR loops altered recognition and cover a large footprint. Unlike murine γδ TCR recognition of the MHC class Ib T22 protein, there was no CDR3δ motif required for recognition because only 1 residue is required. Instead, the length and sequence of CDR3γ was key. Although a potential prenyl pyrophosphate-binding site was defined by Lys109 in Jγ1.2 and Arg51 in CDR2δ, the area outlined by critical mutations is much larger. These results show that prenyl pyrophosphate recognition is primarily by germline-encoded regions of the γδ TCR, allowing a high proportion of Vγ2Vδ2 TCRs to respond. This underscores its parallels to innate immune receptors. Our results also provide strong evidence for the existence of an antigen-presenting molecule for prenyl pyrophosphates. This is an author-produced version of a manuscript accepted for publication in The Journal of Immunology (The JI). The American Association of Immunologists, Inc. (AAI), publisher of The JI, holds the copyright to this manuscript. This version of the manuscript has not yet been copyedited or subjected to editorial proofreading by The JI; hence, it may differ from the final version published in The JI (online and in print). AAI (The JI) is not liable for errors or omissions in this author-produced version of the manuscript or in any version derived from it by the U.S. National Institutes of Health or any other third party. The final, citable version of record can be found at www.jimmunol.org. PMID:20483784

  8. The Vasopressin 1b Receptor Antagonist A-988315 Blocks Stress Effects on the Retrieval of Object-Recognition Memory

    PubMed Central

    Barsegyan, Areg; Atsak, Piray; Hornberger, Wilfried B; Jacobson, Peer B; van Gaalen, Marcel M; Roozendaal, Benno

    2015-01-01

    Stress-induced activation of the hypothalamo–pituitary–adrenocortical (HPA) axis and high circulating glucocorticoid levels are well known to impair the retrieval of memory. Vasopressin can activate the HPA axis by stimulating vasopressin 1b (V1b) receptors located on the pituitary. In the present study, we investigated the effect of A-988315, a selective and highly potent non-peptidergic V1b-receptor antagonist with good pharmacokinetic properties, in blocking stress effects on HPA-axis activity and memory retrieval. To study cognitive performance, male Sprague-Dawley rats were trained on an object-discrimination task during which they could freely explore two identical objects. Memory for the objects and their location was tested 24 h later. A-988315 (20 or 60 mg/kg) or water was administered orally 90 min before retention testing, followed 60 min later by stress of footshock exposure. A-988315 dose-dependently dampened stress-induced increases in corticosterone plasma levels, but did not significantly alter HPA-axis activity of non-stressed control rats. Most importantly, A-988315 administration prevented stress-induced impairment of memory retrieval on both the object-recognition and the object-location tasks. A-988315 did not alter the retention of non-stressed rats and did not influence the total time spent exploring the objects or experimental context in either stressed or non-stressed rats. Thus, these findings indicate that direct antagonism of V1b receptors is an effective treatment to block stress-induced activation of the HPA axis and the consequent impairment of retrieval of different aspects of recognition memory. PMID:25669604

  9. Activation of G-protein-coupled receptor 30 is sufficient to enhance spatial recognition memory in ovariectomized rats.

    PubMed

    Hawley, Wayne R; Grissom, Elin M; Moody, Nicole M; Dohanich, Gary P; Vasudevan, Nandini

    2014-04-01

    In ovariectomized rats, administration of estradiol, or selective estrogen receptor agonists that activate either the α or β isoforms, have been shown to enhance spatial cognition on a variety of learning and memory tasks, including those that capitalize on the preference of rats to seek out novelty. Although the effects of the putative estrogen G-protein-coupled receptor 30 (GPR30) on hippocampus-based tasks have been reported using food-motivated tasks, the effects of activation of GPR30 receptors on tasks that depend on the preference of rats to seek out spatial novelty remain to be determined. Therefore, the aim of the current study was to determine if short-term treatment of ovariectomized rats with G-1, an agonist for GPR30, would mimic the effects on spatial recognition memory observed following short-term estradiol treatment. In Experiment 1, ovariectomized rats treated with a low dose (1 μg) of estradiol 48 h and 24 h prior to the information trial of a Y-maze task exhibited a preference for the arm associated with the novel environment on the retention trial conducted 48 h later. In Experiment 2, treatment of ovariectomized rats with G-1 (25 μg) 48 h and 24 h prior to the information trial of a Y-maze task resulted in a greater preference for the arm associated with the novel environment on the retention trial. Collectively, the results indicated that short-term treatment of ovariectomized rats with a GPR30 agonist was sufficient to enhance spatial recognition memory, an effect that also occurred following short-term treatment with a low dose of estradiol.

  10. Dopamine receptor D4 (DRD4) gene modulates the influence of informational masking on speech recognition

    PubMed Central

    Xie, Zilong; Maddox, W. Todd; Knopik, Valerie S.; McGeary, John E.; Chandrasekaran, Bharath

    2014-01-01

    Listeners vary substantially in their ability to recognize speech in noisy environments. Here we examined the role of genetic variation on individual differences in speech recognition in various noise backgrounds. Background noise typically varies in the levels of energetic masking (EM) and informational masking (IM) imposed on target speech. Relative to EM, release from IM is hypothesized to place greater demand on executive function to selectively attend to target speech while ignoring competing noises. Recent evidence suggests that the long allele variant in exon III of the DRD4 gene, primarily expressed in the prefrontal cortex, may be associated with enhanced selective attention to goal-relevant high-priority information even in the face of interference. We investigated the extent to which this polymorphism is associated with speech recognition in IM and EM conditions. In an unscreened adult sample (Experiment 1) and a larger screened replication sample (Experiment 2), we demonstrate that individuals with the DRD4 long variant show better recognition performance in noise conditions involving significant IM, but not in EM conditions. In Experiment 2, we also obtained neuropsychological measures to assess the underlying mechanisms. Mediation analyses revealed that this listening condition-specific advantage was mediated by enhanced executive attention/working memory capacity in individuals with the long allele variant. These findings suggest that DRD4 may contribute specifically to individual differences in speech recognition ability in noise conditions that place demands on executive function. PMID:25497692

  11. Dopamine receptor D4 (DRD4) gene modulates the influence of informational masking on speech recognition.

    PubMed

    Xie, Zilong; Maddox, W Todd; Knopik, Valerie S; McGeary, John E; Chandrasekaran, Bharath

    2015-01-01

    Listeners vary substantially in their ability to recognize speech in noisy environments. Here we examined the role of genetic variation on individual differences in speech recognition in various noise backgrounds. Background noise typically varies in the levels of energetic masking (EM) and informational masking (IM) imposed on target speech. Relative to EM, release from IM is hypothesized to place greater demand on executive function to selectively attend to target speech while ignoring competing noises. Recent evidence suggests that the long allele variant in exon III of the DRD4 gene, primarily expressed in the prefrontal cortex, may be associated with enhanced selective attention to goal-relevant high-priority information even in the face of interference. We investigated the extent to which this polymorphism is associated with speech recognition in IM and EM conditions. In an unscreened adult sample (Experiment 1) and a larger screened replication sample (Experiment 2), we demonstrate that individuals with the DRD4 long variant show better recognition performance in noise conditions involving significant IM, but not in EM conditions. In Experiment 2, we also obtained neuropsychological measures to assess the underlying mechanisms. Mediation analysis revealed that this listening condition-specific advantage was mediated by enhanced executive attention/working memory capacity in individuals with the long allele variant. These findings suggest that DRD4 may contribute specifically to individual differences in speech recognition ability in noise conditions that place demands on executive function.

  12. Pattern Recognition Scavenger Receptor A/CD204 Regulates Airway Inflammatory Homeostasis Following Organic Dust Extract Exposures

    PubMed Central

    Poole, Jill A.; Anderson, Leigh; Gleason, Angela M.; West, William W.; Romberger, Debra J.; Wyatt, Todd A.

    2014-01-01

    Exposure to agriculture organic dusts, comprised of a diversity of pathogen-associated molecular patterns, results in chronic airway diseases. The multi-functional class A macrophage scavenger receptor (SRA)/CD204 has emerged as an important class of pattern recognition receptors with broad ligand binding ability. Our objective was to determine the role of SRA in mediating repetitive and post-inflammatory organic dust extract (ODE)-induced airway inflammation. Wild-type (WT) and SRA knockout (KO) mice were intra-nasally treated with ODE or saline daily for 3 wk and immediately euthanized or allowed to recover for 1 wk. Results show that lung histopathologic changes were increased in SRA KO mice as compared to WT following repetitive ODE exposures marked predominately by increased size and distribution of lymphoid aggregates. After a 1-wk recovery from daily ODE treatments, there was significant resolution of lung injury in WT mice, but not SRA KO animals. The increased lung histopathology induced by ODE treatment was associated with decreased accumulation of neutrophils, but greater accumulation of CD4+ T-cells. The lung cytokine milieu induced by ODE was consistent with a TH1/TH17 polarization in both WT and SRA KO mice. Overall, our data demonstrate that SRA/CD204 plays an important role in the normative inflammatory lung response to ODE as evidenced by the enhanced dust-mediated injury viewed in the absence of this receptor. PMID:24491035

  13. Structures of the Signal Recognition Particle Receptor From the Archaeon Pyrococcus Furiosus: Implications for the Targeting Step at the Membrane

    SciTech Connect

    Egea, P.F.; Tsuruta, H.; Leon, G.P.de; Napetschnig, J.; Walter, P.; Stroud, R.M.

    2009-05-18

    In all organisms, a ribonucleoprotein called the signal recognition particle (SRP) and its receptor (SR) target nascent proteins from the ribosome to the translocon for secretion or membrane insertion. We present the first X-ray structures of an archeal FtsY, the receptor from the hyper-thermophile Pyrococcus furiosus (Pfu), in its free and GDP {center_dot} magnesium-bound forms. The highly charged N-terminal domain of Pfu-FtsY is distinguished by a long N-terminal helix. The basic charges on the surface of this helix are likely to regulate interactions at the membrane. A peripheral GDP bound near a regulatory motif could indicate a site of interaction between the receptor and ribosomal or SRP RNAs. Small angle X-ray scattering and analytical ultracentrifugation indicate that the crystal structure of Pfu-FtsY correlates well with the average conformation in solution. Based on previous structures of two sub-complexes, we propose a model of the core of archeal and eukaryotic SRP {center_dot} SR targeting complexes.

  14. Selective recognition of sulfate anions by a cyclopeptide-derived receptor in aqueous phosphate buffer.

    PubMed

    Schaly, Astrid; Belda, Raquel; García-España, Enrique; Kubik, Stefan

    2013-12-20

    A cyclopeptide-based anion receptor containing alternating 6-aminopicolinic acid and substituted (4R)-4-aminoproline subunits with appended β-alanine residues binds sulfate anions in water. Importantly, appreciable sulfate binding is even observed in phosphate buffer, hence in the presence of anions of similar structure but with a different degree of protonation. The cause for the high selectivity of this receptor is related to the mode of action of the sulfate-binding protein.

  15. Danger- and pathogen-associated molecular patterns recognition by pattern-recognition receptors and ion channels of the transient receptor potential family triggers the inflammasome activation in immune cells and sensory neurons.

    PubMed

    Santoni, Giorgio; Cardinali, Claudio; Morelli, Maria Beatrice; Santoni, Matteo; Nabissi, Massimo; Amantini, Consuelo

    2015-02-03

    An increasing number of studies show that the activation of the innate immune system and inflammatory mechanisms play an important role in the pathogenesis of numerous diseases. The innate immune system is present in almost all multicellular organisms and its activation occurs in response to pathogens or tissue injury via pattern-recognition receptors (PRRs) that recognize pathogen-associated molecular patterns (PAMPs) or danger-associated molecular patterns (DAMPs). Intracellular pathways, linking immune and inflammatory response to ion channel expression and function, have been recently identified. Among ion channels, the transient receptor potential (TRP) channels are a major family of non-selective cation-permeable channels that function as polymodal cellular sensors involved in many physiological and pathological processes. In this review, we summarize current knowledge of interactions between immune cells and PRRs and ion channels of TRP families with PAMPs and DAMPs to provide new insights into the pathogenesis of inflammatory diseases. TRP channels have been found to interfere with innate immunity via both nuclear factor-kB and procaspase-1 activation to generate the mature caspase-1 that cleaves pro-interleukin-1β cytokine into the mature interleukin-1β.Sensory neurons are also adapted to recognize dangers by virtue of their sensitivity to intense mechanical, thermal and irritant chemical stimuli. As immune cells, they possess many of the same molecular recognition pathways for danger. Thus, they express PRRs including Toll-like receptors 3, 4, 7, and 9, and stimulation by Toll-like receptor ligands leads to induction of inward currents and sensitization in TRPs. In addition, the expression of inflammasomes in neurons and the involvement of TRPs in central nervous system diseases strongly support a role of TRPs in inflammasome-mediated neurodegenerative pathologies. This field is still at its beginning and further studies may be required.Overall, these

  16. Preparation and characterization of novel molecularly imprinted polymers based on thiourea receptors for nitrocompounds recognition.

    PubMed

    Athikomrattanakul, Umporn; Katterle, Martin; Gajovic-Eichelmann, Nenad; Scheller, Frieder W

    2011-04-15

    Molecularly imprinted polymers (MIPs) for the recognition of nitro derivatives are prepared from three different (thio)urea-bearing functional monomers. The binding capability of the polymers is characterized by a batch binding experiment. The imprinting factors and affinity constants (K) of the imprinted polymers exhibit the same tendency as the binding constants (K(a)) of the functional monomers to the target substance in solution. Not only nitrofurantoin is efficiently bound by these MIPs but also a broad spectrum of other nitro compounds is bound with at the intermediate level, addressing that these (thio)urea-based monomers can be utilized to prepare a family of MIPs for various nitro compounds, which can be applied as recognition elements in separation and analytical application.

  17. Molecular recognition of ketamine by a subset of olfactory G protein–coupled receptors

    PubMed Central

    Saven, Jeffery G.; Matsunami, Hiroaki; Eckenhoff, Roderic G.

    2015-01-01

    Ketamine elicits various neuropharmacological effects, including sedation, analgesia, general anesthesia, and antidepressant activity. Through an in vitro screen, we identified four mouse olfactory receptors (ORs) that responded to ketamine. In addition to their presence in the olfactory epithelium, these G protein (heterotrimeric guanine nucleotide–binding protein)–coupled receptors (GPCRs) are distributed throughout the central nervous system. To better understand the molecular basis of the interactions between ketamine and ORs, we used sequence comparison and molecular modeling to design mutations that (i) increased, reduced, or abolished ketamine responsiveness in responding receptors, and (ii) rendered non-responding receptors responsive to ketamine. We showed that olfactory sensory neurons (OSNs) that expressed distinct ORs responded to ketamine in vivo, suggesting that ORs may serve as functional targets for ketamine. The ability to both abolish and introduce responsiveness to ketamine in GPCRs enabled us to identify and confirm distinct interaction loci in the binding site, which suggested a signature ketamine-binding pocket that may guide exploration of additional receptors for this general anesthetic drug. PMID:25829447

  18. Activation by SLAM Family Receptors Contributes to NK Cell Mediated "Missing-Self" Recognition.

    PubMed

    Alari-Pahissa, Elisenda; Grandclément, Camille; Jeevan-Raj, Beena; Leclercq, Georges; Veillette, André; Held, Werner

    2016-01-01

    Natural Killer (NK) cells attack normal hematopoietic cells that do not express inhibitory MHC class I (MHC-I) molecules, but the ligands that activate NK cells remain incompletely defined. Here we show that the expression of the Signaling Lymphocyte Activation Molecule (SLAM) family members CD48 and Ly9 (CD229) by MHC-I-deficient tumor cells significantly contributes to NK cell activation. When NK cells develop in the presence of T cells or B cells that lack inhibitory MHC-I but express activating CD48 and Ly9 ligands, the NK cells' ability to respond to MHC-I-deficient tumor cells is severely compromised. In this situation, NK cells express normal levels of the corresponding activation receptors 2B4 (CD244) and Ly9 but these receptors are non-functional. This provides a partial explanation for the tolerance of NK cells to MHC-I-deficient cells in vivo. Activating signaling via 2B4 is restored when MHC-I-deficient T cells are removed, indicating that interactions with MHC-I-deficient T cells dominantly, but not permanently, impair the function of the 2B4 NK cell activation receptor. These data identify an important role of SLAM family receptors for NK cell mediated "missing-self" reactivity and suggest that NK cell tolerance in MHC-I mosaic mice is in part explained by an acquired dysfunction of SLAM family receptors.

  19. Buggy Creek virus (Togaviridae: Alphavirus) upregulates expression of pattern recognition receptors and interferons in House Sparrows (Passer domesticus).

    PubMed

    Fassbinder-Orth, Carol A; Barak, Virginia A; Rainwater, Ellecia L; Altrichter, Ashley M

    2014-06-01

    Birds serve as reservoirs for at least 10 arthropod-borne viruses, yet specific immune responses of birds to arboviral infections are relatively unknown. Here, adult House Sparrows were inoculated with an arboviral alphavirus, Buggy Creek virus (BCRV), or saline, and euthanized between 1 and 3 days postinoculation. Virological dynamics and gene expression dynamics were investigated. Birds did not develop viremia postinoculation, but cytopathic virus was found in the skeletal muscle and spleen of birds 1 and 3 days postinoculation (DPI). Viral RNA was detected in the blood of BCRV-infected birds 1 and 2 DPI, in oral swabs 1-3 DPI, and in brain, heart, skeletal muscle, and spleen 1-3 DPI. Multiple genes were significantly upregulated following BCRV infection, including pattern recognition receptors (TLR7, TLR15, RIG-1), type I interferon (IFN-α), and type II interferon (IFN-γ). This is the first study to report avian immunological gene expression profiles following an arboviral infection.

  20. Transcriptome Profiling Reveals Higher Vertebrate Orthologous of Intra-Cytoplasmic Pattern Recognition Receptors in Grey Bamboo Shark

    PubMed Central

    Gupta, Ravi; Gopal, Dhinakar Raj; Rajesh, Preeti; Chidambaram, Balachandran; Kalyanasundaram, Aravindan; Angamuthu, Raja

    2014-01-01

    From an immunologist perspective, sharks are an important group of jawed cartilaginous fishes and survey of the public database revealed a great gap in availability of large-scale sequence data for the group of Chondrichthyans the elasmobranchs. In an attempt to bridge this deficit we generated the transcriptome from the spleen and kidney tissues (a total of 1,606,172 transcripts) of the shark, Chiloscyllium griseum using the Illumina HiSeq2000 platform. With a cut off of > = 300 bp and an expression value of >1RPKM we used 43,385 transcripts for BLASTX analysis which revealed 17,548 transcripts matching to the NCBI nr database with an E-value of < = 10−5 and similarity score of 40%. The longest transcript was 16,974 bases with matched to HECT domain containing E3 ubiqutin protein ligase. MEGAN4 annotation pipeline revealed immune and signalling pathways including cell adhesion molecules, cytokine-cytokine receptor interaction, T-cell receptor signalling pathway and chemokine signaling pathway to be highly expressed in spleen, while different metabolism pathways such as amino acid metabolism, carbohydrate metabolism, lipid metabolism and xenobiotic biodegradation were highly expressed in kidney. Few of the candidate genes were selected to analyze their expression levels in various tissues by real-time PCR and also localization of a receptor by in-situ PCR to validate the prediction. We also predicted the domains structures of some of the identified pattern recognition receptors, their phylogenetic relationship with lower and higher vertebrates and the complete downstream signaling mediators of classical dsRNA signaling pathway. The generated transcriptome will be a valuable resource to further genetic and genomic research in elasmobranchs. PMID:24956167

  1. Recognition and sensing of low-epitope targets via ternary complexes with oligonucleotides and synthetic receptors

    NASA Astrophysics Data System (ADS)

    Yang, Kyung-Ae; Barbu, Mihaela; Halim, Marlin; Pallavi, Payal; Kim, Benjamin; Kolpashchikov, Dmitry M.; Pecic, Stevan; Taylor, Steven; Worgall, Tilla S.; Stojanovic, Milan N.

    2014-11-01

    Oligonucleotide-based receptors or aptamers can interact with small molecules, but the ability to achieve high-affinity and specificity of these interactions depends strongly on functional groups or epitopes displayed by the binding targets. Some classes of targets are particularly challenging: for example, monosaccharides have scarce functionalities and no aptamers have been reported to recognize, let alone distinguish from each other, glucose and other hexoses. Here we report aptamers that differentiate low-epitope targets such as glucose, fructose or galactose by forming ternary complexes with high-epitope organic receptors for monosaccharides. In a follow-up example, we expand this method to isolate high-affinity oligonucleotides against aromatic amino acids complexed in situ with a nonspecific organometallic receptor. The method is general and enables broad clinical use of aptamers for the detection of small molecules in mix-and-measure assays, as demonstrated by monitoring postprandial waves of phenylalanine in human subjects.

  2. NMR structure of the natural killer cell receptor 2B4 (CD244): implications for ligand recognition.

    PubMed

    Ames, James B; Vyas, Vinay; Lusin, Jacqueline D; Mariuzza, Roy

    2005-05-03

    2B4, a transmembrane receptor expressed primarily on natural killer (NK) cells and on a subset of CD8(+) T cells, plays an important role in activating NK-mediated cytotoxicity through its interaction with CD48 on target cells. We report here the atomic-resolution structure of the ligand-binding (D1) domain of 2B4 in solution determined by nuclear magnetic resonance (NMR) spectroscopy. The overall main chain structure resembles an immunoglobulin variable (V) domain fold, very similar to that seen previously for domain 1 of CD2 and CD4. The structure contains nine beta-strands assembled into two beta-sheets conventionally labeled DEB and AGFCC'C' '. The six-stranded sheet (AGFCC'C' ') contains structural features that may have implications for ligand recognition and receptor function. A noncanonical disulfide bridge between Cys2 and Cys99 stabilizes a long and parallel beta-structure between strand A (residues 3-12) and strand G (residues 100-108). A beta-bulge at residues Glu45 and Ile46 places a bend in the middle of strand C' that orients two conserved and adjacent hydrophobic residues (Ile46 and Leu47) inside the beta-sandwich as seen in other V domains. Finally, the FG-loop (implicated in ligand recognition in the CD2-CD58 complex) is dynamically disordered in 2B4 in the absence of a ligand. We propose that ligand binding to 2B4 might stabilize the structure of the FG-loop in the ligand complex.

  3. Chemokine production and pattern recognition receptor (PRR) expression in whole blood stimulated with pathogen-associated molecular patterns (PAMPs).

    PubMed

    Møller, Anne-Sophie W; Ovstebø, Reidun; Haug, Kari Bente F; Joø, Gun Britt; Westvik, Ase-Brit; Kierulf, Peter

    2005-12-21

    Recognition of conserved bacterial structures called pathogen-associated molecular patterns (PAMPs) by pattern recognition receptors (PRRs), may lead to induction of a variety of "early immediate genes" such as chemokines. In the current study, we have in an ex vivo whole blood model studied the induction of the chemokines MIP-1alpha, MCP-1 and IL-8 by various PAMPs. The rate of appearance of Escherichia coli-Lipopolysaccharide (LPS) induced chemokines differed. The production of MIP-1alpha and IL-8 was after 1 h of stimulation significantly higher when compared to unstimulated whole blood, whereas MCP-1 was not significantly elevated until after 3 h. At peak levels the MIP-1alpha concentration induced by E. coli-LPS was 3-5-fold higher than MCP-1 and IL-8. By specific cell depletion, we demonstrated that all three chemokines were mainly produced by monocytes. However, the mRNA results showed that IL-8 was induced in both monocytes and granulocytes. The production of all three chemokines, induced by the E. coli-LPS and Neisseria meningitidis-LPS, was significantly inhibited by antibodies against CD14 and TLR4, implying these receptors to be of importance for the effects of LPS in whole blood. The chemokine production induced by lipoteichoic acid (LTA) and non-mannose-capped lipoarabinomannan (AraLAM) was, however, less efficiently blocked by antibodies against CD14 and TLR2. E. coli-LPS and LTA induced a dose-dependent increase of CD14, TLR2 and TLR4 expression on monocytes in whole blood. These data show that PAMPs may induce chemokine production in whole blood and that antibodies against PRRs inhibit the production to different extent.

  4. T cell receptor recognition of a 'super-bulged' major histocompatibility complex class I-bound peptide

    SciTech Connect

    Tynan, Fleur E; Burrows, Scott R; Buckle, Ashley M; Clements, Craig S; Borg, Natalie A; Miles, John J; Beddoe, Travis; Whisstock, James C; Wilce, Matthew C; Silins, Sharon L; Burrows, Jacqueline M; Kjer-Nielsen, Lars; Kostenko, Lyudmila; Purcell, Anthony W; McCluskey, James; Rossjohn, Jamie

    2010-07-20

    Unusually long major histocompatibility complex (MHC) class I-restricted epitopes are important in immunity, but their 'bulged' conformation represents a potential obstacle to {alpha}{beta} T cell receptor (TCR)-MHC class I docking. To elucidate how such recognition is achieved while still preserving MHC restriction, we have determined here the structure of a TCR in complex with HLA-B*3508 presenting a peptide 13 amino acids in length. This complex was atypical of TCR-peptide-MHC class I interactions, being dominated at the interface by peptide-mediated interactions. The TCR assumed two distinct orientations, swiveling on top of the centrally bulged, rigid peptide such that only limited contacts were made with MHC class I. Although the TCR-peptide recognition resembled an antibody-antigen interaction, the TCR-MHC class I contacts defined a minimal 'generic footprint' of MHC-restriction. Thus our findings simultaneously demonstrate the considerable adaptability of the TCR and the 'shape' of MHC restriction.

  5. Molecular determinants of the recognition of ulipristal acetate by oxo-steroid receptors.

    PubMed

    Petit-Topin, I; Fay, M; Resche-Rigon, M; Ulmann, A; Gainer, E; Rafestin-Oblin, M-E; Fagart, J

    2014-10-01

    The human progesterone receptor (PR) plays a key role in reproductive function in women. PR antagonists have numerous applications in female health care including regular and emergency contraception, and treatment of hormone-related pathological conditions such as breast cancer, endometriosis, and leiomyoma. The main factor limiting their long-term administration is the fact that they cross-bind to other oxo-steroid receptors. Ulipristal acetate (UPA), a highly potent PR antagonist, has recently come onto the market and is much more selective for PR than the other oxo-steroid receptors (androgen, AR, glucocorticoid, GR, and mineralocorticoid, MR receptors) and, remarkably, it displays lower GR-inactivating potency than RU486. We adopted a structural approach to characterizing the binding of UPA to the oxo-steroid receptors at the molecular level. We solved the X-ray crystal structure of the ligand-binding domain (LBD) of the human PR complexed with UPA and a peptide from the transcriptional corepressor SMRT. We used the X-ray crystal structure of the GR in its antagonist conformation to dock UPA within its ligand-binding cavity. Finally, we generated three-dimensional models of the LBD of androgen and mineralocorticoid receptors (AR and MR) in an antagonist conformation and docked UPA within them. Comparing the structures revealed that the network of stabilizing contacts between the UPA C11 aryl group and the LBD is responsible for its high PR antagonist potency. It also showed that it is the inability of UPA to contact Gln642 in GR that explains why it has lower potency in inactivating GR than RU486. Finally, we found that the binding pockets of AR and MR are too small to accommodate UPA, and allowed us to propose that the extremely low sensitivity of MR to UPA is due to inappropriate interactions with the C11 substituent. All these findings open new avenues for designing new PR antagonist compounds displaying greater selectivity.

  6. Structural basis for bifunctional peptide recognition at human δ-opioid receptor

    DOE PAGES

    Fenalti, Gustavo; Zatsepin, Nadia A.; Betti, Cecilia; ...

    2015-02-16

    Bi-functional μ- and δ- opioid receptor (OR) ligands are potential therapeutic alternatives to alkaloid opiate analgesics with diminished side effects. We solved the structure of human δ-OR bound to the bi-functional δ-OR antagonist and μ-OR agonist tetrapeptide H-Dmt-Tic-Phe-Phe-NH2 (DIPP-NH2) by serial femtosecond crystallography, revealing a cis-peptide bond between H-Dmt and Tic. In summary, the observed receptor-peptide interactions are critical to understand the pharmacological profiles of opioid peptides, and to develop improved analgesics.

  7. Structural basis for bifunctional peptide recognition at human δ-opioid receptor

    SciTech Connect

    Fenalti, Gustavo; Zatsepin, Nadia A.; Betti, Cecilia; Giguere, Patrick; Han, Gye Won; Ishchenko, Andrii; Liu, Wei; Guillemyn, Karel; Zhang, Haitao; James, Daniel; Wang, Dingjie; Weierstall, Uwe; Spence, John C. H.; Boutet, Sébastien; Messerschmidt, Marc; Williams, Garth J.; Gati, Cornelius; Yefanov, Oleksandr M.; White, Thomas A.; Oberthuer, Dominik; Metz, Markus; Yoon, Chun Hong; Barty, Anton; Chapman, Henry N.; Basu, Shibom; Coe, Jesse; Conrad, Chelsie E.; Fromme, Raimund; Fromme, Petra; Tourwé, Dirk; Schiller, Peter W.; Roth, Bryan L.; Ballet, Steven; Katritch, Vsevolod; Stevens, Raymond C.; Cherezov, Vadim

    2015-02-16

    Bi-functional μ- and δ- opioid receptor (OR) ligands are potential therapeutic alternatives to alkaloid opiate analgesics with diminished side effects. We solved the structure of human δ-OR bound to the bi-functional δ-OR antagonist and μ-OR agonist tetrapeptide H-Dmt-Tic-Phe-Phe-NH2 (DIPP-NH2) by serial femtosecond crystallography, revealing a cis-peptide bond between H-Dmt and Tic. In summary, the observed receptor-peptide interactions are critical to understand the pharmacological profiles of opioid peptides, and to develop improved analgesics.

  8. Structural basis for bifunctional peptide recognition at human δ-Opioid receptor

    PubMed Central

    Fenalti, Gustavo; Zatsepin, Nadia A.; Betti, Cecilia; Giguere, Patrick; Han, Gye Won; Ishchenko, Andrii; Liu, Wei; Guillemyn, Karel; Zhang, Haitao; James, Daniel; Wang, Dingjie; Weierstall, Uwe; Spence, John C.H.; Boutet, Sébastien; Messerschmidt, Marc; Williams, Garth J.; Gati, Cornelius; Yefanov, Oleksandr M.; White, Thomas A.; Oberthuer, Dominik; Metz, Markus; Yoon, Chun Hong; Barty, Anton; Chapman, Henry N.; Basu, Shibom; Coe, Jesse; Conrad, Chelsie E.; Fromme, Raimund; Fromme, Petra; Tourwé, Dirk; Schiller, Peter W.; Roth, Bryan L.; Ballet, Steven; Katritch, Vsevolod; Stevens, Raymond C.; Cherezov, Vadim

    2015-01-01

    Bi-functional μ- and δ- opioid receptor (OR) ligands are potential therapeutic alternatives to alkaloid opiate analgesics with diminished side effects. We solved the structure of human δ-OR bound to the bi-functional δ-OR antagonist and μ-OR agonist tetrapeptide H-Dmt(1)-Tic(2)-Phe(3)-Phe(4)-NH2 (DIPP-NH2) by serial femtosecond crystallography, revealing a cis-peptide bond between H-Dmt(1) and Tic(2). The observed receptor-peptide interactions are critical to understand the pharmacological profiles of opioid peptides, and to develop improved analgesics. PMID:25686086

  9. Calix[4]tetrahydrothiophenopyrrole: a ditopic receptor displaying a split personality for ion recognition.

    PubMed

    Saha, Indrajit; Park, Kyung Hwa; Han, Mina; Kim, Sung Kuk; Lynch, Vincent M; Sessler, Jonathan L; Lee, Chang-Hee

    2014-10-17

    A calix[4]pyrrole fused with 2,5-dihydrothiophene, possessing both a deep, π-electron-rich pocket upon anion binding and chelating ligands on the periphery, was developed. The receptor selectively forms an ion-pair complex with CsF through H-bonding and a cation-π interaction. In the process, it adopt a conformationally fixed cone conformation. The receptor displays exceptionally high affinity toward the Hg(II) ion and forms stable complexes while maintaining a rigid 1,3-alternate conformation. This metal ion-induced conformational locking is unprecedented in calix[4]pyrrole chemistry.

  10. Importance of the extracellular loops in G protein-coupled receptors for ligand recognition and receptor activation.

    PubMed

    Peeters, M C; van Westen, G J P; Li, Q; IJzerman, A P

    2011-01-01

    G protein-coupled receptors (GPCRs) are the major drug target of medicines on the market today. Therefore, much research is and has been devoted to the elucidation of the function and three-dimensional structure of this large family of membrane proteins, which includes multiple conserved transmembrane domains connected by intra- and extracellular loops. In the last few years, the less conserved extracellular loops have garnered increasing interest, particularly after the publication of several GPCR crystal structures that clearly show the extracellular loops to be involved in ligand binding. This review will summarize the recent progress made in the clarification of the ligand binding and activation mechanism of class-A GPCRs and the role of extracellular loops in this process.

  11. Structures of human folate receptors reveal biological trafficking states and diversity in folate and antifolate recognition

    PubMed Central

    Wibowo, Ardian S.; Singh, Mirage; Reeder, Kristen M.; Carter, Joshua J.; Kovach, Alexander R.; Meng, Wuyi; Ratnam, Manohar; Zhang, Faming; Dann, Charles E.

    2013-01-01

    Antifolates, folate analogs that inhibit vitamin B9 (folic acid)-using cellular enzymes, have been used over several decades for the treatment of cancer and inflammatory diseases. Cellular uptake of the antifolates in clinical use occurs primarily via widely expressed facilitative membrane transporters. More recently, human folate receptors (FRs), high affinity receptors that transport folate via endocytosis, have been proposed as targets for the specific delivery of new classes of antifolates or folate conjugates to tumors or sites of inflammation. The development of specific, FR-targeted antifolates would be accelerated if additional biophysical data, particularly structural models of the receptors, were available. Here we describe six distinct crystallographic models that provide insight into biological trafficking of FRs and distinct binding modes of folate and antifolates to these receptors. From comparison of the structures, we delineate discrete structural conformations representative of key stages in the endocytic trafficking of FRs and propose models for pH-dependent conformational changes. Additionally, we describe the molecular details of human FR in complex with three clinically prevalent antifolates, pemetrexed (also Alimta), aminopterin, and methotrexate. On the whole, our data form the basis for rapid design and implementation of unique, FR-targeted, folate-based drugs for the treatment of cancer and inflammatory diseases. PMID:23934049

  12. Problem-Solving Test: Vitellogenin and Vitellogenin-Receptor Recognition: An Example of Protein Interaction

    ERIC Educational Resources Information Center

    Hernandez-Cortes, Patricia

    2012-01-01

    Vitellogenin (Vtg) is a lipid transfer protein that carries yolk to the ovary. The vitellogenin receptor (VtgR) mediates the uptake of Vtg into the oocyte of oviparous animals; its structure includes eight ligand-binding repeats (LBR). The binding site of VtgR and Vtg and the location of the interaction within the molecules are at these LBR.…

  13. Versatility in ligand recognition by LDL receptor family proteins: advances and frontiers.

    PubMed

    Blacklow, Stephen C

    2007-08-01

    Proteins of the low-density lipoprotein receptor family transport cholesterol-carrying particles into cells, clear protease-inhibitor complexes from the circulation, participate in biological signaling cascades, and even serve as viral receptors. These receptors utilize clusters of cysteine-rich LDL receptor type-A (LA) modules to bind many of their ligands. Recent structures show that these modules typically exhibit a characteristic binding mode to recognize their partners, relying primarily on electrostatic complementarity and avidity effects. The dominant contribution of electrostatic interactions with small interface areas in these complexes allows binding to be regulated by changes in pH via at least two distinct mechanisms. The structure of the subtilisin/kexin family protease PCSK9, a newly identified molecular partner of the LDLR also implicated in LDL-cholesterol homeostasis, also raises the possibility that the LDLR and its related family members may employ other strategies for pH-sensitive binding that have yet to be uncovered.

  14. Structures of human folate receptors reveal biological trafficking states and diversity in folate and antifolate recognition.

    PubMed

    Wibowo, Ardian S; Singh, Mirage; Reeder, Kristen M; Carter, Joshua J; Kovach, Alexander R; Meng, Wuyi; Ratnam, Manohar; Zhang, Faming; Dann, Charles E

    2013-09-17

    Antifolates, folate analogs that inhibit vitamin B9 (folic acid)-using cellular enzymes, have been used over several decades for the treatment of cancer and inflammatory diseases. Cellular uptake of the antifolates in clinical use occurs primarily via widely expressed facilitative membrane transporters. More recently, human folate receptors (FRs), high affinity receptors that transport folate via endocytosis, have been proposed as targets for the specific delivery of new classes of antifolates or folate conjugates to tumors or sites of inflammation. The development of specific, FR-targeted antifolates would be accelerated if additional biophysical data, particularly structural models of the receptors, were available. Here we describe six distinct crystallographic models that provide insight into biological trafficking of FRs and distinct binding modes of folate and antifolates to these receptors. From comparison of the structures, we delineate discrete structural conformations representative of key stages in the endocytic trafficking of FRs and propose models for pH-dependent conformational changes. Additionally, we describe the molecular details of human FR in complex with three clinically prevalent antifolates, pemetrexed (also Alimta), aminopterin, and methotrexate. On the whole, our data form the basis for rapid design and implementation of unique, FR-targeted, folate-based drugs for the treatment of cancer and inflammatory diseases.

  15. Viral infection and innate pattern recognition receptors in induction of Hashimoto's thyroiditis.

    PubMed

    Morohoshi, Kazuki; Takahashi, Yurie; Mori, Kouki

    2011-12-01

    Hashimoto's thyroiditis, a common organ-specific autoimmune disease, is multifactorial in which both genetic susceptibility and environmental factors including infection play a critical role in its pathogenesis. Viral infection activates both the innate and adaptive immunity and is implicated as a trigger of Hashimoto's thyroiditis. Candidate viruses include hepatitis C virus and human parvovirus B19. Viral components, which are recognized by innate receptors including Toll-like receptors (TLRs), are detected in thyroid tissues and sera of patients with Hashimoto's thyroiditis. While conflicting results have been obtained regarding the role of TLRs in autoimmune diseases, our preliminary study suggested a contribution of TLR2 and dectin-1 in combination, TLR4, or TLR7 to the production of anti-thyroglobulin antibody in nonobese diabetic mice, a mouse model of Hashimoto's thyroiditis. Despite interesting circumstantial evidence, however, whether viral infection and innate receptors are involved in the development of Hashimoto's thyroiditis remains largely unclear. In this review, we summarize our knowledge regarding the role of viral infection and innate receptors in the etiology of Hashimoto's thyroiditis.

  16. Controlling Cesium Cation Recognition via Cation Metathesis within and Ion Pair Receptor

    SciTech Connect

    Kim, Sung Kuk; Vargas-Zuniga, Gabriela; Hay, Benjamin; Young, Neil J; Delmau, Laetitia Helene; Lee, Prof. Chang-Hee; Kim, Jong Seung; Lynch, Vincent M.; Sessler, Jonathan L.

    2012-01-01

    Ion pair receptor 3 bearing an anion binding site and multiple cation binding sites has been synthesized and shown to function in a novel binding-release cycle that does not necessarily require displacement to effect release. The receptor forms stable complexes with the test cesium salts, CsCl and CsNO{sub 3}, in solution (10% methanol-d{sub 4} in chloroform-d) as inferred from {sup 1}H NMR spectroscopic analyses. The addition of KClO{sub 4} to these cesium salt complexes leads to a novel type of cation metathesis in which the 'exchanged' cations occupy different binding sites. Specifically, K{sup +} becomes bound at the expense of the Cs{sup +} cation initially present in the complex. Under liquid-liquid conditions, receptor 3 is able to extract CsNO{sub 3} and CsCl from an aqueous D{sub 2}O layer into nitrobenzene-d{sub 5} as inferred from {sup 1}H NMR spectroscopic analyses and radiotracer measurements. The Cs{sup +} cation of the CsNO{sub 3} extracted into the nitrobenzene phase by receptor 3 may be released into the aqueous phase by contacting the loaded nitrobenzene phase with an aqueous KClO{sub 4} solution. Additional exposure of the nitrobenzene layer to chloroform and water gives 3 in its uncomplexed, ion-free form. This allows receptor 3 to be recovered for subsequent use. Support for the underlying complexation chemistry came from single-crystal X-ray diffraction analyses and gas-phase energy-minimization studies.

  17. Specificity of Staphyloferrin B recognition by the SirA receptor from Staphylococcus aureus.

    PubMed

    Grigg, Jason C; Cheung, Johnson; Heinrichs, David E; Murphy, Michael E P

    2010-11-05

    Many organisms use sophisticated systems to acquire growth-limiting iron. Iron limitation is especially apparent in bacterial pathogens of mammalian hosts where free iron concentrations are physiologically negligible. A common strategy is to secrete low molecular weight iron chelators, termed siderophores, and express high affinity receptors for the siderophore-iron complex. Staphylococcus aureus, a widespread pathogen, produces two siderophores, staphyloferrin A (SA) and staphyloferrin B (SB). We have determined the crystal structure of the staphyloferrin B receptor, SirA, at high resolution in both the apo and Fe(III)-SB (FeSB)-bound forms. SirA, a member of the class III binding protein family of metal receptors, has N- and C-terminal domains, each composed of mainly a β-stranded core and α-helical periphery. The domains are bridged by a single α-helix and together form the FeSB binding site. SB coordinates Fe(III) through five oxygen atoms and one nitrogen atom in distorted octahedral geometry. SirA undergoes conformational change upon siderophore binding, largely securing two loops from the C-terminal domain to enclose FeSB with a low nanomolar dissociation constant. The staphyloferrin A receptor, HtsA, homologous to SirA, also encloses its cognate siderophore (FeSA); however, the largest conformational rearrangements involve a different region of the C-terminal domain. FeSB is uniquely situated in the binding pocket of SirA with few of the contacting residues being conserved with those of HtsA interacting with FeSA. Although both SirA and HtsA bind siderophores from the same α-hydroxycarboxylate class, the unique structural features of each receptor provides an explanation for their distinct specificity.

  18. Transgenic Expression of the Dicotyledonous Pattern Recognition Receptor EFR in Rice Leads to Ligand-Dependent Activation of Defense Responses

    PubMed Central

    Thomas, Nicolas; Holton, Nicolas; Nekrasov, Vladimir; Ruan, Deling; Canlas, Patrick E.; Daudi, Arsalan; Petzold, Christopher J.; Singan, Vasanth R.; Kuo, Rita; Chovatia, Mansi; Daum, Christopher; Heazlewood, Joshua L.; Zipfel, Cyril; Ronald, Pamela C.

    2015-01-01

    Plant plasma membrane localized pattern recognition receptors (PRRs) detect extracellular pathogen-associated molecules. PRRs such as Arabidopsis EFR and rice XA21 are taxonomically restricted and are absent from most plant genomes. Here we show that rice plants expressing EFR or the chimeric receptor EFR::XA21, containing the EFR ectodomain and the XA21 intracellular domain, sense both Escherichia coli- and Xanthomonas oryzae pv. oryzae (Xoo)-derived elf18 peptides at sub-nanomolar concentrations. Treatment of EFR and EFR::XA21 rice leaf tissue with elf18 leads to MAP kinase activation, reactive oxygen production and defense gene expression. Although expression of EFR does not lead to robust enhanced resistance to fully virulent Xoo isolates, it does lead to quantitatively enhanced resistance to weakly virulent Xoo isolates. EFR interacts with OsSERK2 and the XA21 binding protein 24 (XB24), two key components of the rice XA21-mediated immune response. Rice-EFR plants silenced for OsSERK2, or overexpressing rice XB24 are compromised in elf18-induced reactive oxygen production and defense gene expression indicating that these proteins are also important for EFR-mediated signaling in transgenic rice. Taken together, our results demonstrate the potential feasibility of enhancing disease resistance in rice and possibly other monocotyledonous crop species by expression of dicotyledonous PRRs. Our results also suggest that Arabidopsis EFR utilizes at least a subset of the known endogenous rice XA21 signaling components. PMID:25821973

  19. Transgenic expression of the dicotyledonous pattern recognition receptor EFR in rice leads to ligand-dependent activation of defense responses

    DOE PAGES

    Schwessinger, Benjamin; Bahar, Ofir; Thomas, Nicolas; ...

    2015-03-30

    Plant plasma membrane localized pattern recognition receptors (PRRs) detect extracellular pathogen-associated molecules. PRRs such as Arabidopsis EFR and rice XA21 are taxonomically restricted and are absent from most plant genomes. Here we show that rice plants expressing EFR or the chimeric receptor EFR::XA21, containing the EFR ectodomain and the XA21 intracellular domain, sense both Escherichia coli- and Xanthomonas oryzae pv. oryzae (Xoo)-derived elf18 peptides at sub-nanomolar concentrations. Treatment of EFR and EFR::XA21 rice leaf tissue with elf18 leads to MAP kinase activation, reactive oxygen production and defense gene expression. Although expression of EFR does not lead to robust enhanced resistancemore » to fully virulent Xoo isolates, it does lead to quantitatively enhanced resistance to weakly virulent Xoo isolates. EFR interacts with OsSERK2 and the XA21 binding protein 24 (XB24), two key components of the rice XA21-mediated immune response. Rice-EFR plants silenced for OsSERK2, or overexpressing rice XB24 are compromised in elf18-induced reactive oxygen production and defense gene expression indicating that these proteins are also important for EFR-mediated signaling in transgenic rice. Taken together, our results demonstrate the potential feasibility of enhancing disease resistance in rice and possibly other monocotyledonous crop species by expression of dicotyledonous PRRs. Our results also suggest that Arabidopsis EFR utilizes at least a subset of the known endogenous rice XA21 signaling components.« less

  20. Modeling ligand recognition at the P2Y12 receptor in light of X-ray structural information

    NASA Astrophysics Data System (ADS)

    Paoletta, Silvia; Sabbadin, Davide; von Kügelgen, Ivar; Hinz, Sonja; Katritch, Vsevolod; Hoffmann, Kristina; Abdelrahman, Aliaa; Straßburger, Jens; Baqi, Younis; Zhao, Qiang; Stevens, Raymond C.; Moro, Stefano; Müller, Christa E.; Jacobson, Kenneth A.

    2015-08-01

    The G protein-coupled P2Y12 receptor (P2Y12R) is an important antithrombotic target and of great interest for pharmaceutical discovery. Its recently solved, highly divergent crystallographic structures in complex either with nucleotides (full or partial agonist) or with a nonnucleotide antagonist raise the question of which structure is more useful to understand ligand recognition. Therefore, we performed extensive molecular modeling studies based on these structures and mutagenesis, to predict the binding modes of major classes of P2Y12R ligands previously reported. Various nucleotide derivatives docked readily to the agonist-bound P2Y12R, but uncharged nucleotide-like antagonist ticagrelor required a hybrid receptor resembling the agonist-bound P2Y12R except for the top portion of TM6. Supervised molecular dynamics (SuMD) of ticagrelor binding indicated interactions with the extracellular regions of P2Y12R, defining possible meta-binding sites. Ureas, sulfonylureas, sulfonamides, anthraquinones and glutamic acid piperazines docked readily to the antagonist-bound P2Y12R. Docking dinucleotides at both agonist- and antagonist-bound structures suggested interactions with two P2Y12R pockets. Thus, our structure-based approach consistently rationalized the main structure-activity relationships within each ligand class, giving useful information for designing improved ligands.

  1. Structural basis for chemokine recognition and activation of a viral G protein-coupled receptor

    SciTech Connect

    Burg, John S.; Ingram, Jessica R.; Venkatakrishnan, A.J.; Jude, Kevin M.; Dukkipati, Abhiram; Feinberg, Evan N.; Angelini, Alessandro; Waghray, Deepa; Dror, Ron O.; Ploegh, Hidde L.; Garcia, K. Christopher

    2015-03-05

    Chemokines are small proteins that function as immune modulators through activation of chemokine G protein-coupled receptors (GPCRs). Several viruses also encode chemokines and chemokine receptors to subvert the host immune response. How protein ligands activate GPCRs remains unknown. We report the crystal structure at 2.9 angstrom resolution of the human cytomegalovirus GPCR US28 in complex with the chemokine domain of human CX3CL1 (fractalkine). The globular body of CX3CL1 is perched on top of the US28 extracellular vestibule, whereas its amino terminus projects into the central core of US28. The transmembrane helices of US28 adopt an active-state-like conformation. Atomic-level simulations suggest that the agonist-independent activity of US28 may be due to an amino acid network evolved in the viral GPCR to destabilize the receptor’s inactive state.

  2. Recognition of SUMO-modified PCNA requires tandem receptor motifs in Srs2

    SciTech Connect

    Armstrong, Anthony A.; Mohideen, Firaz; Lima, Christopher D.

    2013-04-08

    Ubiquitin (Ub) and ubiquitin-like (Ubl) modifiers such as SUMO (also known as Smt3 in Saccharomyces cerevisiae) mediate signal transduction through post-translational modification of substrate proteins in pathways that control differentiation, apoptosis and the cell cycle, and responses to stress such as the DNA damage response. In yeast, the proliferating cell nuclear antigen PCNA (also known as Pol30) is modified by ubiquitin in response to DNA damage and by SUMO during S phase. Whereas Ub-PCNA can signal for recruitment of translesion DNA polymerases, SUMO-PCNA signals for recruitment of the anti-recombinogenic DNA helicase Srs2. It remains unclear how receptors such as Srs2 specifically recognize substrates after conjugation to Ub and Ubls. Here we show, through structural, biochemical and functional studies, that the Srs2 carboxy-terminal domain harbors tandem receptor motifs that interact independently with PCNA and SUMO and that both motifs are required to recognize SUMO-PCNA specifically. The mechanism presented is pertinent to understanding how other receptors specifically recognize Ub- and Ubl-modified substrates to facilitate signal transduction.

  3. Synthetic Receptors for the High‐Affinity Recognition of O‐GlcNAc Derivatives

    PubMed Central

    Rios, Pablo; Carter, Tom S.; Crump, Matthew P.; Lisbjerg, Micke; Pittelkow, Michael; Supekar, Nitin T.; Boons, Geert‐Jan

    2016-01-01

    Abstract The combination of a pyrenyl tetraamine with an isophthaloyl spacer has led to two new water‐soluble carbohydrate receptors (“synthetic lectins”). Both systems show outstanding affinities for derivatives of N‐acetylglucosamine (GlcNAc) in aqueous solution. One receptor binds the methyl glycoside GlcNAc‐β‐OMe with K a≈20 000 m −1, whereas the other one binds an O‐GlcNAcylated peptide with K a≈70 000 m −1. These values substantially exceed those usually measured for GlcNAc‐binding lectins. Slow exchange on the NMR timescale enabled structural determinations for several complexes. As expected, the carbohydrate units are sandwiched between the pyrenes, with the alkoxy and NHAc groups emerging at the sides. The high affinity of the GlcNAcyl–peptide complex can be explained by extra‐cavity interactions, raising the possibility of a family of complementary receptors for O‐GlcNAc in different contexts. PMID:26822115

  4. Single cell molecular recognition of migrating and invading tumor cells using a targeted fluorescent probe to receptor PTPmu.

    PubMed

    Burden-Gulley, Susan M; Qutaish, Mohammed Q; Sullivant, Kristin E; Tan, Mingqian; Craig, Sonya E L; Basilion, James P; Lu, Zheng-Rong; Wilson, David L; Brady-Kalnay, Susann M

    2013-04-01

    Detection of an extracellular cleaved fragment of a cell-cell adhesion molecule represents a new paradigm in molecular recognition and imaging of tumors. We previously demonstrated that probes that recognize the cleaved extracellular domain of receptor protein tyrosine phosphatase mu (PTPmu) label human glioblastoma brain tumor sections and the main tumor mass of intracranial xenograft gliomas. In this article, we examine whether one of these probes, SBK2, can label dispersed glioma cells that are no longer connected to the main tumor mass. Live mice with highly dispersive glioma tumors were injected intravenously with the fluorescent PTPmu probe to test the ability of the probe to label the dispersive glioma cells in vivo. Analysis was performed using a unique three-dimensional (3D) cryo-imaging technique to reveal highly migratory and invasive glioma cell dispersal within the brain and the extent of colabeling by the PTPmu probe. The PTPmu probe labeled the main tumor site and dispersed cells up to 3.5 mm away. The cryo-images of tumors labeled with the PTPmu probe provide a novel, high-resolution view of molecular tumor recognition, with excellent 3D detail regarding the pathways of tumor cell migration. Our data demonstrate that the PTPmu probe recognizes distant tumor cells even in parts of the brain where the blood-brain barrier is likely intact. The PTPmu probe has potential translational significance for recognizing tumor cells to facilitate molecular imaging, a more complete tumor resection and to serve as a molecular targeting agent to deliver chemotherapeutics to the main tumor mass and distant dispersive tumor cells.

  5. Positive allosteric modulation of alpha 7 nicotinic acetylcholine receptors enhances recognition memory and cognitive flexibility in rats.

    PubMed

    Nikiforuk, Agnieszka; Kos, Tomasz; Potasiewicz, Agnieszka; Popik, Piotr

    2015-08-01

    A wide body of preclinical and clinical data suggests that alpha 7 nicotinic acetylcholine receptors (α7-nAChRs) may represent useful targets for cognitive improvement in schizophrenia and Alzheimer׳s disease. A promising recent approach is based on the use of positive allosteric modulators (PAMs) of α7-nAChRs due to their several advantages over the direct agonists. Nevertheless, the behavioural effects of this class of compounds, particularly with regard to higher-order cognitive functions, have not been broadly characterised. The aim of the present study was to evaluate the procognitive efficacies of type I and type II α7-nAChRs PAMs, N-(4-chlorophenyl)-[[(4-chlorophenyl)amino]methylene]-3-methyl-5-isoxazoleacet-amide (CCMI) and N-(5-Chloro-2,4-dimethoxyphenyl)-N'-(5-methyl-3-isoxazolyl)urea (PNU-120596) in the novel object recognition task (NORT), attentional set-shifting task (ASST) and five-choice serial reaction time task (5-CSRTT) in rats. Additionally, the effects of galantamine, an acetylcholinesterase inhibitor that also allosterically modulates nAChRs, were assessed. We report that CCMI (0.3-3mg/kg), PNU-120596 (0.3-3mg/kg) and galantamine (1-3mg/kg) attenuated the delay-induced impairment in NORT performance and facilitated cognitive flexibility in the ASST. Methyllycaconitine (3mg/kg) blocked the actions of CCMI, PNU-120596 and galantamine in the NORT and ASST, suggesting that the procognitive effects of these compounds are α7-nAChRs-dependent. However, none of the compounds tested affected the rats' attentional performance in the 5-CSRTT. The present findings confirm and extend the observations indicating that the positive allosteric modulation of α7-nAChRs enhances recognition memory and cognitive flexibility in preclinical tasks. Therefore, the present study supports the utility of α7-nAChRs PAMs as a potential cognitive enhancing therapy.

  6. Cellular Recognition and Trafficking of Amorphous Silica Nanoparticles by Macrophage Scavenger Receptor A

    SciTech Connect

    Orr, Galya; Chrisler, William B.; Cassens, Kaylyn J.; Tan, Ruimin; Tarasevich, Barbara J.; Markillie, Lye Meng; Zangar, Richard C.; Thrall, Brian D.

    2011-09-01

    The internalization of engineered nanoparticles (ENPs) into cells is known to involve active transport mechanisms, yet the precise biological molecules involved are poorly understood. We demonstrate that the uptake of amorphous silica ENPs (92 nm) by macrophage cells is strongly inhibited by silencing expression of scavenger receptor A (SR-A). In addition, ENP uptake is augmented by introducing SR-A expression into human cells that are normally non-phagocytic. Confocal fluorescent microscopy analyses show that the majority of single or small clusters of silica ENPs co-localize intracellularly with SR-A and are internalized through a pathway characteristic of clathrin-dependent endocytosis. In contrast, larger silica NP agglomerates (>500 nm) are poorly co-localized with the receptor, suggesting independent trafficking or internalization pathways are involved. SR-A silencing also caused decreased cellular secretion of pro-inflammatory cytokines in response to silica ENPs. As SR-A is expressed in macrophages throughout the reticulo-endothelial system, this pathway is likely an important determinant of the biodistribution of, and cellular response to ENPs.

  7. Cross-neutralizing human anti-poliovirus antibodies bind the recognition site for cellular receptor

    PubMed Central

    Chen, Zhaochun; Fischer, Elizabeth R.; Kouiavskaia, Diana; Hansen, Bryan T.; Ludtke, Steven J.; Bidzhieva, Bella; Makiya, Michelle; Agulto, Liane; Purcell, Robert H.; Chumakov, Konstantin

    2013-01-01

    Most structural information about poliovirus interaction with neutralizing antibodies was obtained in the 1980s in studies of mouse monoclonal antibodies. Recently we have isolated a number of human/chimpanzee anti-poliovirus antibodies and demonstrated that one of them, MAb A12, could neutralize polioviruses of both serotypes 1 and 2. This communication presents data on isolation of an additional cross-neutralizing antibody (F12) and identification of a previously unknown epitope on the surface of poliovirus virions. Epitope mapping was performed by sequencing of antibody-resistant mutants and by cryo-EM of complexes of virions with Fab fragments. The results have demonstrated that both cross-neutralizing antibodies bind the site located at the bottom of the canyon surrounding the fivefold axis of symmetry that was previously shown to interact with cellular poliovirus receptor CD155. However, the same antibody binds to serotypes 1 and 2 through different specific interactions. It was also shown to interact with type 3 poliovirus, albeit with about 10-fold lower affinity, insufficient for effective neutralization. Antibody interaction with the binding site of the cellular receptor may explain its broad reactivity and suggest that further screening or antibody engineering could lead to a universal antibody capable of neutralizing all three serotypes of poliovirus. PMID:24277851

  8. Structural basis for receptor recognition by New World hemorrhagic fever arenaviruses

    SciTech Connect

    Abraham, Jonathan; Corbett, Kevin D.; Farzan, Michael; Choe, Hyeryun; Harrison, Stephen C.

    2010-08-18

    New World hemorrhagic fever arenaviruses are rodent-borne agents that cause severe human disease. The GP1 subunit of the surface glycoprotein mediates cell attachment through transferrin receptor 1 (TfR1). We report the structure of Machupo virus (MACV) GP1 bound with human TfR1. Atomic details of the GP1-TfR1 interface clarify the importance of TfR1 residues implicated in New World arenavirus host specificity. Analysis of sequence variation among New World arenavirus GP1s and their host-species receptors, in light of the molecular structure, indicates determinants of viral zoonotic transmission. Infectivities of pseudoviruses in cells expressing mutated TfR1 confirm that contacts at the tip of the TfR1 apical domain determine the capacity of human TfR1 to mediate infection by particular New World arenaviruses. We propose that New World arenaviruses that are pathogenic to humans fortuitously acquired affinity for human TfR1 during adaptation to TfR1 of their natural hosts.

  9. Phosphatidylserine recognition and induction of apoptotic cell clearance by Drosophila engulfment receptor Draper.

    PubMed

    Tung, Tran Thanh; Nagaosa, Kaz; Fujita, Yu; Kita, Asana; Mori, Hiroki; Okada, Ryo; Nonaka, Saori; Nakanishi, Yoshinobu

    2013-05-01

    The membrane phospholipid phosphatidylserine is exposed on the cell surface during apoptosis and acts as an eat-me signal in the phagocytosis of apoptotic cells in mammals and nematodes. However, whether this is also true in insects was unclear. When milk fat globule-epidermal growth factor 8, a phosphatidylserine-binding protein of mammals, was ectopically expressed in Drosophila, the level of phagocytosis was reduced, whereas this was not the case for the same protein lacking a domain responsible for the binding to phosphatidylserine. We found that the extracellular region of Draper, an engulfment receptor of Drosophila, binds to phosphatidylserine in an enzyme-linked immunosorbent assay-like solid-phase assay and in an assay for surface plasmon resonance. A portion of Draper containing domains EMI and NIM located close to the N-terminus was required for binding to phosphatidylserine, and a Draper protein lacking this region was not active in Drosophila. Finally, the level of tyrosine-phosphorylated Draper, indicative of the activation of Draper, in a hemocyte-derived cell line was increased after treatment with phosphatidylserine-containing liposome. These results indicated that phosphatidylserine serves as an eat-me signal in the phagocytic removal of apoptotic cells in Drosophila and that Draper is a phosphatidylserine-binding receptor for phagocytosis.

  10. Structural Basis for Ligand Recognition and Functional Selectivity at Angiotensin Receptor*♦

    PubMed Central

    Zhang, Haitao; Unal, Hamiyet; Desnoyer, Russell; Han, Gye Won; Patel, Nilkanth; Katritch, Vsevolod; Karnik, Sadashiva S.; Cherezov, Vadim; Stevens, Raymond C.

    2015-01-01

    Angiotensin II type 1 receptor (AT1R) is the primary blood pressure regulator. AT1R blockers (ARBs) have been widely used in clinical settings as anti-hypertensive drugs and share a similar chemical scaffold, although even minor variations can lead to distinct therapeutic efficacies toward cardiovascular etiologies. The structural basis for AT1R modulation by different peptide and non-peptide ligands has remained elusive. Here, we report the crystal structure of the human AT1R in complex with an inverse agonist olmesartan (BenicarTM), a highly potent anti-hypertensive drug. Olmesartan is anchored to the receptor primarily by the residues Tyr-351.39, Trp-842.60, and Arg-167ECL2, similar to the antagonist ZD7155, corroborating a common binding mode of different ARBs. Using docking simulations and site-directed mutagenesis, we identified specific interactions between AT1R and different ARBs, including olmesartan derivatives with inverse agonist, neutral antagonist, or agonist activities. We further observed that the mutation N1113.35A in the putative sodium-binding site affects binding of the endogenous peptide agonist angiotensin II but not the β-arrestin-biased peptide TRV120027. PMID:26420482

  11. Binding of thyroglobulin to bovine thyroid membranes. Role of specific amino acids in receptor recognition.

    PubMed

    Shifrin, S; Kohn, L D

    1981-10-25

    Bovine thyroglobulin was treated with increasing ratios of succinic anhydride, trinitrobenzene sulfonic acid, tetranitromethane, and N-acetylimidazole in an attempt to assess the role of lysine or tyrosine residues in binding to thyroid membrane receptors. Extensive succinylation results in dissociation to 12 S thyroglobulin with retention of a considerable portion of the three-dimensional structure. Only 25% of the lysine residues can be modified by trinitrophenylation without affecting inter-subunit interactions. Succinylation as well as trinitrophenylation increases the affinity of thyroglobulin for the membrane receptor by a factor of 2. The binding of thyroglobulin to the membrane was reduced after nitration of 30% of the tyrosyl residues with tetranitromethane. O-Acetylation of 40-70% of the tyrosyl residues by N-acetylimidazole nearly abolished the ability of thyroglobulin to bind to the membrane. Removal of the O-acetyl group with hydroxylamine restored the binding properties. The results indicate that tyrosyl residues play an important role in thyroglobulin interactions with thyroid membranes.

  12. Receptor recognition of transferrin bound to lanthanides and actinides: a discriminating step in cellular acquisition of f-block metals

    PubMed Central

    Deblonde, Gauthier J.-P.; Sturzbecher-Hoehne, Manuel; Mason, Anne B.; Abergel, Rebecca J.

    2013-01-01

    Following an internal contamination event, the transport of actinide and lanthanide metal ions through the body is facilitated by endogenous ligands such as the human iron-transport protein transferrin (Tf). The recognition of resulting metallo-transferrin complexes (M2Tf) by the cognate transferrin receptor (TfR) is therefore a critical step for cellular uptake of these metal ions. A high performance liquid chromatography-based method has been used to probe the binding of M2Tf with TfR, yielding a direct measurement of the successive thermodynamic constants that correspond to the dissociation of TfR(M2Tf)2 and TfR(M2Tf) complexes for Fe3+, Ga3+, La3+, Nd3+, Gd3+, Yb3+, Lu3+, 232Th4+, 238UO22+, and 242Pu4+. Important features of this method are (i) its ability to distinguish both 1:1 and 1:2 complexes formed between the receptor and the metal-bound transferrin, and (ii) the requirement for very small amounts of each binding partner (<1 nmol of protein per assay). Consistent with previous reports, the strongest receptor affinity is found for Fe2Tf (Kd1 = 5 nM and Kd2 = 20 nM), while the lowest affinity was measured for Pu2Tf (Kd1 = 0.28 µM and Kd2 = 1.8 µM) binding to the TfR. Other toxic metal ions such as ThIV and UVI, when bound to Tf, are well recognized by the TfR. Under the described experimental conditions, the relative stabilities of TfR:(MxTf)y adducts follow the order Fe3+ >> Th4+ □ UO22+ □ Cm3+ > Ln3+ □ Ga3+ >>> Yb3+ □ Pu4+. This study substantiates a role for Tf in binding lanthanide fission products and actinides, and transporting them into cells by receptor mediated endocytosis. PMID:23446908

  13. Receptor recognition of transferrin bound to lanthanides and actinides: a discriminating step in cellular acquisition of f-block metals.

    PubMed

    Deblonde, Gauthier J-P; Sturzbecher-Hoehne, Manuel; Mason, Anne B; Abergel, Rebecca J

    2013-06-01

    Following an internal contamination event, the transport of actinide (An) and lanthanide (Ln) metal ions through the body is facilitated by endogenous ligands such as the human iron-transport protein transferrin (Tf). The recognition of resulting metallo-transferrin complexes (M2Tf) by the cognate transferrin receptor (TfR) is therefore a critical step for cellular uptake of these metal ions. A high performance liquid chromatography-based method has been used to probe the binding of M2Tf with TfR, yielding a direct measurement of the successive thermodynamic constants that correspond to the dissociation of TfR(M2Tf)2 and TfR(M2Tf) complexes for Fe(3+), Ga(3+), La(3+), Nd(3+), Gd(3+), Yb(3+), Lu(3+), (232)Th(4+), (238)UO2(2+), and (242)Pu(4+). Important features of this method are (i) its ability to distinguish both 1 : 1 and 1 : 2 complexes formed between the receptor and the metal-bound transferrin, and (ii) the requirement for very small amounts of each binding partner (<1 nmol of protein per assay). Consistent with previous reports, the strongest receptor affinity is found for Fe2Tf (Kd1 = 5 nM and Kd2 = 20 nM), while the lowest affinity was measured for Pu2Tf (Kd1 = 0.28 μM and Kd2 = 1.8 μM) binding to the TfR. Other toxic metal ions such as Th(IV) and U(VI), when bound to Tf, are well recognized by the TfR. Under the described experimental conditions, the relative stabilities of TfR:(MxTf)y adducts follow the order Fe(3+) > Th(4+) ~ UO2(2+) ~ Cm(3+) > Ln(3+) ~ Ga(3+) > Yb(3+) ~ Pu(4+). This study substantiates a role for Tf in binding lanthanide fission products and actinides, and transporting them into cells by receptor-mediated endocytosis.

  14. Quorum sensing in Chromobacterium violaceum: DNA recognition and gene regulation by the CviR receptor.

    PubMed

    Stauff, Devin L; Bassler, Bonnie L

    2011-08-01

    The bacterial pathogen Chromobacterium violaceum uses a LuxIR-type quorum-sensing system to detect and respond to changes in cell population density. CviI synthesizes the autoinducer C(10)-homoserine lactone (C(10)-HSL), and CviR is a cytoplasmic DNA binding transcription factor that activates gene expression following binding to C(10)-HSL. A number of behaviors are controlled by quorum sensing in C. violaceum. However, few genes have been shown to be directly controlled by CviR, in part because the DNA motif bound by CviR is not well characterized. Here, we define the DNA sequence required for promoter recognition by CviR. Using in vivo data generated from a library of point mutations in a CviR-regulated promoter, we find that CviR binds to a palindrome with the ideal sequence CTGNCCNNNNGGNCAG. We constructed a position weight matrix using these in vivo data and scanned the C. violaceum genome to predict CviR binding sites. We measured direct activation of the identified promoters by CviR and found that CviR controls the expression of the promoter for a chitinase, a type VI secretion-related gene, a transcriptional regulator gene, a guanine deaminase gene, and cviI. Indeed, regulation of cviI expression by CviR generates a canonical quorum-sensing positive-feedback loop.

  15. The Mannose Receptor (CD206) is an important pattern recognition receptor (PRR) in the detection of the infective stage of the helminth Schistosoma mansoni and modulates IFNγ production.

    PubMed

    Paveley, Ross A; Aynsley, Sarah A; Turner, Joseph D; Bourke, Claire D; Jenkins, Stephen J; Cook, Peter C; Martinez-Pomares, Luisa; Mountford, Adrian P

    2011-11-01

    In this study, infective larvae of the parasitic helminth Schistosoma mansoni were shown to contain a large number of glycosylated components specific for the Mannose Receptor (MR; CD206), which is an important pattern recognition receptor (PRR) of the innate immune system. MR ligands were particularly rich in excretory/secretory (E/S) material released during transformation of cercariae into schistosomula, a process critical for infection of the host. E/S material from carboxyfluorescein diacetate succinimidyl ester (CFDA-SE)-labelled cercariae showed enhanced binding by cells lines that over-express the MR. Conversely, uptake was significantly lower by bone marrow-derived macrophages (MΦ) from MR(-/-) mice, although they were more active as judged by enhanced pro-inflammatory cytokine production and CD40 expression. After natural percutaneous infection of MR(-/-) mice with CFDA-SE-labelled parasites, there were fewer cells in the skin and draining lymph nodes that were CFDA-SE(+) compared with wild-type mice, implying reduced uptake and presentation of larval parasite antigen. However, antigen-specific proliferation of skin draining lymph node cells was significantly enhanced and they secreted markedly elevated levels of IFNγ but decreased levels of IL-4. In conclusion, we show that the MR on mononuclear phagocytic cells, which are plentiful in the skin, plays a significant role in internalising E/S material released by the invasive stages of the parasite which in turn modulates their production of pro-inflammatory cytokines. In the absence of the MR, antigen-specific CD4(+) cells are Th1 biased, suggesting that ligation of the MR by glycosylated E/S material released by schistosome larvae modulates the production of CD4(+) cell specific IFNγ.

  16. Cancer cell-selective promoter recognition accompanies antitumor effect by glucocorticoid receptor-targeted gold nanoparticle

    NASA Astrophysics Data System (ADS)

    Sau, Samaresh; Agarwalla, Pritha; Mukherjee, Sudip; Bag, Indira; Sreedhar, Bojja; Pal-Bhadra, Manika; Patra, Chitta Ranjan; Banerjee, Rajkumar

    2014-05-01

    Nanoparticles, such as gold nanoparticles (GNP), upon convenient modifications perform multi tasks catering to many biomedical applications. However, GNP or any other type of nanoparticles is yet to achieve the feat of intracellular regulation of endogenous genes of choice such as through manipulation of a gene-promoter in a chromosome. As for gene modulation and delivery, GNP (or other nanoparticles) showed only limited gene therapy potential, which relied on the delivery of `exogenous' genes invoking gene knockdown or replacement. Practically, there are no instances for the nanoparticle-mediated promoter regulation of `endogenous' genes, more so, as a cancer selective phenomenon. In this regard, we report the development of a simple, easily modifiable GNP-formulation, which promoted/up-regulated the expression of a specific category of `endogenous' genes, the glucocorticoid responsive genes. This genetic up-regulation was induced in only cancer cells by modified GNP-mediated transcriptional activation of its cytoplasmic receptor, glucocorticoid receptor (GR). Normal cells and their GR remained primarily unperturbed by this GNP-formulation. The most potent gene up-regulating GNP-formulation down-regulated a cancer-specific proliferative signal, phospho-Akt in cancer cells, which accompanied retardation of tumor growth in the murine melanoma model. We show that GR-targeted GNPs may find potential use in the targeting and modulation of genetic information in cancer towards developing novel anticancer therapeutics.Nanoparticles, such as gold nanoparticles (GNP), upon convenient modifications perform multi tasks catering to many biomedical applications. However, GNP or any other type of nanoparticles is yet to achieve the feat of intracellular regulation of endogenous genes of choice such as through manipulation of a gene-promoter in a chromosome. As for gene modulation and delivery, GNP (or other nanoparticles) showed only limited gene therapy potential, which relied

  17. Computational evaluation of unsaturated carbonitriles as neutral receptor model for beryllium(II) recognition.

    PubMed

    Rosli, Ahmad Nazmi; Ahmad, Mohd Rais; Alias, Yatimah; Zain, Sharifuddin Md; Lee, Vannajan Sanghiran; Woi, Pei Meng

    2014-12-01

    Design of neutral receptor molecules (ionophores) for beryllium(II) using unsaturated carbonitrile models has been carried out via density functional theory, G3, and G4 calculations. The first part of this work focuses on gas phase binding energies between beryllium(II) and 2-cyano butadiene (2-CN BD), 3-cyano propene (3-CN P), and simpler models with two separate fragments; acrylonitrile and ethylene. Interactions between beryllium(II) and cyano nitrogen and terminal olefin in the models have been examined in terms of geometrical changes, distribution of charge over the entire π-system, and rehybridization of vinyl carbon orbitals. NMR shieldings and vibrational frequencies probed charge centers and strength of interactions. The six-membered cyclic complexes have planar structures with the rehybridized carbon slightly out of plane (16° in 2-CN BD). G3 results show that in 2-CN BD complex participation of vinyl carbon further stabilizes the cyclic adduct by 16.3 kcal mol(-1), whereas, in simpler models, interaction between beryllium(II) and acetonitrile is favorable by 46.4 kcal mol(-1) compared with that of ethylene. The terminal vinyl carbon in 2-CN BD rehybridizes to sp (3) with an increase of 7 % of s character to allow interaction with beryllium(II). G4 calculations show that the Be(II) and 2-CN BD complex is more strongly bound than those with Mg(II) and Ca(II) by 98.5 and 139.2 kcal mol(-1) (-1), respectively. QST2 method shows that the cyclic and acyclic forms of Be(II)-2-CN BD complexes are separated by 12.3 kcal mol(-1) barrier height. Overlap population analysis reveals that Ca(II) can be discriminated based on its tendency to form ionic interaction with the receptor models.

  18. Functional Roles of Pattern Recognition Receptors That Recognize Virus Nucleic Acids in Human Adipose-Derived Mesenchymal Stem Cells

    PubMed Central

    Wang, Fangchao; Yang, Can; Liu, Guoyan; Song, Xiangfeng

    2016-01-01

    Human adipose-derived mesenchymal stem cells (hAD-MSCs) are mesenchymal stem cells with the capability to modulate immune responses. Evidence showing that hAD-MSCs could mediate innate immune responses through pattern recognition receptors (PRRs) is increasing. However, the roles of PRRs in regulating the innate sensing of virus nucleic acids (RNA and DNA) in hAD-MSCs have not yet been investigated. This study focused on the abundant expression of PRRs, including Toll-like receptor 3 (TLR3) and retinoic acid-inducible gene I (RIG-I), which recognize viral RNA, and gamma-interferon inducible protein 16 (IFI16), which recognizes viral DNA in hAD-MSCs. Poly(I:C), a synthetic dsRNA analogy, activated TLR3 and RIG-I and induced the expression of type I interferons (IFN-α/β) and antivirus proteins, including IFN-stimulating gene 15, 2′5′-oligoadenylate synthetase, and Mx GTPase 1 in hAD-MSCs, which were attenuated by the knockdown of each TLR3 or RIG-I. Synthetic herpes simplex viral DNA (HSV60) activated IFI16 and induced the expression of IFN-α/β and antivirus proteins in hAD-MSCs, which were inhibited by the knockdown of IFI16. Both poly(I:C) and HSV60 induced the expression of IFN-α/β through the phosphorylation of IFN-regulatory factor 3. All these results indicated that PRRs recognizing virus nucleic acids were expressed and can mediate antivirus responses in hAD-MSCs. PMID:28105439

  19. Functional Roles of Pattern Recognition Receptors That Recognize Virus Nucleic Acids in Human Adipose-Derived Mesenchymal Stem Cells.

    PubMed

    Yu, Lili; Xu, Yongtao; Wang, Fangchao; Yang, Can; Liu, Guoyan; Song, Xiangfeng

    2016-01-01

    Human adipose-derived mesenchymal stem cells (hAD-MSCs) are mesenchymal stem cells with the capability to modulate immune responses. Evidence showing that hAD-MSCs could mediate innate immune responses through pattern recognition receptors (PRRs) is increasing. However, the roles of PRRs in regulating the innate sensing of virus nucleic acids (RNA and DNA) in hAD-MSCs have not yet been investigated. This study focused on the abundant expression of PRRs, including Toll-like receptor 3 (TLR3) and retinoic acid-inducible gene I (RIG-I), which recognize viral RNA, and gamma-interferon inducible protein 16 (IFI16), which recognizes viral DNA in hAD-MSCs. Poly(I:C), a synthetic dsRNA analogy, activated TLR3 and RIG-I and induced the expression of type I interferons (IFN-α/β) and antivirus proteins, including IFN-stimulating gene 15, 2'5'-oligoadenylate synthetase, and Mx GTPase 1 in hAD-MSCs, which were attenuated by the knockdown of each TLR3 or RIG-I. Synthetic herpes simplex viral DNA (HSV60) activated IFI16 and induced the expression of IFN-α/β and antivirus proteins in hAD-MSCs, which were inhibited by the knockdown of IFI16. Both poly(I:C) and HSV60 induced the expression of IFN-α/β through the phosphorylation of IFN-regulatory factor 3. All these results indicated that PRRs recognizing virus nucleic acids were expressed and can mediate antivirus responses in hAD-MSCs.

  20. Transgenic expression of the dicotyledonous pattern recognition receptor EFR in rice leads to ligand-dependent activation of defense responses

    SciTech Connect

    Schwessinger, Benjamin; Bahar, Ofir; Thomas, Nicolas; Holton, Nicolas; Nekrasov, Vladimir; Ruan, Deling; Canlas, Patrick E.; Daudi, Arsalan; Petzold, Christopher J.; Singan, Vasanth R.; Kuo, Rita; Chovatia, Mansi; Daum, Christopher; Heazlewood, Joshua L.; Zipfel, Cyril; Ronald, Pamela C.

    2015-03-30

    Plant plasma membrane localized pattern recognition receptors (PRRs) detect extracellular pathogen-associated molecules. PRRs such as Arabidopsis EFR and rice XA21 are taxonomically restricted and are absent from most plant genomes. Here we show that rice plants expressing EFR or the chimeric receptor EFR::XA21, containing the EFR ectodomain and the XA21 intracellular domain, sense both Escherichia coli- and Xanthomonas oryzae pv. oryzae (Xoo)-derived elf18 peptides at sub-nanomolar concentrations. Treatment of EFR and EFR::XA21 rice leaf tissue with elf18 leads to MAP kinase activation, reactive oxygen production and defense gene expression. Although expression of EFR does not lead to robust enhanced resistance to fully virulent Xoo isolates, it does lead to quantitatively enhanced resistance to weakly virulent Xoo isolates. EFR interacts with OsSERK2 and the XA21 binding protein 24 (XB24), two key components of the rice XA21-mediated immune response. Rice-EFR plants silenced for OsSERK2, or overexpressing rice XB24 are compromised in elf18-induced reactive oxygen production and defense gene expression indicating that these proteins are also important for EFR-mediated signaling in transgenic rice. Taken together, our results demonstrate the potential feasibility of enhancing disease resistance in rice and possibly other monocotyledonous crop species by expression of dicotyledonous PRRs. Our results also suggest that Arabidopsis EFR utilizes at least a subset of the known endogenous rice XA21 signaling components.

  1. Microglial activatory (immunoreceptor tyrosine-based activation motif)- and inhibitory (immunoreceptor tyrosine-based inhibition motif)-signaling receptors for recognition of the neuronal glycocalyx.

    PubMed

    Linnartz, Bettina; Neumann, Harald

    2013-01-01

    Microglia sense intact or lesioned cells of the central nervous system (CNS) and respond accordingly. To fulfill this task, microglia express a whole set of recognition receptors. Fc receptors and DAP12 (TYROBP)-associated receptors such as microglial triggering receptor expressed on myeloid cells-2 (TREM2) and the complement receptor-3 (CR3, CD11b/CD18) trigger the immunoreceptor tyrosine-based activation motif (ITAM)-signaling cascade, resulting in microglial activation, migration, and phagocytosis. Those receptors are counter-regulated by immunoreceptor tyrosine-based inhibition motif (ITIM)-signaling receptors, such as sialic acid-binding immunoglobulin superfamily lectins (Siglecs). Siglecs recognize the sialic acid cap of healthy neurons thus leading to an ITIM signaling that turns down microglial immune responses and phagocytosis. In contrast, desialylated neuronal processes are phagocytosed by microglial CR3 signaling via an adaptor protein containing an ITAM. Thus, the aberrant terminal glycosylation of neuronal surface glycoproteins and glycolipids could serve as a flag for microglia, which display a multitude of diverse carbohydrate-binding receptors that monitor the neuronal physical condition and respond via their ITIM- or ITAM-signaling cascade accordingly.

  2. The soluble pattern recognition receptor PTX3 links humoral innate and adaptive immune responses by helping marginal zone B cells

    PubMed Central

    Sintes, Jordi; Polentarutti, Nadia; Walland, A. Cooper; Yeiser, John R.; Cunha, Cristina; Lacerda, João F.; Salvatori, Giovanni; Blander, J. Magarian

    2016-01-01

    Pentraxin 3 (PTX3) is a fluid-phase pattern recognition receptor of the humoral innate immune system with ancestral antibody-like properties but unknown antibody-inducing function. In this study, we found binding of PTX3 to splenic marginal zone (MZ) B cells, an innate-like subset of antibody-producing lymphocytes strategically positioned at the interface between the circulation and the adaptive immune system. PTX3 was released by a subset of neutrophils that surrounded the splenic MZ and expressed an immune activation–related gene signature distinct from that of circulating neutrophils. Binding of PTX3 promoted homeostatic production of IgM and class-switched IgG antibodies to microbial capsular polysaccharides, which decreased in PTX3-deficient mice and humans. In addition, PTX3 increased IgM and IgG production after infection with blood-borne encapsulated bacteria or immunization with bacterial carbohydrates. This immunogenic effect stemmed from the activation of MZ B cells through a neutrophil-regulated pathway that elicited class switching and plasmablast expansion via a combination of T cell–independent and T cell–dependent signals. Thus, PTX3 may bridge the humoral arms of the innate and adaptive immune systems by serving as an endogenous adjuvant for MZ B cells. This property could be harnessed to develop more effective vaccines against encapsulated pathogens. PMID:27621420

  3. Epitope recognition and T cell receptors in recurrent autoimmune anterior uveitis in Lewis rats immunized with myelin basic protein.

    PubMed

    Adamus, G; Manczak, M; Sugden, B; Arendt, A; Hargrave, P A; Offner, H

    2000-08-01

    Lewis rats immunized with myelin basic protein (MBP) develop experimental autoimmune encephalomyelitis (EAE) and associated anterior uveitis (AU). Rats recover and become resistant to further reinduction of EAE. We investigated whether the resistance to reinduction of EAE was associated with the resistance to AU in LEW rats reinjected with MBP. We demonstrated that while rats remained resistant to EAE, they become susceptible to uveitis after recovery, and suffered a second episode of disease. The susceptibility to reinduced disease was associated with the recognition of new MBP epitopes. In contrast to the initial episode of AU, TCR Vbeta8.2 predominance was not observed in the iris/ciliary body. Our results suggest that T cells specific for MBP, which are rapidly reactivated when re-exposed to antigen, are sufficient to induce clinical uveitis in LEW rats. This process may involve a shifting of T cell specificity from the major encephalitogenic peptide utilizing the Vbeta8.2 receptor to a more diverse cell repertoire.

  4. Binding Sites for Acylated Trehalose Analogs of Glycolipid Ligands on an Extended Carbohydrate Recognition Domain of the Macrophage Receptor Mincle*

    PubMed Central

    Feinberg, Hadar; Rambaruth, Neela D. S.; Jégouzo, Sabine A. F.; Jacobsen, Kristian M.; Djurhuus, Rasmus; Poulsen, Thomas B.; Weis, William I.; Taylor, Maureen E.; Drickamer, Kurt

    2016-01-01

    The macrophage receptor mincle binds to trehalose dimycolate on the surface of Mycobacterium tuberculosis. Signaling initiated by this interaction leads to cytokine production, which underlies the ability of mycobacteria to evade the immune system and also to function as adjuvants. In previous work the mechanism for binding of the sugar headgroup of trehalose dimycolate to mincle has been elucidated, but the basis for enhanced binding to glycolipid ligands, in which hydrophobic substituents are attached to the 6-hydroxyl groups, has been the subject of speculation. In the work reported here, the interaction of trehalose derivatives with bovine mincle has been probed with a series of synthetic mimics of trehalose dimycolate in binding assays, in structural studies by x-ray crystallography, and by site-directed mutagenesis. Binding studies reveal that, rather than reflecting specific structural preference, the apparent affinity of mincle for ligands with hydrophobic substituents correlates with their overall size. Structural and mutagenesis analysis provides evidence for interaction of the hydrophobic substituents with multiple different portions of the surface of mincle and confirms the presence of three Ca2+-binding sites. The structure of an extended portion of the extracellular domain of mincle, beyond the minimal C-type carbohydrate recognition domain, also constrains the way the binding domains may interact on the surface of macrophages. PMID:27542410

  5. Lysophospholipid presentation by CD1d and recognition by a human Natural Killer T-cell receptor

    PubMed Central

    López-Sagaseta, Jacinto; Sibener, Leah V; Kung, Jennifer E; Gumperz, Jenny; Adams, Erin J

    2012-01-01

    Invariant Natural Killer T (iNKT) cells use highly restricted αβ T cell receptors (TCRs) to probe the repertoire of lipids presented by CD1d molecules. Here, we describe our studies of lysophosphatidylcholine (LPC) presentation by human CD1d and its recognition by a native, LPC-specific iNKT TCR. Human CD1d presenting LPC adopts an altered conformation from that of CD1d presenting glycolipid antigens, with a shifted α1 helix resulting in an open A' pocket. Binding of the iNKT TCR requires a 7-Å displacement of the LPC headgroup but stabilizes the CD1d–LPC complex in a closed conformation. The iNKT TCR CDR loop footprint on CD1d–LPC is anchored by the conserved positioning of the CDR3α loop, whereas the remaining CDR loops are shifted, due in part to amino-acid differences in the CDR3β and Jβ segment used by this iNKT TCR. These findings provide insight into how lysophospholipids are presented by human CD1d molecules and how this complex is recognized by some, but not all, human iNKT cells. PMID:22395072

  6. Lysophospholipid presentation by CD1d and recognition by a human Natural Killer T-cell receptor

    SciTech Connect

    López-Sagaseta, Jacinto; Sibener, Leah V.; Kung, Jennifer E.; Gumperz, Jenny; Adams, Erin J.

    2014-10-02

    Invariant Natural Killer T (iNKT) cells use highly restricted {alpha}{beta} T cell receptors (TCRs) to probe the repertoire of lipids presented by CD1d molecules. Here, we describe our studies of lysophosphatidylcholine (LPC) presentation by human CD1d and its recognition by a native, LPC-specific iNKT TCR. Human CD1d presenting LPC adopts an altered conformation from that of CD1d presenting glycolipid antigens, with a shifted {alpha}1 helix resulting in an open A pocket. Binding of the iNKT TCR requires a 7-{angstrom} displacement of the LPC headgroup but stabilizes the CD1d-LPC complex in a closed conformation. The iNKT TCR CDR loop footprint on CD1d-LPC is anchored by the conserved positioning of the CDR3{alpha} loop, whereas the remaining CDR loops are shifted, due in part to amino-acid differences in the CDR3{beta} and J{beta} segment used by this iNKT TCR. These findings provide insight into how lysophospholipids are presented by human CD1d molecules and how this complex is recognized by some, but not all, human iNKT cells.

  7. Oxytocin (OT) and arginine-vasopressin (AVP) act on OT receptors and not AVP V1a receptors to enhance social recognition in adult Syrian hamsters (Mesocricetus auratus).

    PubMed

    Song, Zhimin; Larkin, Tony E; Malley, Maureen O'; Albers, H Elliott

    2016-05-01

    Social recognition is a fundamental requirement for all forms of social relationships. A majority of studies investigating the neural mechanisms underlying social recognition in rodents have investigated relatively neutral social stimuli such as juveniles or ovariectomized females over short time intervals (e.g., 2h). The present study developed a new testing model to study social recognition among adult males using a potent social stimulus. Flank gland odors are used extensively in social communication in Syrian hamsters and convey important information such as dominance status. We found that the recognition of flank gland odors after a 3min exposure lasted for at least 24h, substantially longer than the recognition of other social cues in rats and mice. Intracerebroventricular injections of OT and AVP prolonged the recognition of flank gland odor for up to 48h. Selective OTR but not V1aR agonists, mimicked these enhancing effects of OT and AVP. Similarly, selective OTR but not V1aR antagonists blocked recognition of the odors after 20min. In contrast, the recognition of non-social stimuli was not blocked by either the OTR or the V1aR antagonists. Our findings suggest both OT and AVP enhance social recognition via acting on OTRs and not V1aRs and that the recognition enhancing effects of OT and AVP are limited to social stimuli.

  8. Vortioxetine dose-dependently reverses 5-HT depletion-induced deficits in spatial working and object recognition memory: a potential role for 5-HT1A receptor agonism and 5-HT3 receptor antagonism.

    PubMed

    du Jardin, Kristian Gaarn; Jensen, Jesper Bornø; Sanchez, Connie; Pehrson, Alan L

    2014-01-01

    We previously reported that the investigational multimodal antidepressant, vortioxetine, reversed 5-HT depletion-induced memory deficits while escitalopram and duloxetine did not. The present report studied the effects of vortioxetine and the potential impact of its 5-HT1A receptor agonist and 5-HT3 receptor antagonist properties on 5-HT depletion-induced memory deficits. Recognition and spatial working memory were assessed in the object recognition (OR) and Y-maze spontaneous alternation (SA) tests, respectively. 5-HT depletion was induced in female Long-Evans rats using 4-cholro-DL-phenylalanine methyl ester HCl (PCPA) and receptor occupancies were determined by ex vivo autoradiography. Rats were acutely dosed with vortioxetine, ondansetron (5-HT3 receptor antagonist) or flesinoxan (5-HT1A receptor agonist). The effects of chronic vortioxetine administration on 5-HT depletion-induced memory deficits were also assessed. 5-HT depletion reliably impaired memory performance in both the tests. Vortioxetine reversed PCPA-induced memory deficits dose-dependently with a minimal effective dose (MED) ≤0.1mg/kg (∼80% 5-HT3 receptor occupancy; OR) and ≤3.0mg/kg (5-HT1A, 5-HT1B, 5-HT3 receptor occupancy: ∼15%, 60%, 95%) in SA. Ondansetron exhibited a MED ≤3.0μg/kg (∼25% 5-HT3 receptor occupancy; OR), but was inactive in the SA test. Flesinoxan had a MED ≤1.0mg/kg (∼25% 5-HT1A receptor occupancy; SA); only 1.0mg/kg ameliorated deficits in the NOR. Chronic p.o. vortioxetine administration significantly improved memory performance in OR and occupied 95%, 66%, and 9.5% of 5-HT3, 5-HT1B, and 5-HT1A receptors, respectively. Vortioxetine's effects on SA performance may involve 5-HT1A receptor agonism, but not 5-HT3 receptor antagonism, whereas the effects on OR performance may involve 5-HT3 receptor antagonism and 5-HT1A receptor agonism.

  9. Both IIC and IID Components of Mannose Phosphotransferase System Are Involved in the Specific Recognition between Immunity Protein PedB and Bacteriocin-Receptor Complex

    PubMed Central

    Wang, Chunmei; Ren, Fazheng; Hao, Yanling

    2016-01-01

    Upon exposure to exogenous pediocin-like bacteriocins, immunity proteins specifically bind to the target receptor of the mannose phosphotransferase system components (man-PTS IIC and IID), therefore preventing bacterial cell death. However, the specific recognition of immunity proteins and its associated target receptors remains poorly understood. In this study, we constructed hybrid receptors to identify the domains of IIC and/or IID recognized by the immunity protein PedB, which confers immunity to pediocin PA-1. Using Lactobacillus plantarum man-PTS EII mutant W903, the IICD components of four pediocin PA-1-sensitive strains (L. plantarum WQ0815, Leuconostoc mesenteroides 05–43, Lactobacillus salivarius REN and Lactobacillus acidophilus 05–172) were respectively co-expressed with the immunity protein PedB. Well-diffusions assays showed that only the complex formed by LpIICD from L. plantarum WQ0815 with pediocin PA-1 could be recognized by PedB. In addition, a two-step PCR approach was used to construct hybrid receptors by combining LpIIC or LpIID recognized by PedB with the other three heterologous IID or IIC compounds unrecognized by PedB, respectively. The results showed that all six hybrid receptors were recognized by pediocin PA-1. However, when IIC or IID of L. plantarum WQ0815 was replaced with any corresponding IIC or IID component from L. mesenteroides 05–43, L. salivarius REN and L. acidophilus 05–172, all the hybrid receptors could not be recognized by PedB. Taken altogether, we concluded that both IIC and IID components of the mannose phosphotransferase system play an important role in the specific recognition between the bacteriocin-receptor complex and the immunity protein PedB. PMID:27776158

  10. The metabotropic glutamate 2/3 receptor agonist LY379268 counteracted ketamine-and apomorphine-induced performance deficits in the object recognition task, but not object location task, in rats.

    PubMed

    Pitsikas, Nikolaos; Markou, Athina

    2014-10-01

    Experimental evidence indicates that the non competitive N-methyl-d-aspartate (NMDA) receptor antagonist ketamine and the mixed dopamine (DA) D1/D2 receptor agonist apomorphine induce schizophrenia-like symptoms in rodents, including cognitive deficits. Activation of Group II metabotropic glutamate 2/3 (mGlu2/3) receptors reduces the excessive glutamate release that is hypothesized to be associated with psychiatric disorders. Thus, mGlu2/3 receptor agonists may reverse deficits induced by excessive glutamate or DA release induced by administration of NMDA receptor antagonists and DA receptor agonists, respectively, and potentially those seen in schizophrenia. LY379268 is a selective mGlu2/3 receptor agonist that has shown to be effective in several animal models of stroke, epilepsy, and drug abuse. The present study investigated whether LY379268 antagonizes non-spatial and spatial recognition memory deficits induced by ketamine and apomorphine administration in rats. To assess the effects of the compounds on non-spatial and spatial recognition memory, the object recognition task and object location task were used. Post-training administration of LY379268 (1-3 mg/kg, i.p.) counteracted ketamine (3 mg/kg, i.p.) and apomorphine (1 mg/kg, i.p.)-induced performance deficits in the object recognition task. In contrast, LY379268 (1-3 mg/kg, i.p.) did not attenuate spatial recognition memory deficits produced by ketamine (3 mg/kg, i.p.) or apomorphine (1 mg/kg, i.p.) in the object location task. The present data show that the mGlu2/3 receptor agonist LY379268 reversed non-spatial, but not spatial, recognition memory deficits induced by NMDA receptor blockade or DA receptor agonism in rodents. Thus, such mGlu2/3 receptor agonists may be efficacious in reversing some memory deficits seen in schizophrenia patients.

  11. Object-in-place associative recognition memory depends on glutamate receptor neurotransmission within two defined hippocampal-cortical circuits: a critical role for AMPA and NMDA receptors in the hippocampus, perirhinal, and prefrontal cortices.

    PubMed

    Barker, Gareth Robert Issac; Warburton, Elizabeth Clea

    2015-02-01

    Object-in-place associative recognition memory depends on an interaction between the hippocampus (HPC), perirhinal (PRH), and medial prefrontal (mPFC) cortices, yet the contribution of glutamate receptor neurotransmission to these interactions is unknown. NMDA receptors (NMDAR) in the HPC were critical for encoding of object-in-place memory but not for single-item object recognition. Next, a disconnection procedure was used to examine the importance of "concurrent" glutamate neurotransmission in the HPC-mPFC and HPC-PRH. Contralateral unilateral infusions of NBQX (AMPAR antagonist), into the HPC-mPFC, or HPC-PRH, either before acquisition or test, impaired object-in-place performance. Thus, both circuits are necessary for encoding and retrieval. Crossed unilateral AP5 (NMDAR antagonist) infusions into the HPC-mPFC or HPC-PRH impaired encoding, but not retrieval. Specifically crossed HPC-mPFC infusions impaired both short-term (5 min) and longer term (1 h) memory while HPC-PRH infusions impaired longer term memory only. This delay-dependent effect of AP5 in the HPC-PRH on object-in-place memory, accords with its effects in the PRH, on single item object recognition memory, thereby suggesting that a single PRH synaptic plasticity mechanism underpins different recognition memory processes. Further, blocking excitatory neurotransmission in any pair of structures within the networks impaired "both" encoding and retrieval, thus object-in-place memory clearly requires network interdependency across multiple structures.

  12. Receptor recognition mechanism of human influenza A H1N1 (1918), avian influenza A H5N1 (2004), and pandemic H1N1 (2009) neuraminidase.

    PubMed

    Jongkon, Nipa; Sangma, Chak

    2012-01-01

    Influenza A neuraminidase (NA) is a target for anti-influenza drugs. The function of this enzyme is to cleave a glycosidic linkage of a host cell receptor that links sialic acid (Sia) to galactose (Gal), to allow the virus to leave an infected cell and propagate. The receptor is an oligosaccharide on the host cell surface. There are two types of oligosaccharide receptor; the first, which is found mainly on avian epithelial cell surfaces, links Sia with Gal by an α2,3 glycosidic linkage; in the second, found mainly on human epithelial cell surfaces, linkage is via an α2,6 linkage. Some researchers believe that NAs from different viruses show selectivity for each type of linkage, but there is limited information available to confirm this hypothesis. To see if the linkage type is more specific to any particular NA, a number of NA-receptor complexes of human influenza A H1N1 (1918), avian influenza A H5N1 (2004), and a pandemic strain of H1N1 (2009) were constructed using homology modeling and molecular dynamics simulation. The results show that the two types of receptor analogues bound to NAs use different mechanisms. Moreover, it was found that a residue unique to avian virus NA is responsible for the recognition of the Siaα2,3Gal receptor, and a residue unique to human virus NA is responsible for the recognition of Siaα2,6Gal. We believe that this finding could explain how NAs of different virus origins always possess some unique residues.

  13. Crystal structure of LGR4-Rspo1 complex: insights into the divergent mechanisms of ligand recognition by leucine-rich repeat G-protein-coupled receptors (LGRs).

    PubMed

    Xu, Jin-Gen; Huang, Chunfeng; Yang, Zhengfeng; Jin, Mengmeng; Fu, Panhan; Zhang, Ni; Luo, Jian; Li, Dali; Liu, Mingyao; Zhou, Yan; Zhu, Yongqun

    2015-01-23

    Leucine-rich repeat G-protein-coupled receptors (LGRs) are a unique class of G-protein-coupled receptors characterized by a large extracellular domain to recognize ligands and regulate many important developmental processes. Among the three groups of LGRs, group B members (LGR4-6) recognize R-spondin family proteins (Rspo1-4) to stimulate Wnt signaling. In this study, we successfully utilized the "hybrid leucine-rich repeat technique," which fused LGR4 with the hagfish VLR protein, to obtain two recombinant human LGR4 proteins, LGR415 and LGR49. We determined the crystal structures of ligand-free LGR415 and the LGR49-Rspo1 complex. LGR4 exhibits a twisted horseshoe-like structure. Rspo1 adopts a flat and β-fold architecture and is bound in the concave surface of LGR4 in the complex through electrostatic and hydrophobic interactions. All the Rspo1-binding residues are conserved in LGR4-6, suggesting that LGR4-6 bind R-spondins through an identical surface. Structural analysis of our LGR4-Rspo1 complex with the previously determined LGR4 and LGR5 structures revealed that the concave surface of LGR4 is the sole binding site for R-spondins, suggesting a one-site binding model of LGR4-6 in ligand recognition. The molecular mechanism of LGR4-6 is distinct from the two-step mechanism of group A receptors LGR1-3 and the multiple-interface binding model of group C receptors LGR7-8, suggesting LGRs utilize the divergent mechanisms for ligand recognition. Our structures, together with previous reports, provide a comprehensive understanding of the ligand recognition by LGRs.

  14. Recognition and sensing of biologically relevant anions in alcohol and mixed alcohol-aqueous solutions using charge neutral cleft-like glycol-derived pyridyl-amidothiourea receptors.

    PubMed

    Duke, Rebecca M; McCabe, Thomas; Schmitt, Wolfgang; Gunnlaugsson, Thorfinnur

    2012-04-06

    In this paper, the synthesis and the spectroscopic investigation of new colorimetric receptors for anions 3-6, possessing a glycol chain at the 4-position of the pyridyl ring, and 1 and 2, which lack such a chain, and the X-ray crystal structure of 2 is presented. Structures 3-6 are able to bind to anions in competitive media, such as alcohol or in a mixture of methanol and water, where the anion recognition gives rise to changes in the absorption spectra, which is red-shifted, in 1:1 or 1:2 (sensor/anion) stoichiometry. The anion recognition for 1 and 2 was also investigated in organic solvents and in a 4:1 mixture of DMSO/H(2)O. The binding of 1 to anions such as acetate, phosphate, and fluoride was also evaluated using (1)H NMR in DMSO-d(6).

  15. Substituent directed selectivity in anion recognition by a new class of simple osmium-pyrazole derived receptors.

    PubMed

    Das, Ankita; Mondal, Prasenjit; Dasgupta, Moumita; Kishore, Nand; Lahiri, Goutam Kumar

    2016-02-14

    The present article deals with the structurally, spectroscopically and electrochemically characterised osmium-bipyridyl derived complexes [(bpy)2Os(II)(HL1)Cl]ClO4 [1]ClO4 and [(bpy)2Os(II)(HL2)Cl]ClO4 [2]ClO4 incorporating neutral and monodentate pyrazole derivatives (HL) with one free NH function (bpy = 2,2'-bipyridine, HL1 = pyrazole, HL2 = 3,5-dimethylpyrazole). The crystal structures of [1]ClO4 and [2]ClO4 reveal intramolecular hydrogen bonding interactions between the free NH proton of HL and the equatorially placed Cl(-) ligand (N-HCl) with donor-acceptor distances of 3.114(7) Å and 3.153(6) Å as well as intermolecular hydrogen bonding interactions between the NH proton and one of the oxygen atoms of ClO4(-) (N-HO) with donor-acceptor distances of 2.870(10) Å and 3.024(8) Å, respectively. The effect of hydrogen bonding interactions has translated into the less acidic nature of the NH proton of the coordinated HL with estimated pKa > 12. 1(+) and 2(+) exhibit reversible Os(II)/(III) and irreversible Os(III)/(IV) processes in CH3CN within ± 2.0 V versus SCE. The effect of 3,5-dimethyl substituted HL2 on 2(+) has been reflected in the appreciable lowering (40 mV) of the Os(II/III) potential, along with the further decrease in the acidity of the NH proton (pKa > 13.0) with regard to HL1 coordinated 1(+) (pKa: ∼ 12.3). The electronic spectral features of Os(ii) (1(+)/2(+)) and electrochemically generated Os(III) (1(2+)/2(2+)) derived complexes have been analysed by TD-DFT calculations. The efficacy of the 1(+) and 2(+) encompassing free NH proton towards the anion recognition process has been evaluated by different experimental investigations using a wide variety of anions. It however establishes that receptor 1(+) can recognise both F(-) and OAc(-) in acetonitrile solution, while 2(+) is exclusively selective for the F(-) ion.

  16. Crystal Structure of the PAC1R Extracellular Domain Unifies a Consensus Fold for Hormone Recognition by Class B G-Protein Coupled Receptors

    SciTech Connect

    Kumar, Shiva; Pioszak, Augen; Zhang, Chenghai; Swaminathan, Kunchithapadam; Xu, H. Eric

    2012-02-21

    Pituitary adenylate cyclase activating polypeptide (PACAP) is a member of the PACAP/glucagon family of peptide hormones, which controls many physiological functions in the immune, nervous, endocrine, and muscular systems. It activates adenylate cyclase by binding to its receptor, PAC1R, a member of class B G-protein coupled receptors (GPCR). Crystal structures of a number of Class B GPCR extracellular domains (ECD) bound to their respective peptide hormones have revealed a consensus mechanism of hormone binding. However, the mechanism of how PACAP binds to its receptor remains controversial as an NMR structure of the PAC1R ECD/PACAP complex reveals a different topology of the ECD and a distinct mode of ligand recognition. Here we report a 1.9 {angstrom} crystal structure of the PAC1R ECD, which adopts the same fold as commonly observed for other members of Class B GPCR. Binding studies and cell-based assays with alanine-scanned peptides and mutated receptor support a model that PAC1R uses the same conserved fold of Class B GPCR ECD for PACAP binding, thus unifying the consensus mechanism of hormone binding for this family of receptors.

  17. Structure of Natural Killer Receptor 2B4 Bound to CD48 Reveals Basis for Heterophilic Recognition in Signaling Lymphocyte Activation Molecule Family

    SciTech Connect

    Velikovsky,C.; Deng, L.; Chlewicki, L.; Fernandez, M.; Kumar, V.; Mariuzza, R.

    2007-01-01

    Natural killer (NK) cells eliminate virally infected and tumor cells. Among the receptors regulating NK cell function is 2B4 (CD244), a member of the signaling lymphocyte-activation molecule (SLAM) family that binds CD48. 2B4 is the only heterophilic receptor of the SLAM family, whose other members, e.g., NK-T-B-antigen (NTB-A), are self-ligands. We determined the structure of the complex between the N-terminal domains of mouse 2B4 and CD48, as well as the structures of unbound 2B4 and CD48. The complex displayed an association mode related to, yet distinct from, that of the NTB-A dimer. Binding was accompanied by the rigidification of flexible 2B4 regions containing most of the polymorphic residues across different species and receptor isoforms. We propose a model for 2B4-CD48 interactions that permits the intermixing of SLAM receptors with major histocompatibility complex-specific receptors in the NK cell immune synapse. This analysis revealed the basis for heterophilic recognition within the SLAM family.

  18. Adjunctive selective estrogen receptor modulator increases neural activity in the hippocampus and inferior frontal gyrus during emotional face recognition in schizophrenia

    PubMed Central

    Ji, E; Weickert, C S; Lenroot, R; Kindler, J; Skilleter, A J; Vercammen, A; White, C; Gur, R E; Weickert, T W

    2016-01-01

    Estrogen has been implicated in the development and course of schizophrenia with most evidence suggesting a neuroprotective effect. Treatment with raloxifene, a selective estrogen receptor modulator, can reduce symptom severity, improve cognition and normalize brain activity during learning in schizophrenia. People with schizophrenia are especially impaired in the identification of negative facial emotions. The present study was designed to determine the extent to which adjunctive raloxifene treatment would alter abnormal neural activity during angry facial emotion recognition in schizophrenia. Twenty people with schizophrenia (12 men, 8 women) participated in a 13-week, randomized, double-blind, placebo-controlled, crossover trial of adjunctive raloxifene treatment (120 mg per day orally) and performed a facial emotion recognition task during functional magnetic resonance imaging after each treatment phase. Two-sample t-tests in regions of interest selected a priori were performed to assess activation differences between raloxifene and placebo conditions during the recognition of angry faces. Adjunctive raloxifene significantly increased activation in the right hippocampus and left inferior frontal gyrus compared with the placebo condition (family-wise error, P<0.05). There was no significant difference in performance accuracy or reaction time between active and placebo conditions. To the best of our knowledge, this study provides the first evidence suggesting that adjunctive raloxifene treatment changes neural activity in brain regions associated with facial emotion recognition in schizophrenia. These findings support the hypothesis that estrogen plays a modifying role in schizophrenia and shows that adjunctive raloxifene treatment may reverse abnormal neural activity during facial emotion recognition, which is relevant to impaired social functioning in men and women with schizophrenia. PMID:27138794

  19. Adjunctive selective estrogen receptor modulator increases neural activity in the hippocampus and inferior frontal gyrus during emotional face recognition in schizophrenia.

    PubMed

    Ji, E; Weickert, C S; Lenroot, R; Kindler, J; Skilleter, A J; Vercammen, A; White, C; Gur, R E; Weickert, T W

    2016-05-03

    Estrogen has been implicated in the development and course of schizophrenia with most evidence suggesting a neuroprotective effect. Treatment with raloxifene, a selective estrogen receptor modulator, can reduce symptom severity, improve cognition and normalize brain activity during learning in schizophrenia. People with schizophrenia are especially impaired in the identification of negative facial emotions. The present study was designed to determine the extent to which adjunctive raloxifene treatment would alter abnormal neural activity during angry facial emotion recognition in schizophrenia. Twenty people with schizophrenia (12 men, 8 women) participated in a 13-week, randomized, double-blind, placebo-controlled, crossover trial of adjunctive raloxifene treatment (120 mg per day orally) and performed a facial emotion recognition task during functional magnetic resonance imaging after each treatment phase. Two-sample t-tests in regions of interest selected a priori were performed to assess activation differences between raloxifene and placebo conditions during the recognition of angry faces. Adjunctive raloxifene significantly increased activation in the right hippocampus and left inferior frontal gyrus compared with the placebo condition (family-wise error, P<0.05). There was no significant difference in performance accuracy or reaction time between active and placebo conditions. To the best of our knowledge, this study provides the first evidence suggesting that adjunctive raloxifene treatment changes neural activity in brain regions associated with facial emotion recognition in schizophrenia. These findings support the hypothesis that estrogen plays a modifying role in schizophrenia and shows that adjunctive raloxifene treatment may reverse abnormal neural activity during facial emotion recognition, which is relevant to impaired social functioning in men and women with schizophrenia.

  20. Analogs of alpha-melanocyte stimulating hormone with high agonist potency and selectivity at human melanocortin receptor 1b: the role of Trp(9) in molecular recognition.

    PubMed

    Bednarek, Maria A; Macneil, Tanya; Tang, Rui; Fong, Tung M; Angeles Cabello, M; Maroto, Marta; Teran, Ana

    2008-05-01

    alpha-Melanocyte stimulating hormone (alphaMSH), Ac-Ser(1)-Tyr(2)-Ser(3)-Met(4)-Glu(5)-His(6)-Phe(7)-Arg(8)-Trp(9)-Gly(10)-Lys(11)-Pro(12)-Val(13)-NH(2), is an endogenous agonist for the melanocortin receptor 1 (MC1R), the receptor found in the skin, several types of immune cells, and other peripheral sites. Three-dimensional models of complexes of this receptor with alphaMSH and its synthetic analog NDP-alphaMSH, Ac-Ser(1)-Tyr(2)-Ser(3)-Nle(4)-Glu(5)-His(6)-D-Phe(7)-Arg(8)-Trp(9)-Gly(10)-Lys(11)-Pro(12)-Val(13)-NH(2), have been previously proposed. In those models, the 6-9 segment of the ligand was considered essential for the ligand-receptor interactions. In this study, we probed the role of Trp(9) of NDP-alphaMSH in interactions with hMC1bR. Analogs of NDP-alphaMSH with various amino acids in place of Trp(9) were synthesized and tested in vitro in receptor affinity binding and cAMP functional assays at human melanocortin receptors 1b, 3, 4, and 5 (hMC1b,3-5R). Several new compounds displayed high agonist potency at hMC1bR (EC(50) = 0.5-5 nM) and receptor subtype selectivity greater than 2000-fold versus hMC3-5R. The Trp(9) residue of NDP-alphaMSH was determined to be not essential for molecular recognition at hMC1bR.

  1. Kin Recognition in Bacteria.

    PubMed

    Wall, Daniel

    2016-09-08

    The ability of bacteria to recognize kin provides a means to form social groups. In turn these groups can lead to cooperative behaviors that surpass the ability of the individual. Kin recognition involves specific biochemical interactions between a receptor(s) and an identification molecule(s). Recognition specificity, ensuring that nonkin are excluded and kin are included, is critical and depends on the number of loci and polymorphisms involved. After recognition and biochemical perception, the common ensuing cooperative behaviors include biofilm formation, quorum responses, development, and swarming motility. Although kin recognition is a fundamental mechanism through which cells might interact, microbiologists are only beginning to explore the topic. This review considers both molecular and theoretical aspects of bacterial kin recognition. Consideration is also given to bacterial diversity, genetic relatedness, kin selection theory, and mechanisms of recognition.

  2. Selective Blockade of Dopamine D3 Receptors Enhances while D2 Receptor Antagonism Impairs Social Novelty Discrimination and Novel Object Recognition in Rats: A Key Role for the Prefrontal Cortex

    PubMed Central

    Watson, David JG; Loiseau, Florence; Ingallinesi, Manuela; Millan, Mark J; Marsden, Charles A; Fone, Kevin CF

    2012-01-01

    Dopamine D3 receptor antagonists exert pro-cognitive effects in both rodents and primates. Accordingly, this study compared the roles of dopamine D3 vs D2 receptors in social novelty discrimination (SND), which relies on olfactory cues, and novel object recognition (NOR), a visual-recognition task. The dopamine D3 receptor antagonist, S33084 (0.04–0.63 mg/kg), caused a dose-related reversal of delay-dependent impairment in both SND and NOR procedures in adult rats. Furthermore, mice genetically deficient in dopamine D3 receptors displayed enhanced discrimination in the SND task compared with wild-type controls. In contrast, acute treatment with the preferential dopamine D2 receptor antagonist, L741,626 (0.16–5.0 mg/kg), or with the dopamine D3 agonist, PD128,907 (0.63–40 μg/kg), caused a dose-related impairment in performance in rats in both tasks after a short inter-trial delay. Bilateral microinjection of S33084 (2.5 μg/side) into the prefrontal cortex (PFC) of rats increased SND and caused a dose-related (0.63–2.5 μg/side) improvement in NOR, while intra-striatal injection (2.5 μg/side) had no effect on either. In contrast, bilateral microinjection of L741,626 into the PFC (but not striatum) caused a dose-related (0.63–2.5 μg/side) impairment of NOR. These observations suggest that blockade of dopamine D3 receptors enhances both SND and NOR, whereas D3 receptor activation or antagonism of dopamine D2 receptor impairs cognition in these paradigms. Furthermore, these actions are mediated, at least partly, by the PFC. These data have important implications for exploitation of dopaminergic mechanisms in the treatment of schizophrenia and other CNS disorders, and support the potential therapeutic utility of dopamine D3 receptor antagonism. PMID:22030711

  3. Recognition of the Magnaporthe oryzae Effector AVR-Pia by the Decoy Domain of the Rice NLR Immune Receptor RGA5[OPEN

    PubMed Central

    Ortiz, Diana; de Guillen, Karine; Cesari, Stella; Chalvon, Véronique

    2017-01-01

    Nucleotide binding domain and leucine-rich repeat proteins (NLRs) are important receptors in plant immunity that allow recognition of pathogen effectors. The rice (Oryza sativa) NLR RGA5 recognizes the Magnaporthe oryzae effector AVR-Pia through direct interaction. Here, we gained detailed insights into the molecular and structural bases of AVR-Pia-RGA5 interaction and the role of the RATX1 decoy domain of RGA5. NMR titration combined with in vitro and in vivo protein-protein interaction analyses identified the AVR-Pia interaction surface that binds to the RATX1 domain. Structure-informed AVR-Pia mutants showed that, although AVR-Pia associates with additional sites in RGA5, binding to the RATX1 domain is necessary for pathogen recognition but can be of moderate affinity. Therefore, RGA5-mediated resistance is highly resilient to mutations in the effector. We propose a model that explains such robust effector recognition as a consequence, and an advantage, of the combination of integrated decoy domains with additional independent effector-NLR interactions. PMID:28087830

  4. HSV Infection Induces Production of ROS, which Potentiate Signaling from Pattern Recognition Receptors: Role for S-glutathionylation of TRAF3 and 6

    PubMed Central

    Rahbek, Stine H.; Ichijo, Hidenori; Chen, Zhijian J.; Mieyal, John J.; Hartmann, Rune; Paludan, Søren R.

    2011-01-01

    The innate immune response constitutes the first line of defense against infections. Pattern recognition receptors recognize pathogen structures and trigger intracellular signaling pathways leading to cytokine and chemokine expression. Reactive oxygen species (ROS) are emerging as an important regulator of some of these pathways. ROS directly interact with signaling components or induce other post-translational modifications such as S-glutathionylation, thereby altering target function. Applying live microscopy, we have demonstrated that herpes simplex virus (HSV) infection induces early production of ROS that are required for the activation of NF-κB and IRF-3 pathways and the production of type I IFNs and ISGs. All the known receptors involved in the recognition of HSV were shown to be dependent on the cellular redox levels for successful signaling. In addition, we provide biochemical evidence suggesting S-glutathionylation of TRAF family proteins to be important. In particular, by performing mutational studies we show that S-glutathionylation of a conserved cysteine residue of TRAF3 and TRAF6 is important for ROS-dependent activation of innate immune pathways. In conclusion, these findings demonstrate that ROS are essential for effective activation of signaling pathways leading to a successful innate immune response against HSV infection. PMID:21949653

  5. Emerging Bordetella pertussis Strains Induce Enhanced Signaling of Human Pattern Recognition Receptors TLR2, NOD2 and Secretion of IL-10 by Dendritic Cells

    PubMed Central

    Hovingh, Elise S.; van Gent, Marjolein; Hamstra, Hendrik-Jan; Demkes, Marc; Mooi, Frits R.; Pinelli, Elena

    2017-01-01

    Vaccines against pertussis have been available for more than 60 years. Nonetheless, this highly contagious disease is reemerging even in countries with high vaccination coverage. Genetic changes of Bordetella pertussis over time have been suggested to contribute to the resurgence of pertussis, as these changes may favor escape from vaccine-induced immunity. Nonetheless, studies on the effects of these bacterial changes on the immune response are limited. Here, we characterize innate immune recognition and activation by a collection of genetically diverse B. pertussis strains isolated from Dutch pertussis patients before and after the introduction of the pertussis vaccines. For this purpose, we used HEK-Blue cells transfected with human pattern recognition receptors TLR2, TLR4, NOD2 and NOD1 as a high throughput system for screening innate immune recognition of more than 90 bacterial strains. Physiologically relevant human monocyte derived dendritic cells (moDC), purified from peripheral blood of healthy donors were also used. Findings indicate that, in addition to inducing TLR2 and TLR4 signaling, all B. pertussis strains activate the NOD-like receptor NOD2 but not NOD1. Furthermore, we observed a significant increase in TLR2 and NOD2, but not TLR4, activation by strains circulating after the introduction of pertussis vaccines. When using moDC, we observed that the recently circulating strains induced increased activation of these cells with a dominant IL-10 production. In addition, we observed an increased expression of surface markers including the regulatory molecule PD-L1. Expression of PD-L1 was decreased upon blocking TLR2. These in vitro findings suggest that emerging B. pertussis strains have evolved to dampen the vaccine-induced inflammatory response, which would benefit survival and transmission of this pathogen. Understanding how this disease has resurged in a highly vaccinated population is crucial for the design of improved vaccines against pertussis

  6. Selective perrhenate recognition in pure water by halogen bonding and hydrogen bonding alpha-cyclodextrin based receptors.

    PubMed

    Cornes, Stuart P; Sambrook, Mark R; Beer, Paul D

    2017-03-20

    Alpha-cyclodextrin based anion receptors functionalised with pendant arms containing halogen and hydrogen bond donor motifs display selective association of perrhenate in aqueous media at neutral pH. NMR and ITC anion binding investigations reveal the halogen bonding receptor to be the superior host.

  7. Distinct Conformations of Ly49 Natural Killer Cell Receptors Mediate MHC Class I Recognition in Trans and Cis

    SciTech Connect

    Back, J.; Malchiodi, E; Cho, S; Scarpellino, L; Schneider, P; Kerzic, M; Mariuzza, R; Held, W

    2009-01-01

    Certain cell-surface receptors engage ligands expressed on juxtaposed cells and ligands on the same cell. The structural basis for trans versus cis binding is not known. Here, we showed that Ly49 natural killer (NK) cell receptors bound two MHC class I (MHC-I) molecules in trans when the two ligand-binding domains were backfolded onto the long stalk region. In contrast, dissociation of the ligand-binding domains from the stalk and their reorientation relative to the NK cell membrane allowed monovalent binding of MHC-I in cis. The distinct conformations (backfolded and extended) define the structural basis for cis-trans binding by Ly49 receptors and explain the divergent functional consequences of cis versus trans interactions. Further analyses identified specific stalk segments that were not required for MHC-I binding in trans but were essential for inhibitory receptor function. These data identify multiple distinct roles of stalk regions for receptor function.

  8. Oestrogen and Progesterone Receptors and COX-2 Expression in Endometrial Biopsy Samples During Maternal Recognition of Pregnancy in Llamas (Lama glama).

    PubMed

    Bianchi, C P; Meikle, A; Benavente, M A; Álvarez, M A; Trasorras, V L; Miragaya, M H; Rodríguez, E; Aba, M A

    2015-12-01

    Endometrial expression of oestrogen receptor-α (ERα), progesterone receptor (PR) and cyclooxigenase-2 (COX-2) was evaluated in non-pregnant and pregnant llamas during the period when luteolysis/maternal recognition of pregnancy is expected to occur. Females (n = 28) were divided into two groups: non-pregnant llamas were induced to ovulate with a Buserelin injection, and endometrial biopsies were obtained on day 8 (n = 5) or 12 (n = 5) post-induction of ovulation. Animals of the pregnant group (n = 18) were mated with a fertile male. Pregnancy was confirmed by the visualization of the embryo collected by transcervical flushing in 5 of 9 animals on day 8 post-mating and by progesterone profile on day 12 post-mating in 4 of 9 animals, when endometrial biopsies were obtained. An immunohistochemical technique was used to evaluate receptors population and COX-2 expression. Pregnant llamas showed a higher percentage of positive cells and stronger intensity for ERα than for non-pregnant llamas in stroma on day 8 and in the luminal epithelium on day 12 post-induction of ovulation, while a deep decrease in endometrial PR population was reported in pregnant llamas on that day in luminal and glandular epithelia and stroma. In the luminal epithelium, COX-2 expression was lower in pregnant than in non-pregnant animals. Briefly, the increase of ERα in pregnant llamas gives further support to the hypothesis that oestrogens are involved in the mechanism of maternal recognition of pregnancy. Endometrial PR decrease in pregnant llamas might be a necessary event to allow the expression of proteins involved in conceptus attachment, a mechanism widely accepted in other species. Moreover, embryo seems to attenuate maternal PGF(2α) secretion during early pregnancy by decreasing the endometrial expression of COX-2 in the luminal epithelium of pregnant llamas.

  9. Changes in mate recognition through alterations of pheromones and receptors in the multisexual mushroom fungus Schizophyllum commune.

    PubMed Central

    Fowler, T J; Mitton, M F; Vaillancourt, L J; Raper, C A

    2001-01-01

    Schizophyllum commune has thousands of mating types defined in part by numerous lipopeptide pheromones and their G-protein-coupled receptors. These molecules are encoded within multiple versions of two redundantly functioning B mating-type loci, B alpha and B beta. Compatible combinations of pheromones and receptors, produced by individuals of different B mating types, trigger a pathway of fertilization required for sexual development. Analysis of the B beta 2 mating-type locus revealed a large cluster of genes encoding a single pheromone receptor and eight different pheromones. Phenotypic effects of mutations within these genes indicated that small changes in both types of molecules could significantly alter their specificity of interaction. For example, a conservative amino acid substitution in a pheromone resulted in a gain of function toward one receptor and a loss of function with another. A two-amino-acid deletion from a receptor precluded the mutant pheromone from activating the mutant receptor, yet this receptor was activated by other pheromones. Sequence comparisons provided clues toward understanding how so many variants of these multigenic loci could have evolved through duplication and mutational divergence. A three-step model for the origin of new variants comparable to those found in nature is presented. PMID:11514441

  10. Role of the extracellular loops of G protein-coupled receptors in ligand recognition: a molecular modeling study of the human P2Y1 receptor.

    PubMed

    Moro, S; Hoffmann, C; Jacobson, K A

    1999-03-23

    The P2Y1 receptor is a G protein-coupled receptor (GPCR) and is stimulated by extracellular ADP and ATP. Site-directed mutagenesis of the three extracellular loops (ELs) of the human P2Y1 receptor indicates the existence of two essential disulfide bridges (Cys124 in EL1 and Cys202 in EL2; Cys42 in the N-terminal segment and Cys296 in EL3) and several specific ionic and H-bonding interactions (involving Glu209 and Arg287). Through molecular modeling and molecular dynamics simulations, an energetically sound conformational hypothesis for the receptor has been calculated that includes transmembrane (TM) domains (using the electron density map of rhodopsin as a template), extracellular loops, and a truncated N-terminal region. ATP may be docked in the receptor, both within the previously defined TM cleft and within two other regions of the receptor, termed meta-binding sites, defined by the extracellular loops. The first meta-binding site is located outside of the TM bundle, between EL2 and EL3, and the second higher energy site is positioned immediately underneath EL2. Binding at both the principal TM binding site and the lower energy meta-binding sites potentially affects the observed ligand potency. In meta-binding site I, the side chain of Glu209 (EL2) is within hydrogen-bonding distance (2.8 A) of the ribose O3', and Arg287 (EL3) coordinates both alpha- and beta-phosphates of the triphosphate chain, consistent with the insensitivity in potency of the 5'-monophosphate agonist, HT-AMP, to mutation of Arg287 to Lys. Moreover, the selective reduction in potency of 3'NH2-ATP in activating the E209R mutant receptor is consistent with the hypothesis of direct contact between EL2 and nucleotide ligands. Our findings support ATP binding to at least two distinct domains of the P2Y1 receptor, both outside and within the TM core. The two disulfide bridges present in the human P2Y1 receptor play a major role in the structure and stability of the receptor, to constrain the

  11. Perception of the Arabidopsis Danger Signal Peptide 1 Involves the Pattern Recognition Receptor AtPEPR1 and Its Close Homologue AtPEPR2*

    PubMed Central

    Krol, Elzbieta; Mentzel, Tobias; Chinchilla, Delphine; Boller, Thomas; Felix, Georg; Kemmerling, Birgit; Postel, Sandra; Arents, Michael; Jeworutzki, Elena; Al-Rasheid, Khaled A. S.; Becker, Dirk; Hedrich, Rainer

    2010-01-01

    Plasma membrane-borne pattern recognition receptors, which recognize microbe-associated molecular patterns and endogenous damage-associated molecular patterns, provide the first line of defense in innate immunity. In plants, leucine-rich repeat receptor kinases fulfill this role, as exemplified by FLS2 and EFR, the receptors for the microbe-associated molecular patterns flagellin and elongation factor Tu. Here we examined the perception of the damage-associated molecular pattern peptide 1 (AtPep1), an endogenous peptide of Arabidopsis identified earlier and shown to be perceived by the leucine-rich repeat protein kinase PEPR1. Using seedling growth inhibition, elicitation of an oxidative burst and induction of ethylene biosynthesis, we show that wild type plants and the pepr1 and pepr2 mutants, affected in PEPR1 and in its homologue PEPR2, are sensitive to AtPep1, but that the double mutant pepr1/pepr2 is completely insensitive. As a central body of our study, we provide electrophysiological evidence that at the level of the plasma membrane, AtPep1 triggers a receptor-dependent transient depolarization through activation of plasma membrane anion channels, and that this effect is absent in the double mutant pepr1/pepr2. The double mutant also fails to respond to AtPep2 and AtPep3, two distant homologues of AtPep1 on the basis of homology screening, implying that the PEPR1 and PEPR2 are responsible for their perception too. Our findings provide a basic framework to study the biological role of AtPep1-related danger signals and their cognate receptors. PMID:20200150

  12. The role of pattern-recognition receptors in graft-versus-host disease and graft-versus-leukemia after allogeneic stem cell transplantation.

    PubMed

    Heidegger, Simon; van den Brink, Marcel R M; Haas, Tobias; Poeck, Hendrik

    2014-01-01

    Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is the only treatment with curative potential for certain aggressive hematopoietic malignancies. Its success is limited by acute graft-versus-host disease (GVHD), a life-threatening complication that occurs when allo-reactive donor T cells attack recipient organs. There is growing evidence that microbes and innate pattern-recognition receptors (PRRs) such as toll-like receptors (TLR) and nod-like receptors (NLR) are critically involved in the pathogenesis of acute GVHD. Currently, a widely accepted model postulates that intensive chemotherapy and/or total-body irradiation during pre-transplant conditioning results in tissue damage and a loss of epithelial barrier function. Subsequent translocation of bacterial components as well as release of endogenous danger molecules stimulate PRRs of host antigen-presenting cells to trigger the production of pro-inflammatory cytokines (cytokine storm) that modulate T cell allo-reactivity against host tissues, but eventually also the beneficial graft-versus-leukemia (GVL) effect. Given the limitations of existing immunosuppressive therapies, a better understanding of the molecular mechanisms that govern GVHD versus GVL is urgently needed. This may ultimately allow to design modulators, which protect from GvHD but preserve donor T-cell attack on hematologic malignancies. Here, we will briefly summarize current knowledge about the role of innate immunity in the pathogenesis of GVHD and GVL following allo-HSCT.

  13. Evolutionarily conserved recognition and innate immunity to fungal pathogens by the scavenger receptors SCARF1 and CD36

    PubMed Central

    Mylonakis, Eleftherios; Tampakakis, Emmanouil; Colvin, Richard A.; Seung, Edward; Puckett, Lindsay; Tai, Melissa F.; Stewart, Cameron R.; Pukkila-Worley, Read; Hickman, Suzanne E.; Moore, Kathryn J.; Calderwood, Stephen B.; Hacohen, Nir; Luster, Andrew D.; El Khoury, Joseph

    2009-01-01

    Receptors involved in innate immunity to fungal pathogens have not been fully elucidated. We show that the Caenorhabditis elegans receptors CED-1 and C03F11.3, and their mammalian orthologues, the scavenger receptors SCARF1 and CD36, mediate host defense against two prototypic fungal pathogens, Cryptococcus neoformans and Candida albicans. CED-1 and C03F11.1 mediated antimicrobial peptide production and were necessary for nematode survival after C. neoformans infection. SCARF1 and CD36 mediated cytokine production and were required for macrophage binding to C. neoformans, and control of the infection in mice. Binding of these pathogens to SCARF1 and CD36 was β-glucan dependent. Thus, CED-1/SCARF1 and C03F11.3/CD36 are β-glucan binding receptors and define an evolutionarily conserved pathway for the innate sensing of fungal pathogens. PMID:19237602

  14. The GM-CSF/IL-3/IL-5 cytokine receptor family: from ligand recognition to initiation of signaling.

    PubMed

    Broughton, Sophie E; Dhagat, Urmi; Hercus, Timothy R; Nero, Tracy L; Grimbaldeston, Michele A; Bonder, Claudine S; Lopez, Angel F; Parker, Michael W

    2012-11-01

    Granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), and IL-5 are members of a discrete family of cytokines that regulates the growth, differentiation, migration and effector function activities of many hematopoietic cells and immunocytes. These cytokines are involved in normal responses to infectious agents, bridging innate and adaptive immunity. However, in certain cases, the overexpression of these cytokines or their receptors can lead to excessive or aberrant initiation of signaling resulting in pathological conditions, with chronic inflammatory diseases and myeloid leukemias the most notable examples. Recent crystal structures of the GM-CSF receptor ternary complex and the IL-5 binary complex have revealed new paradigms of cytokine receptor activation. Together with a wealth of associated structure-function studies, they have significantly enhanced our understanding of how these receptors recognize cytokines and initiate signals across cell membranes. Importantly, these structures provide opportunities for structure-based approaches for the discovery of novel and disease-specific therapeutics. In addition, recent biochemical evidence has suggested that the GM-CSF/IL-3/IL-5 receptor family is capable of interacting productively with other membrane proteins at the cell surface. Such interactions may afford additional or unique biological activities and might be harnessed for selective modulation of the function of these receptors in disease.

  15. One-trial object recognition memory in the domestic rabbit (Oryctolagus cuniculus) is disrupted by NMDA receptor antagonists.

    PubMed

    Hoffman, Kurt Leroy; Basurto, Enrique

    2013-08-01

    The spontaneous response to novelty is the basis of one-trial object recognition tests for the study of object recognition memory (ORM) in rodents. We describe an object recognition task for the rabbit, based on its natural tendency to scent-mark ("chin") novel objects. The object recognition task comprised a 15min sample phase in which the rabbit was placed into an open field arena containing two similar objects, then removed for a 5-360min delay, and then returned to the same arena that contained one object similar to the original ones ("Familiar") and one that differed from the original ones ("Novel"), for a 15min test phase. Chin-marks directed at each of the objects were registered. Some animals received injections (sc) of saline, ketamine (1mg/kg), or MK-801 (37μg/kg), 5 or 20min before the sample phase. We found that chinning decreased across the sample phase, and that this response showed stimulus specificity, a defining characteristic of habituation: in the test phase, chinning directed at the Novel, but not Familiar, object was increased. Chinning directed preferentially at the novel object, which we interpret as novelty-induced sensitization and the behavioral correlate of ORM, was promoted by tactile/visual and spatial novelty. ORM deficits were induced by pre-treatment with MK-801 and, to a lesser extent, ketamine. Novel object discrimination was not observed after delays longer than 5min. These results suggest that short-term habituation and sensitization, not long-term memory, underlie novel object discrimination in this test paradigm.

  16. Pattern recognition receptor mediated downregulation of microRNA‐650 fine‐tunes MxA expression in dendritic cells infected with influenza A virus

    PubMed Central

    Khatamzas, Elham; Liu, Xiao; Brain, Oliver; Delmiro Garcia, Magno; Leslie, Alasdair; Danis, Benedicte; Mayer, Alice; Baban, Dilair; Ragoussis, Jiannis; Weber, Alexander N. R.; Simmons, Alison

    2015-01-01

    MicroRNAs are important posttranscriptional regulators of gene expression, which have been shown to fine‐tune innate immune responses downstream of pattern recognition receptor (PRR) signaling. This study identifies miR‐650 as a novel PRR‐responsive microRNA that is downregulated upon stimulation of primary human monocyte‐derived dendritic cells (MDDCs) with a variety of different microbe‐associated molecular patterns. A comprehensive target search combining in silico analysis, transcriptional profiling, and reporter assays reveals that miR‐650 regulates several well‐known interferon‐stimulated genes, including IFIT2 and MXA. In particular, downregulation of miR‐650 in influenza A infected MDDCs enhances the expression of MxA and may therefore contribute to the establishment of an antiviral state. Together these findings reveal a novel link between miR‐650 and the innate immune response in human MDDCs. PMID:26460926

  17. Ethanol and Caffeine Effects on Social Interaction and Recognition in Mice: Involvement of Adenosine A2A and A1 Receptors.

    PubMed

    López-Cruz, Laura; San-Miguel, Noemí; Bayarri, Pilar; Baqi, Younis; Müller, Christa E; Salamone, John D; Correa, Mercé

    2016-01-01

    Ethanol and caffeine are frequently consumed in combination and have opposite effects on the adenosine system: ethanol metabolism leads to an increase in adenosine levels, while caffeine is a non-selective adenosine A1/A2A receptor antagonist. These receptors are highly expressed in striatum and olfactory tubercle, brain areas involved in exploration and social interaction in rodents. Ethanol modulates social interaction processes, but the role of adenosine in social behavior is still poorly understood. The present work was undertaken to study the impact of ethanol, caffeine and their combination on social behavior, and to explore the involvement of A1 and A2A receptors on those actions. Male CD1 mice were evaluated in a social interaction three-chamber paradigm, for preference of conspecific vs. object, and also for long-term recognition memory of familiar vs. novel conspecific. Ethanol showed a biphasic effect, with low doses (0.25 g/kg) increasing social contact and higher doses (1.0-1.5 g/kg) reducing social interaction. However, no dose changed social preference; mice always spent more time sniffing the conspecific than the object, independently of the ethanol dose. Ethanol, even at doses that did not change social exploration, produced amnestic effects on social recognition the following day. Caffeine reduced social contact (15.0-60.0 mg/kg), and even blocked social preference at higher doses (30.0-60.0 mg/kg). The A1 antagonist Cyclopentyltheophylline (CPT; 3-9 mg/kg) did not modify social contact or preference on its own, and the A2A antagonist MSX-3 (1.5-6 mg/kg) increased social interaction at all doses. Ethanol at intermediate doses (0.5-1.0 g/kg) was able to reverse the reduction in social exploration induced by caffeine (15.0-30.0 mg/kg). Although there was no interaction between ethanol and CPT or MSX-3 on social exploration in the first day, MSX-3 blocked the amnestic effects of ethanol observed on the following day. Thus, ethanol impairs the

  18. Ethanol and Caffeine Effects on Social Interaction and Recognition in Mice: Involvement of Adenosine A2A and A1 Receptors

    PubMed Central

    López-Cruz, Laura; San-Miguel, Noemí; Bayarri, Pilar; Baqi, Younis; Müller, Christa E.; Salamone, John D.; Correa, Mercé

    2016-01-01

    Ethanol and caffeine are frequently consumed in combination and have opposite effects on the adenosine system: ethanol metabolism leads to an increase in adenosine levels, while caffeine is a non-selective adenosine A1/A2A receptor antagonist. These receptors are highly expressed in striatum and olfactory tubercle, brain areas involved in exploration and social interaction in rodents. Ethanol modulates social interaction processes, but the role of adenosine in social behavior is still poorly understood. The present work was undertaken to study the impact of ethanol, caffeine and their combination on social behavior, and to explore the involvement of A1 and A2A receptors on those actions. Male CD1 mice were evaluated in a social interaction three-chamber paradigm, for preference of conspecific vs. object, and also for long-term recognition memory of familiar vs. novel conspecific. Ethanol showed a biphasic effect, with low doses (0.25 g/kg) increasing social contact and higher doses (1.0–1.5 g/kg) reducing social interaction. However, no dose changed social preference; mice always spent more time sniffing the conspecific than the object, independently of the ethanol dose. Ethanol, even at doses that did not change social exploration, produced amnestic effects on social recognition the following day. Caffeine reduced social contact (15.0–60.0 mg/kg), and even blocked social preference at higher doses (30.0–60.0 mg/kg). The A1 antagonist Cyclopentyltheophylline (CPT; 3–9 mg/kg) did not modify social contact or preference on its own, and the A2A antagonist MSX-3 (1.5–6 mg/kg) increased social interaction at all doses. Ethanol at intermediate doses (0.5–1.0 g/kg) was able to reverse the reduction in social exploration induced by caffeine (15.0–30.0 mg/kg). Although there was no interaction between ethanol and CPT or MSX-3 on social exploration in the first day, MSX-3 blocked the amnestic effects of ethanol observed on the following day. Thus, ethanol

  19. Kinetic interaction analysis of human interleukin 5 receptor alpha mutants reveals a unique binding topology and charge distribution for cytokine recognition.

    PubMed

    Ishino, Tetsuya; Pasut, Gianfranco; Scibek, Jeffery; Chaiken, Irwin

    2004-03-05

    Human interleukin 5 receptor alpha (IL5Ralpha) comprises three fibronectin type III domains (D1, D2, and D3) in the extracellular region. Previous results have indicated that residues in the D1D2 domains are crucial for high affinity interaction with human interleukin 5 (IL5). Yet, it is the D2D3 domains that have sequence homology with the classic cytokine recognition motif that is generally assumed to be the minimum cytokine-recognizing unit. In the present study, we used kinetic interaction analysis of alanine-scanning mutational variants of IL5Ralpha to define the residues involved in IL5 recognition. Soluble forms of IL5Ralpha variants were expressed in S2 cells, selectively captured via their C-terminal V5 tag by anti-V5 tag antibody immobilized onto the sensor chip and examined for IL5 interaction by using a sandwich surface plasmon resonance biosensor method. Marked effects on the interaction kinetics were observed not only in D1 (Asp(55), Asp(56), and Glu(58)) and D2 (Lys(186) and Arg(188)) domains, but also in the D3 (Arg(297)) domain. Modeling of the tertiary structure of IL5Ralpha indicated that these binding residues fell into two clusters. The first cluster consists of D1 domain residues that form a negatively charged patch, whereas the second cluster consists of residues that form a positively charged patch at the interface of D2 and D3 domains. These results suggest that the IL5 x IL5Ralpha system adopts a unique binding topology, in which the cytokine is recognized by a D2D3 tandem domain combined with a D1 domain, to form an extended cytokine recognition interface.

  20. Hemispherand-Strapped Calix[4]pyrrole: An Ion-pair Receptor for the Recognition and Extraction of Lithium Nitrite.

    PubMed

    He, Qing; Zhang, Zhan; Brewster, James T; Lynch, Vincent M; Kim, Sung Kuk; Sessler, Jonathan L

    2016-08-10

    The hemispherand-strapped calix[4]pyrrole (1) acts as an ion pair receptor that exhibits selectivity for lithium salts. In organic media (CD2Cl2 and CD3OD, v/v, 9:1), receptor 1 binds LiCl with high preference relative to NaCl, KCl, and RbCl. DFT calculations provided support for the observed selectivity. Single crystal structures of five different lithium ion-pair complexes of 1 were obtained. In the case of LiCl, a single bridging water molecule between the lithium cation and chloride anion was observed, while tight contact ion pairs were observed in the case of the LiBr, LiI, LiNO3, and LiNO2 salts. Receptor 1 proved effective as an extractant for LiNO2 under both model solid-liquid and liquid-liquid extraction conditions.

  1. Differentiation of human peripheral blood Vδ1+ T cells expressing the natural cytotoxicity receptor NKp30 for recognition of lymphoid leukemia cells.

    PubMed

    Correia, Daniel V; Fogli, Manuela; Hudspeth, Kelly; da Silva, Maria Gomes; Mavilio, Domenico; Silva-Santos, Bruno

    2011-07-28

    The success of cancer immunotherapy depends on productive tumor cell recognition by killer lymphocytes. γδ T cells are a population of innate-like lymphocytes endowed with strong, MHC-unrestricted cytotoxicity against tumor cells. This notwithstanding, we recently showed that a large proportion of human hematologic tumors is resistant to γδ peripheral blood lymphocytes (PBLs) activated with specific agonists to the highly prevalent Vγ9Vδ2 TCR. Although this probably constitutes an important limitation to current γδ T cell-mediated immunotherapy strategies, we describe here the differentiation of a novel subset of Vδ2(-) Vδ1(+) PBLs expressing natural cytotoxicity receptors (NCRs) that directly mediate killing of leukemia cell lines and chronic lymphocytic leukemia patient neoplastic cells. We show that Vδ1(+) T cells can be selectively induced to express NKp30, NKp44 and NKp46, through a process that requires functional phosphatidylinositol 3-kinase (PI-3K)/AKT signaling on stimulation with γ(c) cytokines and TCR agonists. The stable expression of NCRs is associated with high levels of granzyme B and enhanced cytotoxicity against lymphoid leukemia cells. Specific gain-of-function and loss-of-function experiments demonstrated that NKp30 makes the most important contribution to TCR-independent leukemia cell recognition. Thus, NKp30(+) Vδ1(+) T cells constitute a novel, inducible and specialized killer lymphocyte population with high potential for immunotherapy of human cancer.

  2. Duox2 is required for the transcription of pattern recognition receptors in acute viral lung infection: An interferon-independent regulatory mechanism.

    PubMed

    Hong, Seung-No; Kim, Ji Young; Kim, Hanna; Kim, Dong-Young; Won, Tae-Bin; Han, Doo Hee; Rhee, Chae-Seo; Kim, Hyun Jik

    2016-10-01

    The innate immune response, which constitutes the first line of defense against influenza A virus (IAV) infection, is activated by pattern recognition receptors (PRRs) that recognize viral structures. We found that the PRRs, retinoic acid-inducible gene 1 (RIG-I) and melanoma differentiation-associated protein 5 (MDA5), which have been implicated as interferon (IFN)-stimulated genes, were dominantly responsible for the recognition of IAV in lungs of mice at 3 and 7 days post infection (dpi). Intranasal administration of IFNs enhanced RIG-I and MDA5 gene expression after IAV infection and mRNA levels of RIG-I and MDA5 were significantly reduced at 7 dpi in mice with neutralization of secreted IFNs. However, blockade of IFNs did not alter the transcription of RIG-I and MDA5 at 3 dpi. We studied the antiviral effect of Duox2 in vivo lung to elucidate the role of Duox2 in respiratory mucosa. RIG-I and MDA5 mRNA levels were induced to a lower extent in lungs of mice that were inoculated with Duox2 small hairpin RNA regardless of secreted IFNs at 3 dpi. We propose that Duox2 is responsible for IFN-independent signaling for induction of PRRs transcription and can control acute IAV lung infection at the beginning of infection.

  3. Isolation from Gluconacetobacter diazotrophicus cell walls of specific receptors for sugarcane glycoproteins, which act as recognition factors.

    PubMed

    Blanco, Y; Arroyo, M; Legaz, M E; Vicente, C

    2005-11-04

    Glycoproteins from sugarcane stalks have been isolated from plants field-grown by size-exclusion chromatography. Some of these glycoproteins, previously labelled with fluorescein isothiocyanate, are able to bind to the cell wall of the sugarcane endophyte, N2-fixing Gluconacetobacter diazotrophicus, and largely removed after washing the bacterial cells with sucrose. This implies that sugarcane glycoproteins use beta-(1-->2)-fructofuranosyl fructose domains in their glycosidic moiety to bind to specific receptors in the bacterial cell walls. These receptors have been isolated by affinity chromatography on a sugarcane glycoprotein-agarose matrix, desorbed with sucrose and characterized by sodium dodecyl sulfate polyacrylamide gel electrophresisand capillary electrophoresis (CE).

  4. The Glycophosphatidylinositol Anchor of the MCMV Evasin, m157, Facilitates Optimal Cell Surface Expression and Ly49 Receptor Recognition

    PubMed Central

    Carlin, Lindsey E.; Guseva, Natalya V.; Shey, Michael R.; Ballas, Zuhair K.; Heusel, Jonathan W.

    2013-01-01

    The murine cytomegalovirus-encoded protein m157 is a cognate ligand for both inhibitory and activating receptors expressed by natural killer cells. Additionally, m157 is expressed on the surface of infected cells by a glycophosphatidylinositol (GPI) anchor. Although endogenous GPI-anchored proteins are known to be ligands for the NK cell receptor, NKG2D, the contribution of the GPI anchor for viral m157 ligand function is unknown. To determine whether the GPI anchor for m157 is dispensable for m157 function, we generated m157 variants expressed as transmembrane fusion proteins and tested cells expressing transmembrane m157 for the capacity to activate cognate Ly49 receptors. We found that the GPI anchor is required for high-level cell surface expression of m157, and that the transmembrane m157 ligand retains the capacity to activate reporter cells and NK cells expressing Ly49H, as well as Ly49I129 reporter cells, but with reduced potency. Importantly, target cells expressing the transmembrane form of m157 were killed less efficiently and failed to mediate Ly49H receptor downregulation on fresh NK cells compared to targets expressing GPI-anchored m157. Taken together, these results show that the GPI anchor for m157 facilitates robust cell surface expression, and that NK cells are sensitive to the altered cell surface expression of this potent viral evasin. PMID:23840655

  5. Multichannel HSO4- recognition promoted by a bound cation within a ferrocene-based ion pair receptor.

    PubMed

    Alfonso, María; Espinosa, Arturo; Tárraga, Alberto; Molina, Pedro

    2012-07-11

    A ferrocene-based ion pair receptor is shown only to recognise HSO(4)(-) anions in the presence of a cobound Pb(2+) or Zn(2+) cation guest species through a perturbation of the redox potential of the ferrocene unit and a remarkable enhancement of the fluorescence.

  6. Recognition of the Major Histocompatibility Complex (MHC) Class Ib Molecule H2-Q10 by the Natural Killer Cell Receptor Ly49C.

    PubMed

    Sullivan, Lucy C; Berry, Richard; Sosnin, Natasha; Widjaja, Jacqueline M L; Deuss, Felix A; Balaji, Gautham R; LaGruta, Nicole L; Mirams, Michiko; Trapani, Joseph A; Rossjohn, Jamie; Brooks, Andrew G; Andrews, Daniel M

    2016-09-02

    Murine natural killer (NK) cells are regulated by the interaction of Ly49 receptors with major histocompatibility complex class I molecules (MHC-I). Although the ligands for inhibitory Ly49 were considered to be restricted to classical MHC (MHC-Ia), we have shown that the non-classical MHC molecule (MHC-Ib) H2-M3 was a ligand for the inhibitory Ly49A. Here we establish that another MHC-Ib, H2-Q10, is a bona fide ligand for the inhibitory Ly49C receptor. H2-Q10 bound to Ly49C with a marginally lower affinity (∼5 μm) than that observed between Ly49C and MHC-Ia (H-2K(b)/H-2D(d), both ∼1 μm), and this recognition could be prevented by cis interactions with H-2K in situ To understand the molecular details underpinning Ly49·MHC-Ib recognition, we determined the crystal structures of H2-Q10 and Ly49C bound H2-Q10. Unliganded H2-Q10 adopted a classical MHC-I fold and possessed a peptide-binding groove that exhibited features similar to those found in MHC-Ia, explaining the diverse peptide binding repertoire of H2-Q10. Ly49C bound to H2-Q10 underneath the peptide binding platform to a region that encompassed residues from the α1, α2, and α3 domains, as well as the associated β2-microglobulin subunit. This docking mode was conserved with that previously observed for Ly49C·H-2K(b) Indeed, structure-guided mutation of Ly49C indicated that Ly49C·H2-Q10 and Ly49C·H-2K(b) possess similar energetic footprints focused around residues located within the Ly49C β4-stand and L5 loop, which contact the underside of the peptide-binding platform floor. Our data provide a structural basis for Ly49·MHC-Ib recognition and demonstrate that MHC-Ib represent an extended family of ligands for Ly49 molecules.

  7. The endocytic receptor protein LRP also mediates neuronal calcium signaling via N-methyl-d-aspartate receptors

    PubMed Central

    Bacskai, B. J.; Xia, M. Q.; Strickland, D. K.; Rebeck, G. W.; Hyman, B. T.

    2000-01-01

    The low density lipoprotein receptor-related protein (LRP) is an endocytic receptor that is a member of the low density lipoprotein receptor family. We report that the LRP ligand, activated α2-macroglobulin (α2M*), induces robust calcium influx in cultured primary neurons, but not in nonneuronal LRP-containing cells in the same culture. The calcium influx is mediated through N-methyl-d-aspartate receptor channels, which explains the neuron specificity of the response. Microapplication of α2M* leads to a localized response at the site of application that dissipates rapidly, suggesting that the calcium signal is temporally and spatially discrete. Calcium influx to α2M* is blocked by the physiological LRP inhibitor, receptor-associated protein. Bivalent antibodies to the extracellular domain of LRP, but not Fab fragments of the same antibody, cause calcium influx, indicating that the response is specific to LRP and may require dimerization of the receptor. Thus, LRP is an endocytic receptor with a novel signaling role. PMID:11016955

  8. IL-22 is rapidly induced by Pathogen Recognition Receptors Stimulation in Bone-Marrow-derived Dendritic Cells in the Absence of IL-23.

    PubMed

    Fumagalli, Silvia; Torri, Anna; Papagna, Angela; Citterio, Stefania; Mainoldi, Federica; Foti, Maria

    2016-09-22

    In vertebrates, microorganisms are recognized by pathogen recognition receptors (PRRs). Exposure of immune cells to the ligands of these receptors activates intracellular signaling cascades that rapidly induce the expression of a variety of genes. Within these genes, the cytokines family plays a crucial function because of its role in adaptive immunity induction and in tissue-specific functional regulation, such as tissue repair and tissue homeostasis during steady state conditions. Within the myeloid compartment, dendritic cells (DCs) release a variety of inflammatory cytokines in response to microbes. In this study, we show that BMDCs release IL-22 directly upon PRRs activation without the need of IL-23 signaling as reported for other IL22-producing cells. Moreover, we demonstrate that cytokine IL-22 is rapidly released in a cell-specific manner as macrophages are not able to produce IL-22 through the same PRRs system. In addition, we characterize the intracellular signaling cascade required for IL-22 release in BMDCs. Myd88, MEK1/2, NFkb and AhR, but not p38, NFAT, and RORgt, were found to be involved in IL-22 regulation in DCs. Our study suggests that BMDCs possess a unique intracellular molecular plasticity which, once activated, directs different BMDCs functions in a cell-specific manner.

  9. IL-22 is rapidly induced by Pathogen Recognition Receptors Stimulation in Bone-Marrow-derived Dendritic Cells in the Absence of IL-23

    PubMed Central

    Fumagalli, Silvia; Torri, Anna; Papagna, Angela; Citterio, Stefania; Mainoldi, Federica; Foti, Maria

    2016-01-01

    In vertebrates, microorganisms are recognized by pathogen recognition receptors (PRRs). Exposure of immune cells to the ligands of these receptors activates intracellular signaling cascades that rapidly induce the expression of a variety of genes. Within these genes, the cytokines family plays a crucial function because of its role in adaptive immunity induction and in tissue-specific functional regulation, such as tissue repair and tissue homeostasis during steady state conditions. Within the myeloid compartment, dendritic cells (DCs) release a variety of inflammatory cytokines in response to microbes. In this study, we show that BMDCs release IL-22 directly upon PRRs activation without the need of IL-23 signaling as reported for other IL22-producing cells. Moreover, we demonstrate that cytokine IL-22 is rapidly released in a cell-specific manner as macrophages are not able to produce IL-22 through the same PRRs system. In addition, we characterize the intracellular signaling cascade required for IL-22 release in BMDCs. Myd88, MEK1/2, NFkb and AhR, but not p38, NFAT, and RORgt, were found to be involved in IL-22 regulation in DCs. Our study suggests that BMDCs possess a unique intracellular molecular plasticity which, once activated, directs different BMDCs functions in a cell-specific manner. PMID:27652524

  10. The Ile13 residue of microcin J25 is essential for recognition by the receptor FhuA, but not by the inner membrane transporter SbmA.

    PubMed

    Socias, Sergio B; Severinov, Konstantin; Salomon, Raul A

    2009-11-01

    Entry of the peptide antibiotic microcin J25 (MccJ25) into target cells is mediated by the outer membrane receptor FhuA and the inner membrane protein SbmA. The latter also transports MccB17 into the cell cytoplasm. Comparison of MccJ25 and MccB17 revealed a tetrapeptide sequence (VGIG) common to both antibiotics. We speculated that this structural feature in MccJ25 could be a motif recognized by SbmA. To test this hypothesis, we used a MccJ25 variant in which the isoleucine in VGIG (position 13 in the MccJ25 sequence) was replaced by lysine (I13K). In experiments in which the FhuA receptor was bypassed, the substituted microcin showed an inhibitory activity similar to that of the wild-type peptide. Moreover, MccJ25 interfered with colicin M uptake by FhuA in a competition assay, while the I13K mutant did not. From these results, we propose that the Ile(13) residue is only required for interaction with FhuA, and that VGIG is not a major recognition element by SbmA.

  11. Structure-Based Mutagenesis of the Substrate-Recognition Domain of Nrdp1/FLRF Identifies the Binding Site for the Receptor Tyrosine Kinase ErbB3

    SciTech Connect

    Bouyain,S.; Leahy, D.

    2007-01-01

    The E3 ubiquitin ligase neuregulin receptor degrading protein 1 (Nrdp1) mediates the ligand-independent degradation of the epidermal growth factor receptor family member ErbB3/HER3. By regulating cellular levels of ErbB3, Nrdp1 influences ErbB3-mediated signaling, which is essential for normal vertebrate development. Nrdp1 belongs to the tripartite or RBCC (RING, B-box, coiled-coil) family of ubiquitin ligases in which the RING domain is responsible for ubiquitin ligation and a variable C-terminal region mediates substrate recognition. We report here the 1.95 A crystal structure of the C-terminal domain of Nrdp1 and show that this domain is sufficient to mediate ErbB3 binding. Furthermore, we have used site-directed mutagenesis to map regions of the Nrdp1 surface that are important for interacting with ErbB3 and mediating its degradation in transfected cells. The ErbB3-binding site localizes to a region of Nrdp1 that is conserved from invertebrates to vertebrates, in contrast to ErbB3, which is only found in vertebrates. This observation suggests that Nrdp1 uses a common binding site to recognize its targets in different species.

  12. Structure-based mutagenesis of the substrate-recognition domain of Nrdp1/FLRF identifies the binding site for the receptor tyrosine kinase ErbB3

    PubMed Central

    Bouyain, Samuel; Leahy, Daniel J.

    2007-01-01

    The E3 ubiquitin ligase neuregulin receptor degrading protein 1 (Nrdp1) mediates the ligand-independent degradation of the epidermal growth factor receptor family member ErbB3/HER3. By regulating cellular levels of ErbB3, Nrdp1 influences ErbB3-mediated signaling, which is essential for normal vertebrate development. Nrdp1 belongs to the tripartite or RBCC (RING, B-box, coiled-coil) family of ubiquitin ligases in which the RING domain is responsible for ubiquitin ligation and a variable C-terminal region mediates substrate recognition. We report here the 1.95 Å crystal structure of the C-terminal domain of Nrdp1 and show that this domain is sufficient to mediate ErbB3 binding. Furthermore, we have used site-directed mutagenesis to map regions of the Nrdp1 surface that are important for interacting with ErbB3 and mediating its degradation in transfected cells. The ErbB3-binding site localizes to a region of Nrdp1 that is conserved from invertebrates to vertebrates, in contrast to ErbB3, which is only found in vertebrates. This observation suggests that Nrdp1 uses a common binding site to recognize its targets in different species. PMID:17384230

  13. Structure-based mutagenesis of the substrate-recognition domain of Nrdp1/FLRF identifies the binding site for the receptor tyrosine kinase ErbB3.

    PubMed

    Bouyain, Samuel; Leahy, Daniel J

    2007-04-01

    The E3 ubiquitin ligase neuregulin receptor degrading protein 1 (Nrdp1) mediates the ligand-independent degradation of the epidermal growth factor receptor family member ErbB3/HER3. By regulating cellular levels of ErbB3, Nrdp1 influences ErbB3-mediated signaling, which is essential for normal vertebrate development. Nrdp1 belongs to the tripartite or RBCC (RING, B-box, coiled-coil) family of ubiquitin ligases in which the RING domain is responsible for ubiquitin ligation and a variable C-terminal region mediates substrate recognition. We report here the 1.95 A crystal structure of the C-terminal domain of Nrdp1 and show that this domain is sufficient to mediate ErbB3 binding. Furthermore, we have used site-directed mutagenesis to map regions of the Nrdp1 surface that are important for interacting with ErbB3 and mediating its degradation in transfected cells. The ErbB3-binding site localizes to a region of Nrdp1 that is conserved from invertebrates to vertebrates, in contrast to ErbB3, which is only found in vertebrates. This observation suggests that Nrdp1 uses a common binding site to recognize its targets in different species.

  14. Substrate recognition by the Lyn protein-tyrosine kinase. NMR structure of the immunoreceptor tyrosine-based activation motif signaling region of the B cell antigen receptor.

    PubMed

    Gaul, B S; Harrison, M L; Geahlen, R L; Burton, R A; Post, C B

    2000-05-26

    The immunoreceptor tyrosine-based activation motif (ITAM) plays a central role in transmembrane signal transduction in hematopoietic cells by mediating responses leading to proliferation and differentiation. An initial signaling event following activation of the B cell antigen receptor is phosphorylation of the CD79a (Ig-alpha) ITAM by Lyn, a Src family protein-tyrosine kinase. To elucidate the structural basis for recognition between the ITAM substrate and activated Lyn kinase, the structure of an ITAM-derived peptide bound to Lyn was determined using exchange-transferred nuclear Overhauser NMR spectroscopy. The bound substrate structure has an irregular helix-like character. Docking based on the NMR data into the active site of the closely related Lck kinase strongly favors ITAM binding in an orientation similar to binding of cyclic AMP-dependent protein kinase rather than that of insulin receptor tyrosine kinase. The model of the complex provides a rationale for conserved ITAM residues, substrate specificity, and suggests that substrate binds only the active conformation of the Src family tyrosine kinase, unlike the ATP cofactor, which can bind the inactive form.

  15. Discovery of a novel site of opioid action at the innate immune pattern-recognition receptor TLR4 and its role in addiction.

    PubMed

    Jacobsen, Jonathan Henry W; Watkins, Linda R; Hutchinson, Mark R

    2014-01-01

    Opioids have historically, and continue to be, an integral component of pain management. However, despite pharmacokinetic and dynamic optimization over the past 100 years, opioids continue to produce many undesirable side effects such as tolerance, reward, and dependence. As such, opioids are liable for addiction. Traditionally, opioid addiction was viewed as a solely neuronal process, and while substantial headway has been made into understanding the molecular and cellular mechanisms mediating this process, research has however, been relatively ambivalent to how the rest of the central nervous system (CNS) responds to opioids. Evidence over the past 20 years has clearly demonstrated the importance of the immunocompetent cells of the CNS (glia) in many aspects of opioid pharmacology. Particular focus has been placed on microglia and astrocytes, who in response to opioids, become activated and release inflammatory mediators. Importantly, the mechanism underlying immune activation is beginning to be elucidated. Evidence suggests an innate immune pattern-recognition receptor (toll-like receptor 4) as an integral component underlying opioid-induced glial activation. The subsequent proinflammatory response may be viewed akin to neurotransmission creating a process termed central immune signaling. Translationally, we are beginning to appreciate the importance of central immune signaling as it contributes to many behavioral actions of addiction including reward, withdrawal, and craving. As such, the aim of this chapter is to review and integrate the neuronal and central immune signaling perspective of addiction.

  16. Molecular basis of lipo-chitooligosaccharide recognition by the lysin motif receptor-like kinase LYR3 in legumes.

    PubMed

    Malkov, Nikita; Fliegmann, Judith; Rosenberg, Charles; Gasciolli, Virginie; Timmers, Antonius C J; Nurisso, Alessandra; Cullimore, Julie; Bono, Jean-Jacques

    2016-05-15

    LYR3 [LysM (lysin motif) receptor-like kinase 3] of Medicago truncatula is a high-affinity binding protein for symbiotic LCO (lipo-chitooligosaccharide) signals, produced by rhizobia bacteria and arbuscular mycorrhizal fungi. The present study shows that LYR3 from several other legumes, but not from two Lupinus species which are incapable of forming the mycorrhizal symbiosis, bind LCOs with high affinity and discriminate them from COs (chitooligosaccharides). The biodiversity of these proteins and the lack of binding to the Lupinus proteins were used to identify features required for high-affinity LCO binding. Swapping experiments between each of the three LysMs of the extracellular domain of the M. truncatula and Lupinus angustifolius LYR3 proteins revealed the crucial role of the third LysM in LCO binding. Site-directed mutagenesis identified a tyrosine residue, highly conserved in all LYR3 LCO-binding proteins, which is essential for high-affinity binding. Molecular modelling suggests that it may be part of a hydrophobic tunnel able to accommodate the LCO acyl chain. The lack of conservation of these features in the binding site of plant LysM proteins binding COs provides a mechanistic explanation of how LCO recognition might differ from CO perception by structurally related LysM receptors.

  17. Subtle Changes in Peptide Conformation Profoundly Affect Recognition of the Non-Classical MHC Class I Molecule HLA-E by the CD94-NKG2 Natural Killer Cell Receptors

    SciTech Connect

    Hoare, Hilary L; Sullivan, Lucy C; Clements, Craig S; Ely, Lauren K; Beddoe, Travis; Henderson, Kate N; Lin, Jie; Reid, Hugh H; Brooks, Andrew G; Rossjohn, Jamie

    2008-03-31

    Human leukocyte antigen (HLA)-E is a non-classical major histocompatibility complex class I molecule that binds peptides derived from the leader sequences of other HLA class I molecules. Natural killer cell recognition of these HLA-E molecules, via the CD94-NKG2 natural killer family, represents a central innate mechanism for monitoring major histocompatibility complex expression levels within a cell. The leader sequence-derived peptides bound to HLA-E exhibit very limited polymorphism, yet subtle differences affect the recognition of HLA-E by the CD94-NKG2 receptors. To better understand the basis for this peptide-specific recognition, we determined the structure of HLA-E in complex with two leader peptides, namely, HLA-Cw*07 (VMAPRALLL), which is poorly recognised by CD94-NKG2 receptors, and HLA-G*01 (VMAPRTLFL), a high-affinity ligand of CD94-NKG2 receptors. A comparison of these structures, both of which were determined to 2.5-Å resolution, revealed that allotypic variations in the bound leader sequences do not result in conformational changes in the HLA-E heavy chain, although subtle changes in the conformation of the peptide within the binding groove of HLA-E were evident. Accordingly, our data indicate that the CD94-NKG2 receptors interact with HLA-E in a manner that maximises the ability of the receptors to discriminate between subtle changes in both the sequence and conformation of peptides bound to HLA-E.

  18. Functional recognition of a distinct receptor preferential for leukotriene E4 in mice lacking the cysteinyl leukotriene 1 and 2 receptors.

    PubMed

    Maekawa, Akiko; Kanaoka, Yoshihide; Xing, Wei; Austen, K Frank

    2008-10-28

    The cysteinyl leukotrienes (cys-LTs) are a family of potent lipid mediators of inflammation derived from arachidonic acid. Activation of certain cell types results in the biosynthesis and export of leukotriene (LT) C(4), which then undergoes extracellular metabolism to LTD(4) and LTE(4). LTE(4), the most stable cys-LT, is only a weak agonist for the defined type 1 and type 2 cys-LT receptors (CysLT(1)R and CysLT(2)R, respectively). We had recognized a greater potency for LTE(4) than LTC(4) or LTD(4) in constricting guinea pig trachea in vitro and comparable activity in eliciting a cutaneous wheal and flare response in humans. Thus, we hypothesized that a vascular permeability response to LTE(4) in mice lacking both the CysLT(1)R and CysLT(2)R could establish the existence of a separate LTE(4) receptor. We now report that the intradermal injection of LTE(4) into the ear of mice deficient in both CysLT(1)R and CysLT(2)R elicits a vascular leak that exceeds the response to intradermal injection of LTC(4) or LTD(4), and that this response is inhibited by pretreatment of the mice with pertussis toxin or a Rho kinase inhibitor. LTE(4) is approximately 64-fold more potent in the CysLT(1)R/CysLT(2)R double-deficient mice than in sufficient mice. The administration of a CysLT(1)R antagonist augmented the permeability response of the CysLT(1)R/CysLT(2)R double-deficient mice to LTC(4), LTD(4), and LTE(4). Our findings establish the existence of a third receptor, CysLT(E)R, that responds preferentially to LTE(4), the most abundant cys-LT in biologic fluids, and thus reveal a new target for therapeutic intervention.

  19. Blonanserin ameliorates phencyclidine-induced visual-recognition memory deficits: the complex mechanism of blonanserin action involving D₃-5-HT₂A and D₁-NMDA receptors in the mPFC.

    PubMed

    Hida, Hirotake; Mouri, Akihiro; Mori, Kentaro; Matsumoto, Yurie; Seki, Takeshi; Taniguchi, Masayuki; Yamada, Kiyofumi; Iwamoto, Kunihiro; Ozaki, Norio; Nabeshima, Toshitaka; Noda, Yukihiro

    2015-02-01

    Blonanserin differs from currently used serotonin 5-HT₂A/dopamine-D₂ receptor antagonists in that it exhibits higher affinity for dopamine-D₂/₃ receptors than for serotonin 5-HT₂A receptors. We investigated the involvement of dopamine-D₃ receptors in the effects of blonanserin on cognitive impairment in an animal model of schizophrenia. We also sought to elucidate the molecular mechanism underlying this involvement. Blonanserin, as well as olanzapine, significantly ameliorated phencyclidine (PCP)-induced impairment of visual-recognition memory, as demonstrated by the novel-object recognition test (NORT) and increased extracellular dopamine levels in the medial prefrontal cortex (mPFC). With blonanserin, both of these effects were antagonized by DOI (a serotonin 5-HT₂A receptor agonist) and 7-OH-DPAT (a dopamine-D₃ receptor agonist), whereas the effects of olanzapine were antagonized by DOI but not by 7-OH-DPAT. The ameliorating effect was also antagonized by SCH23390 (a dopamine-D₁ receptor antagonist) and H-89 (a protein kinase A (PKA) inhibitor). Blonanserin significantly remediated the decrease in phosphorylation levels of PKA at Thr(197) and of NR1 (an essential subunit of N-methyl-D-aspartate (NMDA) receptors) at Ser(897) by PKA in the mPFC after a NORT training session in the PCP-administered mice. There were no differences in the levels of NR1 phosphorylated at Ser(896) by PKC in any group. These results suggest that the ameliorating effect of blonanserin on PCP-induced cognitive impairment is associated with indirect functional stimulation of the dopamine-D₁-PKA-NMDA receptor pathway following augmentation of dopaminergic neurotransmission due to inhibition of both dopamine-D₃ and serotonin 5-HT₂A receptors in the mPFC.

  20. Blonanserin Ameliorates Phencyclidine-Induced Visual-Recognition Memory Deficits: the Complex Mechanism of Blonanserin Action Involving D3-5-HT2A and D1-NMDA Receptors in the mPFC

    PubMed Central

    Hida, Hirotake; Mouri, Akihiro; Mori, Kentaro; Matsumoto, Yurie; Seki, Takeshi; Taniguchi, Masayuki; Yamada, Kiyofumi; Iwamoto, Kunihiro; Ozaki, Norio; Nabeshima, Toshitaka; Noda, Yukihiro

    2015-01-01

    Blonanserin differs from currently used serotonin 5-HT2A/dopamine-D2 receptor antagonists in that it exhibits higher affinity for dopamine-D2/3 receptors than for serotonin 5-HT2A receptors. We investigated the involvement of dopamine-D3 receptors in the effects of blonanserin on cognitive impairment in an animal model of schizophrenia. We also sought to elucidate the molecular mechanism underlying this involvement. Blonanserin, as well as olanzapine, significantly ameliorated phencyclidine (PCP)-induced impairment of visual-recognition memory, as demonstrated by the novel-object recognition test (NORT) and increased extracellular dopamine levels in the medial prefrontal cortex (mPFC). With blonanserin, both of these effects were antagonized by DOI (a serotonin 5-HT2A receptor agonist) and 7-OH-DPAT (a dopamine-D3 receptor agonist), whereas the effects of olanzapine were antagonized by DOI but not by 7-OH-DPAT. The ameliorating effect was also antagonized by SCH23390 (a dopamine-D1 receptor antagonist) and H-89 (a protein kinase A (PKA) inhibitor). Blonanserin significantly remediated the decrease in phosphorylation levels of PKA at Thr197 and of NR1 (an essential subunit of N-methyl-D-aspartate (NMDA) receptors) at Ser897 by PKA in the mPFC after a NORT training session in the PCP-administered mice. There were no differences in the levels of NR1 phosphorylated at Ser896 by PKC in any group. These results suggest that the ameliorating effect of blonanserin on PCP-induced cognitive impairment is associated with indirect functional stimulation of the dopamine-D1-PKA-NMDA receptor pathway following augmentation of dopaminergic neurotransmission due to inhibition of both dopamine-D3 and serotonin 5-HT2A receptors in the mPFC. PMID:25120077

  1. Neonatal paternal deprivation impairs social recognition and alters levels of oxytocin and estrogen receptor α mRNA expression in the MeA and NAcc, and serum oxytocin in mandarin voles.

    PubMed

    Cao, Yan; Wu, Ruiyong; Tai, Fadao; Zhang, Xia; Yu, Peng; An, Xiaolei; Qiao, Xufeng; Hao, Ping

    2014-01-01

    Paternal care is necessary for the healthy development of social behavior in monogamous rodents and social recognition underpins social behavior in these animals. The effects of paternal care on the development of social recognition and underlying neuroendocrine mechanisms, especially the involvement of oxytocin and estrogen pathways, remain poorly understood. We investigated the effects of paternal deprivation (PD: father was removed from neonatal pups and mother alone raised the offspring) on social recognition in mandarin voles (Microtus mandarinus), a socially monogamous rodent. Paternal deprivation was found to inhibit the development of social recognition in female and male offspring according to a habituation-dishabituation paradigm. Paternal deprivation resulted in increased inactivity and reduced investigation during new encounters with other animals. Paternal deprivation reduced oxytocin receptor (OTR) and estrogen receptor α (ERα) mRNA expression in the medial amygdala and nucleus accumbens. Paternal deprivation reduced serum oxytocin (OT) concentration in females, but had no effect on males. Our results provide substantial evidence that paternal deprivation inhibits the development of social recognition in female and male mandarin voles and alters social behavior later in life. This is possibly the result of altered expression of central OTR and ERα and serum OT levels caused by paternal deprivation.

  2. Allosteric modulation of GABA(A) receptor subtypes:effects on visual recognition and visuospatial working memory in rhesus monkeys [corrected].

    PubMed

    Soto, Paul L; Ator, Nancy A; Rallapalli, Sundari K; Biawat, Poonam; Clayton, Terry; Cook, James M; Weed, Michael R

    2013-10-01

    Non-selective positive allosteric modulators (PAMs) of GABAA receptors (GABAARs) are known to impair anterograde memory. The role of the various GABAAR subtypes in the memory-impairing effects of non-selective GABAAR PAMs has not been fully elucidated. The current study assessed, in rhesus monkeys, effects of modulation of α1, α2/3, and α5GABAARs on visual recognition and spatial working memory using delayed matching-to-sample (DMTS) and self-ordered spatial search (SOSS) procedures, respectively. The DMTS procedure (n=8) involved selecting a previously presented 'sample' image from a set of multiple images presented after a delay. The SOSS procedure (n=6) involved touching a number of boxes without repeats. The non-selective GABAAR PAM triazolam and the α1GABAA preferential PAMS zolpidem and zaleplon reduced accuracy in both procedures, whereas the α5GABAA preferential PAMs SH-053-2'F-R-CH3 and SH-053-2'F-S-CH3, and the α2/3GABAA preferential PAM TPA023B were without effects on accuracy or trial completion. The low-efficacy α5GABAAR negative allosteric modulator (NAM) PWZ-029 slightly increased only DMTS accuracy, whereas the high-efficacy α5GABAAR NAMs RY-23 and RY-24 did not affect accuracy under either procedure. Finally, the slopes of the accuracy dose-effect curves for triazolam, zolpidem, and zaleplon increased with box number in the SOSS procedure, but were equivalent across DMTS delays. The present results suggest that (1) α1GABAARs, compared with α2/3 and α5GABAARs, are primarily involved in the impairment, by non-selective GABAAR PAMs, of visual recognition and visuospatial working memory in nonhuman primates; and (2) relative cognitive impairment produced by positive modulation of GABAARs increases with number of locations to be remembered, but not with the delay for remembering.

  3. Therapeutic vaccination with a trivalent T-cell receptor (TCR) peptide vaccine restores deficient FoxP3 expression and TCR recognition in subjects with multiple sclerosis.

    PubMed

    Vandenbark, Arthur A; Culbertson, Nicole E; Bartholomew, Richard M; Huan, Jianya; Agotsch, Marci; LaTocha, Dorian; Yadav, Vijayshree; Mass, Michele; Whitham, Ruth; Lovera, Jesus; Milano, June; Theofan, Georgia; Chou, Yuan K; Offner, Halina; Bourdette, Dennis N

    2008-01-01

    Therapeutic vaccination using T-cell receptor (TCR) peptides from V genes commonly expressed by potentially pathogenic T cells remains an approach of interest for treatment of multiple sclerosis (MS) and other autoimmune diseases. We developed a trivalent TCR vaccine containing complementarity determining region (CDR) 2 peptides from BV5S2, BV6S5 and BV13S1 emulsified in incomplete Freund's adjuvant that reliably induced high frequencies of TCR-specific T cells. To evaluate induction of regulatory T-cell subtypes, immunological and clinical parameters were followed in 23 treatment-naïve subjects with relapsing-remitting or progressive MS who received 12 monthly injections of the trivalent peptide vaccine over 1 year in an open-label study design. Prior to vaccination, subjects had reduced expression of forkhead box (Fox) P3 message and protein, and reduced recognition of the expressed TCR repertoire by TCR-reactive cells compared with healthy control donors. After three or four injections, most vaccinated MS subjects developed high frequencies of circulating interleukin (IL)-10-secreting T cells specific for the injected TCR peptides and significantly enhanced expression of FoxP3 by regulatory T cells present in both 'native' CD4+ CD25+ and 'inducible' CD4+ CD25- peripheral blood mononuclear cells (PBMC). At the end of the trial, PBMC from vaccinated MS subjects retained or further increased FoxP3 expression levels, exhibited significantly enhanced recognition of the TCR V gene repertoire apparently generated by perturbation of the TCR network, and significantly suppressed neuroantigen but not recall antigen responses. These findings demonstrate that therapeutic vaccination using only three commonly expressed BV gene determinants can induce an expanded immunoregulatory network in vivo that may optimally control complex autoreactive responses that characterize the inflammatory phase of MS.

  4. Structural basis for antibody recognition in the receptor-binding domains of toxins A and B from Clostridium difficile.

    PubMed

    Murase, Tomohiko; Eugenio, Luiz; Schorr, Melissa; Hussack, Greg; Tanha, Jamshid; Kitova, Elena N; Klassen, John S; Ng, Kenneth K S

    2014-01-24

    Clostridium difficile infection is a serious and highly prevalent nosocomial disease in which the two large, Rho-glucosylating toxins TcdA and TcdB are the main virulence factors. We report for the first time crystal structures revealing how neutralizing and non-neutralizing single-domain antibodies (sdAbs) recognize the receptor-binding domains (RBDs) of TcdA and TcdB. Surprisingly, the complexes formed by two neutralizing antibodies recognizing TcdA do not show direct interference with the previously identified carbohydrate-binding sites, suggesting that neutralization of toxin activity may be mediated by mechanisms distinct from steric blockage of receptor binding. A camelid sdAb complex also reveals the molecular structure of the TcdB RBD for the first time, facilitating the crystallization of a strongly negatively charged protein fragment that has resisted previous attempts at crystallization and structure determination. Electrospray ionization mass spectrometry measurements confirm the stoichiometries of sdAbs observed in the crystal structures. These studies indicate how key epitopes in the RBDs from TcdA and TcdB are recognized by sdAbs, providing molecular insights into toxin structure and function and providing for the first time a basis for the design of highly specific toxin-specific therapeutic and diagnostic agents.

  5. Structural Basis for Antibody Recognition in the Receptor-binding Domains of Toxins A and B from Clostridium difficile*

    PubMed Central

    Murase, Tomohiko; Eugenio, Luiz; Schorr, Melissa; Hussack, Greg; Tanha, Jamshid; Kitova, Elena N.; Klassen, John S.; Ng, Kenneth K. S.

    2014-01-01

    Clostridium difficile infection is a serious and highly prevalent nosocomial disease in which the two large, Rho-glucosylating toxins TcdA and TcdB are the main virulence factors. We report for the first time crystal structures revealing how neutralizing and non-neutralizing single-domain antibodies (sdAbs) recognize the receptor-binding domains (RBDs) of TcdA and TcdB. Surprisingly, the complexes formed by two neutralizing antibodies recognizing TcdA do not show direct interference with the previously identified carbohydrate-binding sites, suggesting that neutralization of toxin activity may be mediated by mechanisms distinct from steric blockage of receptor binding. A camelid sdAb complex also reveals the molecular structure of the TcdB RBD for the first time, facilitating the crystallization of a strongly negatively charged protein fragment that has resisted previous attempts at crystallization and structure determination. Electrospray ionization mass spectrometry measurements confirm the stoichiometries of sdAbs observed in the crystal structures. These studies indicate how key epitopes in the RBDs from TcdA and TcdB are recognized by sdAbs, providing molecular insights into toxin structure and function and providing for the first time a basis for the design of highly specific toxin-specific therapeutic and diagnostic agents. PMID:24311789

  6. Structural Insights into Immune Recognition of the Severe Acute Respiratory Syndrome Coronavirus S Protein Receptor Binding Domain

    SciTech Connect

    Pak, J.; Sharon, C; Satkunarajah, M; Thierry, C; Cameron, C; Kelvin, D; Seetharaman, J; Cochrane, A; Plummer, F; et. al.

    2009-01-01

    The spike (S) protein of the severe acute respiratory syndrome coronavirus (SARS-CoV) is responsible for host cell attachment and fusion of the viral and host cell membranes. Within S the receptor binding domain (RBD) mediates the interaction with angiotensin-converting enzyme 2 (ACE2), the SARS-CoV host cell receptor. Both S and the RBD are highly immunogenic and both have been found to elicit neutralizing antibodies. Reported here is the X-ray crystal structure of the RBD in complex with the Fab of a neutralizing mouse monoclonal antibody, F26G19, elicited by immunization with chemically inactivated SARS-CoV. The RBD-F26G19 Fab complex represents the first example of the structural characterization of an antibody elicited by an immune response to SARS-CoV or any fragment of it. The structure reveals that the RBD surface recognized by F26G19 overlaps significantly with the surface recognized by ACE2 and, as such, suggests that F26G19 likely neutralizes SARS-CoV by blocking the virus-host cell interaction.

  7. Proliferation of thymic stem cells with and without receptors for interleukin 2. Implications for intrathymic antigen recognition

    PubMed Central

    1985-01-01

    We have tested the dividing cells in the mouse thymus for expression of interleukin 2 (IL-2) receptors (IL-2-R) using the rat monoclonal antibody 7D4. A discrete subpopulation of the lymphoblasts clearly expressed IL-2-R at levels comparable to those on mitogen-activated peripheral T cells. This subpopulation, however, represented a small minority of the proliferating cells. IL-2-R-bearing cells were depleted from the PNA+ (peanut agglutinin) lymphoblast population, which contains the direct precursors of most of the cells in the thymus. The majority of receptor-bearing cells were found in the PNA- lymphoblast population, where they constituted only approximately 12% of the cells. Thus, virtually all the PNA+ and most of the PNA- blast cells were in cycle without detectable IL-2-R expression. This indicates that they were not dividing in response to IL-2, and implies that they were not dividing in response to antigen, but rather to novel thymus-specific mitogenic stimuli. On the other hand, the proliferating cells that do express IL-2-R were enriched 4-5-fold in the rapidly growing neonatal thymus, suggesting that they may also play a key role in T cell development. PMID:3921650

  8. Beta-adrenergic receptors link NO/sGC/PKG signaling to BDNF expression during the consolidation of object recognition long-term memory.

    PubMed

    Furini, Cristiane R; Rossato, Janine I; Bitencourt, Lucas L; Medina, Jorge H; Izquierdo, Iván; Cammarota, Martín

    2010-05-01

    The nitric oxide (NO)/soluble guanylyl cyclase (sGC)/protein kinase G (PKG) pathway is important for memory processing, but the identity of its downstream effectors as well as its actual participation in the consolidation of nonaversive declarative long-term memory (LTM) remain unknown. Here, we show that training rats in an object recognition (OR) learning task rapidly increased nitrites/nitrates (NOx) content in the CA1 region of the dorsal hippocampus while posttraining intra-CA1 microinfusion of the neuronal NO synthase (nNOS) inhibitor L-NN hindered OR LTM retention without affecting memory retrieval or other behavioral variables. The amnesic effect of L-NN was not state dependent, was mimicked by the sGC inhibitor LY83583 and the PKG inhibitor KT-5823, and reversed by coinfusion of the NO donor S-nitroso-N-acetylpenicillamine (SNAP) and the PKG activator 8-bromoguanosine 3',5'-cyclic monophosphate (8Br-cGMP). SNAP did not affect the amnesic effect of LY83583 and KT-5823. Conversely, 8Br-cGMP overturned the amnesia induced by LY83583 but not that caused by KT-5823. Intra-CA1 infusion of the beta-adrenergic receptor blocker timolol right after training hindered OR LTM and, although coadministration of noradrenaline reversed the amnesia caused by L-NN, LY83583, and KT5823, the amnesic effect of timolol was unaffected by coinfusion of 8Br-cGMP or SNAP, indicating that hippocampal beta-adrenergic receptors act downstream NO/sGC/PKG signaling. We also found that posttraining intra-CA1 infusion of function-blocking anti-brain-derived neurotrophic factor (BDNF) antibodies hampered OR LTM retention, whereas OR training increased CA1 BDNF levels in a nNOS- and beta-adrenergic receptor-dependent manner. Taken together, our results demonstrate that NO/sGC/PKG signaling in the hippocampus is essential for OR memory consolidation and suggest that beta-adrenergic receptors link the activation of this pathway to BDNF expression during the consolidation of declarative

  9. Facilitated receptor-recognition and enhanced bioactivity of bone morphogenetic protein-2 on magnesium-substituted hydroxyapatite surface

    PubMed Central

    Huang, Baolin; Yuan, Yuan; Li, Tong; Ding, Sai; Zhang, Wenjing; Gu, Yuantong; Liu, Changsheng

    2016-01-01

    Biomaterial surface functionalized with bone morphogenetic protein-2 (BMP-2) is a promising approach to fabricating successful orthopedic implants/scaffolds. However, the bioactivity of BMP-2 on material surfaces is still far from satisfactory and the mechanism of related protein-surface interaction remains elusive. Based on the most widely used bone-implants/scaffolds material, hydroxyapatite (HAP), we developed a matrix of magnesium-substituted HAP (Mg-HAP, 2.2 at% substitution) to address these issues. Further, we investigated the adsorption dynamics, BMPRs-recruitment, and bioactivity of recombinant human BMP-2 (rhBMP-2) on the HAP and Mg-HAP surfaces. To elucidate the mechanism, molecular dynamic simulations were performed to calculate the preferred orientations, conformation changes, and cysteine-knot stabilities of adsorbed BMP-2 molecules. The results showed that rhBMP-2 on the Mg-HAP surface exhibited greater bioactivity, evidenced by more facilitated BMPRs-recognition and higher ALP activity than on the HAP surface. Moreover, molecular simulations indicated that BMP-2 favoured distinct side-on orientations on the HAP and Mg-HAP surfaces. Intriguingly, BMP-2 on the Mg-HAP surface largely preserved the active protein structure evidenced by more stable cysteine-knots than on the HAP surface. These findings explicitly clarify the mechanism of BMP-2-HAP/Mg-HAP interactions and highlight the promising application of Mg-HAP/BMP-2 matrixes in bone regeneration implants/scaffolds. PMID:27075233

  10. Facilitated receptor-recognition and enhanced bioactivity of bone morphogenetic protein-2 on magnesium-substituted hydroxyapatite surface

    NASA Astrophysics Data System (ADS)

    Huang, Baolin; Yuan, Yuan; Li, Tong; Ding, Sai; Zhang, Wenjing; Gu, Yuantong; Liu, Changsheng

    2016-04-01

    Biomaterial surface functionalized with bone morphogenetic protein-2 (BMP-2) is a promising approach to fabricating successful orthopedic implants/scaffolds. However, the bioactivity of BMP-2 on material surfaces is still far from satisfactory and the mechanism of related protein-surface interaction remains elusive. Based on the most widely used bone-implants/scaffolds material, hydroxyapatite (HAP), we developed a matrix of magnesium-substituted HAP (Mg-HAP, 2.2 at% substitution) to address these issues. Further, we investigated the adsorption dynamics, BMPRs-recruitment, and bioactivity of recombinant human BMP-2 (rhBMP-2) on the HAP and Mg-HAP surfaces. To elucidate the mechanism, molecular dynamic simulations were performed to calculate the preferred orientations, conformation changes, and cysteine-knot stabilities of adsorbed BMP-2 molecules. The results showed that rhBMP-2 on the Mg-HAP surface exhibited greater bioactivity, evidenced by more facilitated BMPRs-recognition and higher ALP activity than on the HAP surface. Moreover, molecular simulations indicated that BMP-2 favoured distinct side-on orientations on the HAP and Mg-HAP surfaces. Intriguingly, BMP-2 on the Mg-HAP surface largely preserved the active protein structure evidenced by more stable cysteine-knots than on the HAP surface. These findings explicitly clarify the mechanism of BMP-2-HAP/Mg-HAP interactions and highlight the promising application of Mg-HAP/BMP-2 matrixes in bone regeneration implants/scaffolds.

  11. T Cell Receptor CDR3 Sequence but Not Recognition Characteristics Distinguish Autoreactive Effector and Foxp3+ Regulatory T Cells

    PubMed Central

    Liu, Xin; Nguyen, Phuong; Liu, Wei; Cheng, Cheng; Steeves, Meredith; Obenauer, John C.; Ma, Jing; Geiger, Terrence L.

    2010-01-01

    SUMMARY The source, specificity, and plasticity of the forkhead box transcription factor 3 (Foxp3)+ regulatory T (Treg) and conventional T (Tconv) cell populations active at sites of autoimmune pathology are not well characterized. To evaluate this, we combined global repertoire analyses and functional assessments of isolated T cell receptors (TCR) from TCRα retrogenic mice with autoimmune encephalomyelitis. Treg and Tconv cell TCR repertoires were distinct, and autoantigen-specific Treg and Tconv cells were enriched in diseased tissue. Autoantigen sensitivity and fine specificity of these cells intersected, implying that differences in responsiveness were not responsible for lineage specification. Notably, autoreactive Treg and Tconv cells could be fully distinguished by an acidic versus aliphatic variation at a single TCR CDR3 residue. Our results imply that ontogenically distinct Treg and Tconv cell repertoires with convergent specificities for autoantigen respond during autoimmunity and argue against more than limited plasticity between Treg and Tconv cells during autoimmune inflammation. PMID:20005134

  12. Recognition of CpG oligodeoxynucleotides by human Toll-like receptor 9 and subsequent cytokine induction.

    PubMed

    Suwarti, Suwarti; Yamazaki, Tomohiko; Svetlana, Chechetka; Hanagata, Nobutaka

    2013-01-25

    Toll-like receptor 9 (TLR9) recognizes a synthetic ligand, oligodeoxynucleotide (ODN) containing cytosine-phosphate-guanine (CpG). Activation of TLR9 by CpG ODN induces a signal transduction cascade that plays a pivotal role in first-line immune defense in the human body. The three-dimensional structure of TLR9 has not yet been reported, and the ligand-binding mechanism of TLR9 is still poorly understood; therefore, the mechanism of human TLR9 (hTLR9) ligand binding needs to be elucidated. In this study, we constructed several hTLR9 mutants, including truncated mutants and single mutants in the predicted CpG ODN-binding site. We used these mutants to analyze the role of potential important regions of hTLR9 in receptor signaling induced by phosphorothioate (PTO)-modified CpG ODN and CpG ODNs only consist entirely of a phosphodiester (PD) backbone, CpG ODN2006x3-PD that we developed. We found truncated mutants of hTLR9 lost the signaling activity, indicating that both the C- and N-termini of the extracellular domain (ECD) are necessary for the function of hTLR9. We identified residues, His505, Gln510, His530, and Tyr554, in the C-terminal of hTLR9-ECD that are essential for hTLR9 activation. These residues might form positive charged clusters with which negatively charged CpG ODN could interact. Furthermore, we observed ODN-PD induced interleukin-6 (IL-6) through TLR9 in a CpG-sequence-dependent manner in human peripheral blood mononuclear cells and B cells, whereas ODN-PTO induced IL-6 in a CpG-sequence-independent manner. These finding are relevant for the mechanism of hTLR9 activation by CpG ODNs.

  13. Pattern recognition receptor expression is not impaired in patients with chronic mucocutanous candidiasis with or without autoimmune polyendocrinopathy candidiasis ectodermal dystrophy

    PubMed Central

    Hong, M; Ryan, K R; Arkwright, P D; Gennery, A R; Costigan, C; Dominguez, M; Denning, D W; McConnell, V; Cant, A J; Abinun, M; Spickett, G P; Swan, D C; Gillespie, C S; Young, D A; Lilic, D

    2009-01-01

    Patients with chronic mucocutaneous candidiasis (CMC) have an unknown primary immune defect and are unable to clear infections with the yeast Candida. CMC includes patients with AIRE gene mutations who have autoimmune polyendocrinopathy candidiasis ectodermal dystrophy (APECED), and patients without known mutations. CMC patients have dysregulated cytokine production, suggesting that defective expression of pattern recognition receptors (PRRs) may underlie disease pathogenesis. In 29 patients with CMC (13 with APECED) and controls, we assessed dendritic cell (DC) subsets and monocyte Toll-like receptor (TLR) expression in blood. We generated and stimulated monocyte-derived (mo)DCs with Candida albicans, TLR-2/6 ligand and lipopolysaccharide and assessed PRR mRNA expression by polymerase chain reaction [TLR-1–10, Dectin-1 and -2, spleen tyrosine kinase (Syk) and caspase recruitment domain (CARD) 9] in immature and mature moDCs. We demonstrate for the first time that CMC patients, with or without APECED, have normal blood levels of plasmocytoid and myeloid DCs and monocyte TLR-2/TLR-6 expression. We showed that in immature moDCs, expression levels of all PRRs involved in anti-Candida responses (TLR-1, -2, -4, -6, Dectin-1, Syk, CARD9) were comparable to controls, implying that defects in PRR expression are not responsible for the increased susceptibility to Candida infections seen in CMC patients. However, as opposed to healthy controls, both groups of CMC patients failed to down-regulate PRR mRNA expression in response to Candida, consistent with defective DC maturation, as we reported recently. Thus, impaired DC maturation and consequent altered regulation of PRR signalling pathways rather than defects in PRR expression may be responsible for inadequate Candida handling in CMC patients. PMID:19196253

  14. Epitope Structure of the Carbohydrate Recognition Domain of Asialoglycoprotein Receptor to a Monoclonal Antibody Revealed by High-Resolution Proteolytic Excision Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Stefanescu, Raluca; Born, Rita; Moise, Adrian; Ernst, Beat; Przybylski, Michael

    2011-01-01

    Recent studies suggest that the H1 subunit of the carbohydrate recognition domain (H1CRD) of the asialoglycoprotein receptor is used as an entry site into hepatocytes by hepatitis A and B viruses and Marburg virus. Thus, molecules binding specifically to the CRD might exert inhibition towards these diseases by blocking the virus entry site. We report here the identification of the epitope structure of H1CRD to a monoclonal antibody by proteolytic epitope excision of the immune complex and high-resolution MALDI-FTICR mass spectrometry. As a prerequisite of the epitope determination, the primary structure of the H1CRD antigen was characterised by ESI-FTICR-MS of the intact protein and by LC-MS/MS of tryptic digest mixtures. Molecular mass determination and proteolytic fragments provided the identification of two intramolecular disulfide bridges (seven Cys residues), and a Cys-mercaptoethanol adduct formed by treatment with β-mercaptoethanol during protein extraction. The H1CRD antigen binds to the monoclonal antibody in both native and Cys-alkylated form. For identification of the epitope, the antibody was immobilized on N-hydroxysuccinimide (NHS)-activated Sepharose. Epitope excision and epitope extraction with trypsin and FTICR-MS of affinity-bound peptides provided the identification of two specific epitope peptides (5-16) and (17-23) that showed high affinity to the antibody. Affinity studies of the synthetic epitope peptides revealed independent binding of each peptide to the antibody.

  15. Transgenic expression of the rice Xa21 pattern-recognition receptor in banana (Musa sp.) confers resistance to Xanthomonas campestris pv. musacearum.

    PubMed

    Tripathi, Jaindra N; Lorenzen, Jim; Bahar, Ofir; Ronald, Pamela; Tripathi, Leena

    2014-08-01

    Banana Xanthomonas wilt (BXW), caused by the bacterium Xanthomonas campestris pv. musacearum (Xcm), is the most devastating disease of banana in east and central Africa. The spread of BXW threatens the livelihood of millions of African farmers who depend on banana for food security and income. There are no commercial chemicals, biocontrol agents or resistant cultivars available to control BXW. Here, we take advantage of the robust resistance conferred by the rice pattern-recognition receptor (PRR), XA21, to the rice pathogen Xanthomonas oryzae pv. oryzae (Xoo). We identified a set of genes required for activation of Xa21-mediated immunity (rax) that were conserved in both Xoo and Xcm. Based on the conservation, we hypothesized that intergeneric transfer of Xa21 would confer resistance to Xcm. We evaluated 25 transgenic lines of the banana cultivar 'Gonja manjaya' (AAB) using a rapid bioassay and 12 transgenic lines in the glasshouse for resistance against Xcm. About 50% of the transgenic lines showed complete resistance to Xcm in both assays. In contrast, all of the nontransgenic control plants showed severe symptoms that progressed to complete wilting. These results indicate that the constitutive expression of the rice Xa21 gene in banana results in enhanced resistance against Xcm. Furthermore, this work demonstrates the feasibility of PRR gene transfer between monocotyledonous species and provides a valuable new tool for controlling the BXW pandemic of banana, a staple food for 100 million people in east Africa.

  16. Pattern-Recognition Receptor Signaling Regulator mRNA Expression in Humans and Mice, and in Transient Inflammation or Progressive Fibrosis

    PubMed Central

    Günthner, Roman; Kumar, Vankayala Ramaiah Santhosh; Lorenz, Georg; Anders, Hans-Joachim; Lech, Maciej

    2013-01-01

    The cell type-, organ-, and species-specific expression of the pattern-recognition receptors (PRRs) are well described but little is known about the respective expression profiles of their negative regulators. We therefore determined the mRNA expression levels of A20, CYLD, DUBA, ST2, CD180, SIGIRR, TANK, SOCS1, SOCS3, SHIP, IRAK-M, DOK1, DOK2, SHP1, SHP2, TOLLIP, IRF4, SIKE, NLRX1, ERBIN, CENTB1, and Clec4a2 in human and mouse solid organs. Humans and mice displayed significant differences between their respective mRNA expression patterns of these factors. Additionally, we characterized their expression profiles in mononuclear blood cells upon bacterial endotoxin, which showed a consistent induction of A20, SOCS3, IRAK-M, and Clec4a2 in human and murine cells. Furthermore, we studied the expression pattern in transient kidney ischemia-reperfusion injury versus post-ischemic atrophy and fibrosis in mice. A20, CD180, ST2, SOCS1, SOCS3, SHIP, IRAK-M, DOK1, DOK2, IRF4, CENTB1, and Clec4a2 were all induced, albeit at different times of injury and repair. Progressive fibrosis was associated with a persistent induction of these factors. Thus, the organ- and species-specific expression patterns need to be considered in the design and interpretation of studies related to PRR-mediated innate immunity, which seems to be involved in tissue injury, tissue regeneration and in progressive tissue scarring. PMID:24009023

  17. Aspergillus fumigatus evades immune recognition during germination through loss of toll-like receptor-4-mediated signal transduction.

    PubMed

    Netea, Mihai G; Warris, Adilia; Van der Meer, Jos W M; Fenton, Matthew J; Verver-Janssen, Trees J G; Jacobs, Liesbeth E H; Andresen, Tonje; Verweij, Paul E; Kullberg, Bart Jan

    2003-07-15

    Peritoneal macrophages from Toll-like receptor (TLR) 4-deficient ScCr mice produced less tumor necrosis factor, interleukin (IL)-1alpha, and IL-1beta than did macrophages of control mice, when stimulated with conidia, but not with hyphae, of Aspergillus fumigatus, a finding suggesting that TLR4-mediated signals are lost during germination. This hypothesis was confirmed by use of a TLR4-specific fibroblast reporter cell line (3E10) that responded to the conidia, but not to the hyphae, of A. fumigatus. In contrast, macrophages from TLR2-knockout mice had a decreased production of proinflammatory cytokines in response to both Aspergillus conidia and Aspergillus hyphae, and these results were confirmed in 3E10 cells transfected with human TLR2. In addition, Aspergillus hyphae, but not Aspergillus conidia, stimulated production of IL-10 through TLR2-dependent mechanisms. In conclusion, TLR4-mediated proinflammatory signals, but not TLR2-induced anti-inflammatory signals, are lost on Aspergillus germination to hyphae. Therefore, phenotypic switching during germination may be an important escape mechanism of A. fumigatus that results in counteracting the host defense.

  18. T helper cell recognition of muscle acetylcholine receptor in myasthenia gravis. Epitopes on the gamma and delta subunits.

    PubMed Central

    Manfredi, A A; Protti, M P; Dalton, M W; Howard, J F; Conti-Tronconi, B M

    1993-01-01

    We tested the response of CD4+ cells and/or total lymphocytes from the blood of 22 myasthenic patients and 10 healthy controls to overlapping synthetic peptides, 20 residues long, to screen the sequence of the gamma and delta subunits of human muscle acetylcholine receptor (AChR). The gamma subunit is part of the AChR expressed in embryonic muscle and is substituted in the AChRs of most adult muscles by an epsilon subunit. The delta subunit is present in both embryonic and adult AChRs. Adult extrinsic ocular muscles, which are preferentially and sometimes uniquely affected by myasthenic symptoms, and thymus, which has a still obscure but important role in the pathogenesis of myasthenia gravis, express the embryonic gamma subunit. Anti-AChR CD4+ responses were more easily detected after CD8+ depletion. All responders recognized epitopes on both the gamma and delta subunits and had severe symptoms. In four patients the CD4+ cell response was tested twice, when the symptoms were severe and during a period of remission. Consistently, the response was only detectable, or larger, when the patients were severely affected. Images PMID:7688757

  19. Clearance of Cell Remnants and Regeneration of Injured Muscle Depend on Soluble Pattern Recognition Receptor PTX3

    PubMed Central

    Vezzoli, Michela; Sciorati, Clara; Campana, Lara; Monno, Antonella; Doglio, Maria Giulia; Rigamonti, Elena; Corna, Gianfranca; Touvier, Thierry; Castiglioni, Alessandra; Capobianco, Annalisa; Mantovani, Alberto; Manfredi, Angelo A; Garlanda, Cecilia; Rovere-Querini, Patrizia

    2016-01-01

    The signals causing resolution of muscle inflammation are only partially characterized. The long pentraxin PTX3, which modulates leukocyte recruitment and activation, could contribute. We analyzed the expression of PTX3 after muscle injury and verified whether hematopoietic precursors are a source of the protein. The kinetics of regeneration and leukocyte infiltration and the accumulation of cell remnants and anti-histidyl-t-RNA synthetase autoantibodies were compared in wild-type and PTX3-deficient mice. PTX3 expression was upregulated 3 d to 5 d after injury and restricted to the extracellular matrix. Cellular debris and leukocytes persisted in the muscle of PTX3-deficient mice for a long time after wild-type animals had healed. PTX3-deficient macrophages expressed receptors involved in apoptotic cell clearance and engulfed dead cells in vitro. Accumulation of cell debris in a proinflammatory microenvironment was not sufficient to elicit autoantibodies. We concluded that PTX3 generated in response to muscle injury prompts clearance of debris and termination of the inflammatory response. PMID:27900389

  20. Highly conserved regions within the spike proteins of human coronaviruses 229E and NL63 determine recognition of their respective cellular receptors.

    PubMed

    Hofmann, Heike; Simmons, Graham; Rennekamp, Andrew J; Chaipan, Chawaree; Gramberg, Thomas; Heck, Elke; Geier, Martina; Wegele, Anja; Marzi, Andrea; Bates, Paul; Pöhlmann, Stefan

    2006-09-01

    We have recently demonstrated that the severe acute respiratory syndrome coronavirus (SARS-CoV) receptor angiotensin converting enzyme 2 (ACE2) also mediates cellular entry of the newly discovered human coronavirus (hCoV) NL63. Here, we show that expression of DC-SIGN augments NL63 spike (S)-protein-driven infection of susceptible cells, while only expression of ACE2 but not DC-SIGN is sufficient for entry into nonpermissive cells, indicating that ACE2 fulfills the criteria of a bona fide hCoV-NL63 receptor. As for SARS-CoV, murine ACE2 is used less efficiently by NL63-S for entry than human ACE2. In contrast, several amino acid exchanges in human ACE2 which diminish SARS-S-driven entry do not interfere with NL63-S-mediated infection, suggesting that SARS-S and NL63-S might engage human ACE2 differentially. Moreover, we observed that NL63-S-driven entry was less dependent on a low-pH environment and activity of endosomal proteases compared to infection mediated by SARS-S, further suggesting differences in hCoV-NL63 and SARS-CoV cellular entry. NL63-S does not exhibit significant homology to SARS-S but is highly related to the S-protein of hCoV-229E, which enters target cells by engaging CD13. Employing mutagenic analyses, we found that the N-terminal unique domain in NL63-S, which is absent in 229E-S, does not confer binding to ACE2. In contrast, the highly homologous C-terminal parts of the NL63-S1 and 229E-S1 subunits in conjunction with distinct amino acids in the central regions of these proteins confer recognition of ACE2 and CD13, respectively. Therefore, despite the high homology of these sequences, they likely form sufficiently distinct surfaces, thus determining receptor specificity.

  1. Fluorescence switch on-off-on receptor constructed of quinoline allied calix[4]arene for selective recognition of Cu2+ from blood serum and F- from industrial waste water.

    PubMed

    Sutariya, Pinkesh G; Pandya, Alok; Lodha, Anand; Menon, Shobhana K

    2013-05-07

    A novel PET with ICT based substituted calix[4]arene fluroionophore was synthesized and used for detection of Cu(2+) and F(-) by emission spectra. The detection limit of the synthesized receptor was found to be 4.16 nM for Cu(2+) and 2.15 nM for F(-). Moreover, this probe has been applied for recognition of Cu(2+) in blood serum and F(-) from waste water.

  2. [The unity of image recognition of tolite homothetic receptors and glycosylation end product receptors in biologic function in case of diabetes].

    PubMed

    Titov, V N; Shiriaeva, Iu K

    2011-08-01

    In compliance with our theory of biologic functions and reactions, pathogenesis of all diseases in formed in the framework of disorder of seven biologic functions: trophology, homeostasis, endoecology, adaptation, locomotion, genus continuation and intelligence. The disorder of biologic function of endoecology ("purity" of intercellular medium in vivo) is the most common basis of pathogenesis of diseases' nosological forms. There are two methods of elimination of disorder of this function. First is the biological reaction of excretion i.e. the removal of minor endogenic phlogogenes (initiators of inflammation) with molecular weightless than 70 amu. The second is the biological function of inflammation i.e. utilization of large endogenic phlogogenes (more than 70 amu) by means of phagocytosis of functional phagocytes. The inherent and acquired immunity are other biological reactions of biological function of endoecology. They overlap with biological reaction of transcytosis and biological reaction of hydrodynamic pressure activating transcytosis. The system of tolite homothetic receptors discriminating molecules of native proteins and endogenic phlogogenes on principle "right-wrong" is the basis of biological function of inherent immunity. In case of hyperglycemia and diabetes glucose becomes the minor biological "refuse". The products of chemical reaction between glucose and proteins and formation of the end products of glycosylation, components of areolar tissue become the major biological "refuse". The accumulation of products of glycosylation of collagen with cross-links in capillaries' wall increases its rigidity and makes it impossible for pericytes to implement the qualities of primary peristaltic pump. Hence the formation of hypoperfusion, condition of hypoxemia in microcirculatory section of blood circulation (muscular type arteriolae, postateriolae with muscular sphincter, metabolic capillaries) resulting in the development of microangiopathies of distal

  3. The Dynamics of the Human Leukocyte Antigen Head Domain Modulates Its Recognition by the T-Cell Receptor.

    PubMed

    García-Guerrero, Estefanía; Pérez-Simón, José Antonio; Sánchez-Abarca, Luis Ignacio; Díaz-Moreno, Irene; De la Rosa, Miguel A; Díaz-Quintana, Antonio

    2016-01-01

    Generating the immune response requires the discrimination of peptides presented by the human leukocyte antigen complex (HLA) through the T-cell receptor (TCR). However, how a single amino acid substitution in the antigen bonded to HLA affects the response of T cells remains uncertain. Hence, we used molecular dynamics computations to analyze the molecular interactions between peptides, HLA and TCR. We compared immunologically reactive complexes with non-reactive and weakly reactive complexes. MD trajectories were produced to simulate the behavior of isolated components of the various p-HLA-TCR complexes. Analysis of the fluctuations showed that p-HLA binding barely restrains TCR motions, and mainly affects the CDR3 loops. Conversely, inactive p-HLA complexes displayed significant drop in their dynamics when compared with its free versus ternary forms (p-HLA-TCR). In agreement, the free non-reactive p-HLA complexes showed a lower amount of salt bridges than the responsive ones. This resulted in differences between the electrostatic potentials of reactive and inactive p-HLA species and larger vibrational entropies in non-elicitor complexes. Analysis of the ternary p-HLA-TCR complexes also revealed a larger number of salt bridges in the responsive complexes. To summarize, our computations indicate that the affinity of each p-HLA complex towards TCR is intimately linked to both, the dynamics of its free species and its ability to form specific intermolecular salt-bridges in the ternary complexes. Of outstanding interest is the emerging concept of antigen reactivity involving its interplay with the HLA head sidechain dynamics by rearranging its salt-bridges.

  4. Mutational analysis identifies leucine-rich repeat insertions crucial for pigeon toll-like receptor 7 recognition and signaling.

    PubMed

    Xiong, Dan; Song, Li; Jiao, Yang; Kang, Xilong; Chen, Xiang; Geng, Shizhong; Pan, Zhiming; Jiao, Xinan

    2015-11-15

    Toll-like receptor 7 (TLR7) is responsible for recognizing viral single-stranded RNA and antiviral imidazoquinoline compounds, leading to the activation of the innate immune response. In this study, mutated pigeon TLR7 fragments, in which the insertion at position 10 of leucine-rich repeat 10 (LRR10) or at position 15 of LRR2/11/13/14 was deleted, were amplified with an overlap-PCR method, and inserted into the expression vector pCMV. The immune functions of the TLR7 mutants were determined with an NF-κB luciferase assay of transfected cells. The deletion of the insertions absolutely abolished TLR7-NF-κB signaling. With quantitative real-time PCR and sandwich enzyme-linked immunosorbent assay, we observed that stimulation with R848 failed to induce the expression of interleukin 8 (IL-8) in any of the mutant-TLR7-transfected cells, consistent with their lack of NF-κB activity. However, the expression of interferon α (IFN-α) and tumor necrosis factor α (TNF-α) was significantly upregulated in the Del10IN10 and Del14IN15 groups. Remarkably, the levels of pigeon TLR7 expression were significantly increased in all the TLR7-mutated groups. Therefore, we speculate that another part of the deficient TLR7 mediates the induction of IFN-α and TNF-α by increasing the expression of TLR7 as compensation. However, the increased expression of TLR7 in the Del11IN15 group failed to induce the production of IFN-α, IL-8, or TNF-α, indicating that a false compensation occurred when the crucial LRR insertion was deleted.

  5. The Dynamics of the Human Leukocyte Antigen Head Domain Modulates Its Recognition by the T-Cell Receptor

    PubMed Central

    García-Guerrero, Estefanía; Pérez-Simón, José Antonio; Sánchez-Abarca, Luis Ignacio; Díaz-Moreno, Irene; De la Rosa, Miguel A.; Díaz-Quintana, Antonio

    2016-01-01

    Generating the immune response requires the discrimination of peptides presented by the human leukocyte antigen complex (HLA) through the T-cell receptor (TCR). However, how a single amino acid substitution in the antigen bonded to HLA affects the response of T cells remains uncertain. Hence, we used molecular dynamics computations to analyze the molecular interactions between peptides, HLA and TCR. We compared immunologically reactive complexes with non-reactive and weakly reactive complexes. MD trajectories were produced to simulate the behavior of isolated components of the various p-HLA-TCR complexes. Analysis of the fluctuations showed that p-HLA binding barely restrains TCR motions, and mainly affects the CDR3 loops. Conversely, inactive p-HLA complexes displayed significant drop in their dynamics when compared with its free versus ternary forms (p-HLA-TCR). In agreement, the free non-reactive p-HLA complexes showed a lower amount of salt bridges than the responsive ones. This resulted in differences between the electrostatic potentials of reactive and inactive p-HLA species and larger vibrational entropies in non-elicitor complexes. Analysis of the ternary p-HLA-TCR complexes also revealed a larger number of salt bridges in the responsive complexes. To summarize, our computations indicate that the affinity of each p-HLA complex towards TCR is intimately linked to both, the dynamics of its free species and its ability to form specific intermolecular salt-bridges in the ternary complexes. Of outstanding interest is the emerging concept of antigen reactivity involving its interplay with the HLA head sidechain dynamics by rearranging its salt-bridges. PMID:27124285

  6. [The role of the class A scavenger receptors, SR-A and MARCO, in the immune system. Part 2. Contribution to recognition and phagocytosis of pathogens as well as induction of immune response].

    PubMed

    Józefowski, Szczepan

    2012-02-29

    Recognition of pathogens by innate immune cells is mediated by pattern recognition receptors (PRR), which include the class A scavenger receptors (SR), SR-A/CD204 and MARCO. It seems that in addition to activating innate immune responses, phagocytosis and inflammation, this initial, PRR-mediated recognition also determines polarization of adaptive immune responses (Th1, Th2, Th17 or Treg). It has been demonstrated that class A SR are major PRR mediating opsonin-independent phagocytosis. SR-A- or MARCO-deficient mice exhibit impaired ability to control bacterial infections, resulting in increased mortality. Our results suggest that in addition to impaired bacterial destruction by macrophages, dysregulation of immune responses may contribute to defective antibacterial defense in class A SR-deficient mice. Using specific receptor ligation with antibodies, we showed that SR-A and MARCO regulate in an opposite manner production of IL-12 in macrophages, the cytokine playing a crucial role in Th1/Th2 polarization of adaptive immune responses. Together with the observation that expression of MARCO is increased by different Th1-polarizing factors and decreased by Th2-polarizing factors, these results suggest that changes in relative expression levels of SR-A and MARCO may be a mechanism of sustained polarization of adaptive immune responses.

  7. Linkage of multiequilibria in DNA recognition by the D53H Escherichia coli cAMP receptor protein.

    PubMed

    Lin, Shwu-Hwa; Lee, J Ching

    2002-12-17

    The transcription factor cyclic AMP receptor protein, CRP, regulates the operons that encode proteins involved in translocation and metabolism of carbohydrates in Escherichia coli. The structure of the CRP-cAMP complex reveals the presence of two sets of cAMP binding sites. Solution biophysical studies show that there are two high-affinity and two low-affinity binding sites, to which the binding of cAMP is characterized by varying degrees of cooperativity. A stoichiometry of four implies that potentially CRP can exist in five conformers with different numbers of bound cAMP. These conformers may exhibit differential affinities for specific DNA sequences. In this study, the affinity between DNA and each conformer of D53H CRP was defined through a dissection of the thermodynamic linkage scheme that included all the conformers. Loading of the high- and low-affinity sites with cAMP leads to high and low affinity for DNA, respectively. The specific magnitude of the binding constants of these conformers is DNA sequence dependent. The various association constants defined by the present study provide a solution to address an enigma of the CRP system, namely, the 3 orders of magnitude difference between the cAMP binding constants determined by in vitro studies and the cAMP concentration regime to which the bacteria respond. Under physiological conditions, the apo-CRP-DNA complex is the dominant species. As a consequence of the 1000-fold stronger affinity of cAMP to the apo-CRP-DNA complex than that to CRP, the relevant reaction is the binding of cAMP to this DNA-protein complex. The binding constant is of the order of 10(7) M(-)(1), the same concentration regime as that of cellular concentration of cAMP. In addition, under physiological conditions the species that binds to the lac and gal operons is predicted to be CRP-(cAMP)(1). A comparison of parameters between the wild type and the mutant CRP shows that the mutation apparently shifts the various thermodynamically linked

  8. Exchange of the H(CC) domain mediating double receptor recognition improves the pharmacodynamic properties of botulinum neurotoxin.

    PubMed

    Rummel, Andreas; Mahrhold, Stefan; Bigalke, Hans; Binz, Thomas

    2011-12-01

    The four-domain structure of botulinum neurotoxins (BoNTs) reflects their multistep intoxication process. The high toxicity of BoNTs primarily results from specific binding and uptake into neurons mediated by their 50-kDa cell-binding fragment (H(C) ). X-ray crystallography data have revealed that the H(C) fragment consists of two domains of equal size, named the 25-kDa N-terminal half of H(C) (H(CN) ) and the 25-kDa C-terminal half of H(C) (H(CC) ). In recent years, the ganglioside-binding sites of all seven BoNT serotypes have been allocated to the H(CC) domain. For BoNT/A, BoNT/B and BoNT/G, the protein receptor-binding site has been also been localized to the H(CC) domain. Here, we demonstrate that the H(CC) serotype can modulate the affinity of the H(C) fragment for neuronal membranes as well as the potency of full-length BoNT by replacing the BoNT/A H(CC) domain with the BoNT/B H(CC) , BoNT/C H(CC) and BoNT/E H(CC) domains, which exhibit higher affinity for synaptosomes. Indeed, the hybrids H(C) AB and H(C) AC display a higher affinity than wild-type H(C) A. Furthermore, the potency of a BoNT/A-based full-length hybrid containing the H(CC) B domain (AAAB; letters represent the serotype origin of the four domains) was quadrupled as compared with wild-type BoNT/A. Analogously, exchange of the H(C) fragment (AABB) yielded a neurotoxin with four-fold higher potency. As BoNT/A and BoNT/B are extensively used to treat neurological disorders, thereby facing the problem of BoNT neutralizing antibody formation, a BoNT with increased potency would lower the repeatedly administered protein dosage while maintaining the clinical benefit. Such a lowered protein load will delay the onset of neurotoxin antibody formation in patients.

  9. A novel junctional adhesion molecule A (CgJAM-A-L) from oyster (Crassostrea gigas) functions as pattern recognition receptor and opsonin.

    PubMed

    Liu, Conghui; Wang, Mengqiang; Jiang, Shuai; Wang, Lingling; Chen, Hao; Liu, Zhaoqun; Qiu, Limei; Song, Linsheng

    2016-02-01

    Junctional adhesion molecule (JAM), a subfamily of immunoglobulin superfamily (IgSF) with a couple of immunoglobulin domains, can act as regulator in homeostasis and inflammation of vertebrates. In the present study, a structural homolog of JAM-A (designated CgJAM-A-L) was screened out from oyster, Crassostrea gigas, through a search of JAM-A D1 domain (N-terminal Ig domain in JAM-A). The cDNA of CgJAM-A-L was of 1188 bp encoding a predicted polypeptide of 395 amino acids. The immunoreactive area of CgJAM-A-L mainly distributed over the plasma membrane of hemocytes. After Vibro splendidus or tumor necrosis factor (CgTNF-1) stimulation, the mRNA transcripts of CgJAM-A-L in hemocytes increased significantly by 4.46-fold and 9.00-fold (p < 0.01) of those in control group, respectively. The recombinant CgJAM-A-L protein (rCgJAM-A-L) could bind multiple PAMPs including lipopolysaccharides (LPS), peptidoglycan (PGN), lipoteichoic acid (LTA), mannose (MAN), β-glucan (GLU) and poly(I:C), and various microorganisms including Micrococcus luteus, Staphylococcus aureus, Escherichia coli, Vibro anguillarum, V. splendidus, Pastoris pastoris and Yarrowia lipolytica. The phagocytic rates of oyster hemocytes towards Gram-negative bacteria V. anguillarum and yeast P. pastoris were significantly enhanced after the incubation of rCgJAM-A-L, and even increased more significantly after the pre-incubation of rCgJAM-A-L with microbes (p < 0.01). The results collectively indicated that CgJAM-A-L functioned as an important pattern recognition receptor (PRR) and opsonin in the immune defense against invading pathogen in oyster. Moreover, as the most primitive specie with homolog of JAMs, the information of CgJAM-A-L in oyster would provide useful clues for the evolutionary study of JAMs and immunoglobulins.

  10. Hepatitis C Virus Frameshift/Alternate Reading Frame Protein Suppresses Interferon Responses Mediated by Pattern Recognition Receptor Retinoic-Acid-Inducible Gene-I

    PubMed Central

    Park, Seung Bum; Seronello, Scott; Mayer, Wasima; Ojcius, David M.

    2016-01-01

    Hepatitis C virus (HCV) actively evades host interferon (IFN) responses but the mechanisms of how it does so are not completely understood. In this study, we present evidence for an HCV factor that contributes to the suppression of retinoic-acid-inducible gene-I (RIG-I)-mediated IFN induction. Expression of frameshift/alternate reading frame protein (F/ARFP) from HCV -2/+1 frame in Huh7 hepatoma cells suppressed type I IFN responses stimulated by HCV RNA pathogen-associated molecular pattern (PAMP) and poly(IC). The suppression occurred independently of other HCV factors; and activation of interferon stimulated genes, TNFα, IFN-λ1, and IFN-λ2/3 was likewise suppressed by HCV F/ARFP. Point mutations in the full-length HCV sequence (JFH1 genotype 2a strain) were made to introduce premature termination codons in the -2/+1 reading frame coding for F/ARFP while preserving the original reading frame, which enhanced IFNα and IFNβ induction by HCV. The potentiation of IFN response by the F/ARFP mutations was diminished in Huh7.5 cells, which already have a defective RIG-I, and by decreasing RIG-I expression in Huh7 cells. Furthermore, adding F/ARFP back via trans-complementation suppressed IFN induction in the F/ARFP mutant. The F/ARFP mutants, on the other hand, were not resistant to exogenous IFNα. Finally, HCV-infected human liver samples showed significant F/ARFP antibody reactivity, compared to HCV-uninfected control livers. Therefore, HCV F/ARFP likely cooperates with other viral factors to suppress type I and III IFN induction occurring through the RIG-I signaling pathway. This study identifies a novel mechanism of pattern recognition receptor modulation by HCV and suggests a biological function of the HCV alternate reading frame in the modulation of host innate immunity. PMID:27404108

  11. Molecular modeling of a T-cell receptor bound to a major histocompatibility complex molecule: implications for T-cell recognition.

    PubMed Central

    Almagro, J. C.; Vargas-Madrazo, E.; Lara-Ochoa, F.; Horjales, E.

    1995-01-01

    The main functions of the T-cell receptor (TCR) involve its specific interaction with short and linear antigenic peptides bound to the major histocompatibility complex (MHC) molecules. In the absence of a 3D structure for TCR and for the TCR/peptide/MHC complex, several attempts to characterize the structural components of the TCR/peptide/MHC interaction have been made. However, this subject is still troublesome. In this paper a computer-based 3D model for a TCR/peptide/MHC complex (5C.C7/moth cytochrome c [MCC] peptide 93-103/I-Ek) was obtained. The complex surface shows a high complementarity between the 5C.C7 structure and the peptide/I-Ek molecule. The mapping of residues involved in the TCR/peptide/MHC interaction shows close agreement with mutational experiments (Jorgensen JL, Reay PA, Ehrich EW, Davis MM, 1992b, Annu Rev Immunol 10:835-873). Moreover, the results are consistent with a recent variability analysis of TCR sequences using three variability indexes (Almagro JC, Zenteno-Cuevas R, Vargas-Madrazo E, Lara-Ochoa F, 1995b, Int J Pept Protein Res 45:180-186). Accordingly, the 3D model of the 5C.C7/MCC peptide 93-103/I-Ek complex provides a framework to generate testable hypotheses about TCR recognition. Thus, starting from this model, the role played by each loop that forms the peptide/MHC binding site of the TCR is discussed. PMID:8528069

  12. Structure-Based Analysis of Toxoplasma gondii Profilin: A Parasite-Specific Motif Is Required for Recognition by Toll-Like Receptor 11

    SciTech Connect

    K Kucera; A Koblansky; L Saunders; K Frederick; E De La Cruz; S Ghosh; Y Modis

    2011-12-31

    Profilins promote actin polymerization by exchanging ADP for ATP on monomeric actin and delivering ATP-actin to growing filament barbed ends. Apicomplexan protozoa such as Toxoplasma gondii invade host cells using an actin-dependent gliding motility. Toll-like receptor (TLR) 11 generates an innate immune response upon sensing T. gondii profilin (TgPRF). The crystal structure of TgPRF reveals a parasite-specific surface motif consisting of an acidic loop, followed by a long {beta}-hairpin. A series of structure-based profilin mutants show that TLR11 recognition of the acidic loop is responsible for most of the interleukin (IL)-12 secretion response to TgPRF in peritoneal macrophages. Deletion of both the acidic loop and the {beta}-hairpin completely abrogates IL-12 secretion. Insertion of the T. gondii acidic loop and {beta}-hairpin into yeast profilin is sufficient to generate TLR11-dependent signaling. Substitution of the acidic loop in TgPRF with the homologous loop from the apicomplexan parasite Cryptosporidium parvum does not affect TLR11-dependent IL-12 secretion, while substitution with the acidic loop from Plasmodium falciparum results in reduced but significant IL-12 secretion. We conclude that the parasite-specific motif in TgPRF is the key molecular pattern recognized by TLR11. Unlike other profilins, TgPRF slows nucleotide exchange on monomeric rabbit actin and binds rabbit actin weakly. The putative TgPRF actin-binding surface includes the {beta}-hairpin and diverges widely from the actin-binding surfaces of vertebrate profilins.

  13. Cannabinoid CB1 receptor recognition of endocannabinoids via the lipid bilayer: molecular dynamics simulations of CB1 transmembrane helix 6 and anandamide in a phospholipid bilayer

    NASA Astrophysics Data System (ADS)

    Lynch, Diane L.; Reggio, Patricia H.

    2006-08-01

    The phospholipid bilayer plays a central role in the lifecycle of the endogenous cannabinoid, N-arachidonoylethanolamine (anandamide, AEA). Therefore, the orientation and location of AEA in the phospholipid bilayer with respect to key membrane associated proteins, is a central issue in understanding the mechanism of endocannabinoid signaling. In this paper, we report a test of the hypothesis that a βXX β motif (formed by beta branching amino acids, V6.43 and I6.46) on the lipid face of the cannabinoid CB1 receptor in its inactive state may serve as an initial CB1 interaction site for AEA. Eight 6 ns NAMD2 molecular dynamics simulations of AEA were conducted in a model system composed of CB1 transmembrane helix 6 (TMH6) in a 1,2-dioleoyl- sn-glycero-3-phosphocholine (DOPC) bilayer. In addition, eight 6 ns NAMD2 molecular dynamics simulations of a low CB1 affinity (20:2, n-6) analog of AEA were conducted in the same model system. AEA was found to exhibit a higher incidence of V6.43/I6.46 groove insertion than did the (20:2, n-6) analog. In certain cases, AEA established a high energy of interaction with TMH6 by first associating with the V6.43/I6.46 groove and then molding itself to the lipid face of TMH6 to establish a hydrogen bonding interaction with the exposed backbone carbonyl of P6.50. Based upon these results, we propose that the formation of this hydrogen bonded AEA/TMH6 complex may be the initial step in CB1 recognition of AEA in the lipid bilayer.

  14. Atypical Antigen Recognition Mode of a Shark Immunoglobulin New Antigen Receptor (IgNAR) Variable Domain Characterized by Humanization and Structural Analysis

    PubMed Central

    Kovalenko, Oleg V.; Olland, Andrea; Piché-Nicholas, Nicole; Godbole, Adarsh; King, Daniel; Svenson, Kristine; Calabro, Valerie; Müller, Mischa R.; Barelle, Caroline J.; Somers, William; Gill, Davinder S.; Mosyak, Lidia; Tchistiakova, Lioudmila

    2013-01-01

    The immunoglobulin new antigen receptors (IgNARs) are a class of Ig-like molecules of the shark immune system that exist as heavy chain-only homodimers and bind antigens by their single domain variable regions (V-NARs). Following shark immunization and/or in vitro selection, V-NARs can be generated as soluble, stable, and specific high affinity monomeric binding proteins of ∼12 kDa. We have previously isolated a V-NAR from an immunized spiny dogfish shark, named E06, that binds specifically and with high affinity to human, mouse, and rat serum albumins. Humanization of E06 was carried out by converting over 60% of non-complementarity-determining region residues to those of a human germ line Vκ1 sequence, DPK9. The resulting huE06 molecules have largely retained the specificity and affinity of antigen binding of the parental V-NAR. Crystal structures of the shark E06 and its humanized variant (huE06 v1.1) in complex with human serum albumin (HSA) were determined at 3- and 2.3-Å resolution, respectively. The huE06 v1.1 molecule retained all but one amino acid residues involved in the binding site for HSA. Structural analysis of these V-NARs has revealed an unusual variable domain-antigen interaction. E06 interacts with HSA in an atypical mode that utilizes extensive framework contacts in addition to complementarity-determining regions that has not been seen previously in V-NARs. On the basis of the structure, the roles of various elements of the molecule are described with respect to antigen binding and V-NAR stability. This information broadens the general understanding of antigen recognition and provides a framework for further design and humanization of shark IgNARs. PMID:23632026

  15. Atypical antigen recognition mode of a shark immunoglobulin new antigen receptor (IgNAR) variable domain characterized by humanization and structural analysis.

    PubMed

    Kovalenko, Oleg V; Olland, Andrea; Piché-Nicholas, Nicole; Godbole, Adarsh; King, Daniel; Svenson, Kristine; Calabro, Valerie; Müller, Mischa R; Barelle, Caroline J; Somers, William; Gill, Davinder S; Mosyak, Lidia; Tchistiakova, Lioudmila

    2013-06-14

    The immunoglobulin new antigen receptors (IgNARs) are a class of Ig-like molecules of the shark immune system that exist as heavy chain-only homodimers and bind antigens by their single domain variable regions (V-NARs). Following shark immunization and/or in vitro selection, V-NARs can be generated as soluble, stable, and specific high affinity monomeric binding proteins of ∼12 kDa. We have previously isolated a V-NAR from an immunized spiny dogfish shark, named E06, that binds specifically and with high affinity to human, mouse, and rat serum albumins. Humanization of E06 was carried out by converting over 60% of non-complementarity-determining region residues to those of a human germ line Vκ1 sequence, DPK9. The resulting huE06 molecules have largely retained the specificity and affinity of antigen binding of the parental V-NAR. Crystal structures of the shark E06 and its humanized variant (huE06 v1.1) in complex with human serum albumin (HSA) were determined at 3- and 2.3-Å resolution, respectively. The huE06 v1.1 molecule retained all but one amino acid residues involved in the binding site for HSA. Structural analysis of these V-NARs has revealed an unusual variable domain-antigen interaction. E06 interacts with HSA in an atypical mode that utilizes extensive framework contacts in addition to complementarity-determining regions that has not been seen previously in V-NARs. On the basis of the structure, the roles of various elements of the molecule are described with respect to antigen binding and V-NAR stability. This information broadens the general understanding of antigen recognition and provides a framework for further design and humanization of shark IgNARs.

  16. Molecular modeling of a T-cell receptor bound to a major histocompatibility complex molecule: implications for T-cell recognition.

    PubMed

    Almagro, J C; Vargas-Madrazo, E; Lara-Ochoa, F; Horjales, E

    1995-09-01

    The main functions of the T-cell receptor (TCR) involve its specific interaction with short and linear antigenic peptides bound to the major histocompatibility complex (MHC) molecules. In the absence of a 3D structure for TCR and for the TCR/peptide/MHC complex, several attempts to characterize the structural components of the TCR/peptide/MHC interaction have been made. However, this subject is still troublesome. In this paper a computer-based 3D model for a TCR/peptide/MHC complex (5C.C7/moth cytochrome c [MCC] peptide 93-103/I-Ek) was obtained. The complex surface shows a high complementarity between the 5C.C7 structure and the peptide/I-Ek molecule. The mapping of residues involved in the TCR/peptide/MHC interaction shows close agreement with mutational experiments (Jorgensen JL, Reay PA, Ehrich EW, Davis MM, 1992b, Annu Rev Immunol 10:835-873). Moreover, the results are consistent with a recent variability analysis of TCR sequences using three variability indexes (Almagro JC, Zenteno-Cuevas R, Vargas-Madrazo E, Lara-Ochoa F, 1995b, Int J Pept Protein Res 45:180-186). Accordingly, the 3D model of the 5C.C7/MCC peptide 93-103/I-Ek complex provides a framework to generate testable hypotheses about TCR recognition. Thus, starting from this model, the role played by each loop that forms the peptide/MHC binding site of the TCR is discussed.

  17. Molecular basis of non-self recognition by the horseshoe crab tachylectins.

    PubMed

    Kawabata, Shun-ichiro; Tsuda, Ryoko

    2002-09-19

    The self/non-self discrimination by innate immunity through simple ligands universally expressed both on pathogens and hosts, such as monosaccharides and acetyl group, depends on the density or clustering patterns of the ligands. The specific recognition by the horseshoe crab tachylectins with a propeller-like fold or a propeller-like oligomeric arrangement is reinforced by the short distance between the individual binding sites that interact with pathogen-associated molecular patterns (PAMPs). There is virtually no conformational change in the main or side chains of tachylectins upon binding with the ligands. This low structural flexibility of the propeller structures must be very important for specific interaction with PAMPs. Mammalian lectins, such as mannose-binding lectin and ficolins, trigger complement activation through the lectin pathway in the form of opsonins. However, tachylectins have no effector collagenous domains and no lectin-associated serine proteases found in the mammalian lectins. Furthermore, no complement-like proteins have been found in horseshoe crabs, except for alpha(2)-macroglobulin. The mystery of the molecular mechanism of the scavenging pathway of pathogens in horseshoe crabs remains to be solved.

  18. A cleavage-resistant urokinase plasminogen activator receptor exhibits dysregulated cell-surface clearance.

    PubMed

    Nieves, Evelyn C; Manchanda, Naveen

    2010-04-23

    Urokinase plasminogen activator receptor (u-PAR) binds urokinase plasminogen activator (u-PA) and participates in plasminogen activation in addition to modulating several cellular processes such as adhesion, proliferation, and migration. u-PAR is susceptible to proteolysis by its cognate ligand and several other proteases. To elucidate the biological significance of receptor cleavage by u-PA, we engineered and expressed a two-chain urokinase plasminogen activator (tcu-PA) cleavage-resistant u-PAR (cr-u-PAR). This mutated receptor was similar to wild-type u-PAR in binding u-PA and initiating plasminogen activation. However, cr-u-PAR exhibited accelerated internalization and resurfacing due to direct association with the endocytic receptor alpha(2)-macroglobulin receptor/low density lipoprotein receptor-related protein in the absence of the enzyme x inhibitor complex of tcu-PA and plasminogen activator inhibitor-1 (tcu-PA.PAI-1). cr-u-PAR-expressing cells had enhanced migration compared with wild-type u-PAR-expressing cells, and cr-u-PAR was less sensitive to chymotrypsin cleavage as compared with wt u-PAR. Our studies suggest that these mutations in the linker region result in a rearrangement within the cr-u-PAR structure that makes it resemble its ligand-bound form. This constitutively active variant may mimic highly glycosylated cleavage-resistant u-PAR expressed in certain highly malignant cancer-cells.

  19. Level of receptor-associated protein moderates cellular susceptibility to pseudomonas exotoxin A.

    PubMed Central

    Mucci, D; Forristal, J; Strickland, D; Morris, R; Fitzgerald, D; Saelinger, C B

    1995-01-01

    Pseudomonas exotoxin A (PE) enters mammalian cells via a receptor-mediated endocytic pathway. The initial step in this pathway is binding to the multiligand receptor termed the alpha 2-macroglobulin receptor/low-density lipoprotein receptor-related protein (LRP). Binding of toxin, and of the many other ligands that bind to LRP, is blocked by the addition of a 39-kDa receptor-associated protein (RAP). Here we show that approximately 40% of the cell-associated LRP is on the surface of toxin-sensitive mouse LM fibroblasts and thus accessible for toxin internalization. The remainder is located intracellularly, primarily in the Golgi region. Mammalian cells exhibit a wide range of sensitivity to PE. To investigate possible reasons for this, we examined the expression levels of both LRP and RAP. Results from a variety of cell lines indicated that there was a positive correlation between LRP expression and toxin sensitivity. In the absence of LRP, cells were as much as 200-fold more resistant to PE compared with sensitive cells. A second group of resistant cells expressed LRP but had a high level of RAP. Thus, a toxin-resistant phenotype would be expected when cells expressed either low levels of LRP or high levels of LRP in the presence of high levels of RAP. We hypothesize that RAP has a pivotal role in moderating cellular susceptibility to PE. PMID:7622212

  20. Crystal Structures of Mouse CD1d-IGb3 Complex And Its Cognate Valpha14 T Cell Receptor Suggest a Model for Dual Recognition of Foreign And Self Glycolipids

    SciTech Connect

    Zajonc, D.M.; Saveage, P.B.; Bendelac, A.; Wilson, I.A.; Teyton, L.

    2009-05-28

    The semi-invariant Valpha14Jalpha18 T cell receptor (TCR) is expressed by regulatory NKT cells and has the unique ability to recognize chemically diverse ligands presented by CD1d. The crystal structure of CD1d complexed to a natural, endogenous ligand, isoglobotrihexosylceramide (iGb3), illustrates the extent of this diversity when compared to the binding of potent, exogenous ligands, such as alpha-galactosylceramide (alpha-GalCer). A single mode of recognition for these two classes of ligands would then appear problematic for a single T cell receptor. However, the Valpha14 TCR adopts two different conformations in the crystal where, in one configuration, the presence of a larger cavity between the two CDR3 regions could accommodate iGb3 and, in the other, a smaller cavity fits alpha-GalCer more snugly. Alternatively, the extended iGb3 headgroup could be 'squashed' upon docking of the TCR and accommodated between the CD1 and TCR surfaces. Thus, the same TCR may adopt alternative modes of recognition for these foreign and self-ligands for NKT cell activation.

  1. Crystal structure of a Staphylococcus aureus protein A domain complexed with the Fab fragment of a human IgM antibody: structural basis for recognition of B-cell receptors and superantigen activity.

    PubMed

    Graille, M; Stura, E A; Corper, A L; Sutton, B J; Taussig, M J; Charbonnier, J B; Silverman, G J

    2000-05-09

    Staphylococcus aureus produces a virulence factor, protein A (SpA), that contains five homologous Ig-binding domains. The interactions of SpA with the Fab region of membrane-anchored Igs can stimulate a large fraction of B cells, contributing to lymphocyte clonal selection. To understand the molecular basis for this activity, we have solved the crystal structure of the complex between domain D of SpA and the Fab fragment of a human IgM antibody to 2.7-A resolution. In the complex, helices II and III of domain D interact with the variable region of the Fab heavy chain (V(H)) through framework residues, without the involvement of the hypervariable regions implicated in antigen recognition. The contact residues are highly conserved in human V(H)3 antibodies but not in other families. The contact residues from domain D also are conserved among all SpA Ig-binding domains, suggesting that each could bind in a similar manner. Features of this interaction parallel those reported for staphylococcal enterotoxins that are superantigens for many T cells. The structural homology between Ig V(H) regions and the T-cell receptor V(beta) regions facilitates their comparison, and both types of interactions involve lymphocyte receptor surface remote from the antigen binding site. However, T-cell superantigens reportedly interact through hydrogen bonds with T-cell receptor V(beta) backbone atoms in a primary sequence-independent manner, whereas SpA relies on a sequence-restricted conformational binding with residue side chains, suggesting that this common bacterial pathogen has adopted distinct molecular recognition strategies for affecting large sets of B and T lymphocytes.

  2. α₄β₂ Nicotinic receptor stimulation of the GABAergic system within the orbitofrontal cortex ameliorates the severe crossmodal object recognition impairment in ketamine-treated rats: implications for cognitive dysfunction in schizophrenia.

    PubMed

    Cloke, Jacob M; Winters, Boyer D

    2015-03-01

    Schizophrenia is associated with atypical multisensory integration. Rats treated sub-chronically with NMDA receptor antagonists to model schizophrenia are severely impaired on a tactile-to-visual crossmodal object recognition (CMOR) task, and this deficit is reversed by systemic nicotine. The current study assessed the receptor specificity of the ameliorative effect of nicotine in the CMOR task, as well as the potential for nicotinic receptor (nAChR) interactions with GABA and glutamate. Male Long-Evans rats were treated sub-chronically for 10 days with ketamine or saline and then tested on the CMOR task after a 10-day washout. Systemic nicotine given before the sample phase of the CMOR task reversed the ketamine-induced impairment, but this effect was blocked by co-administration of the GABAA receptor antagonist bicuculline at a dosage that itself did not cause impairment. Pre-sample systemic co-administration of the NMDA receptor antagonist MK-801 did not block the remediating effect of nicotine in ketamine-treated rats. The selective α7 nAChR agonist GTS-21 and α4β2 nAChR agonist ABT-418 were also tested, with only the latter reversing the ketamine impairment dose-dependently; bicuculline also blocked this effect. Similarly, infusions of nicotine or ABT-418 into the orbitofrontal cortex (OFC) reversed the CMOR impairment in ketamine-treated rats, and systemic bicuculline blocked the effect of intra-OFC ABT-418. These results suggest that nicotine-induced agonism of α4β2 nAChRs within the OFC ameliorates CMOR deficits in ketamine-treated rats via stimulation of the GABAergic system. The findings of this research may have important implications for understanding the nature and potential treatment of cognitive impairment in schizophrenia.

  3. Ternary Complex of Transforming Growth Factor-[beta]1 Reveals Isoform-specific Ligand Recognition and Receptor Recruitment in the Superfamily

    SciTech Connect

    Radaev, Sergei; Zou, Zhongcheng; Huang, Tao; Lafer, Eileen M.; Hinck, Andrew P.; Sun, Peter D.

    2010-11-03

    Transforming growth factor (TGF)-{beta}1, -{beta}2, and -{beta}3 are 25-kDa homodimeric polypeptides that play crucial nonoverlapping roles in embryogenesis, tissue development, carcinogenesis, and immune regulation. Here we report the 3.0-{angstrom} resolution crystal structure of the ternary complex between human TGF-{beta}1 and the extracellular domains of its type I and type II receptors, T{beta}RI and T{beta}RII. The TGF-{beta}1 ternary complex structure is similar to previously reported TGF-{beta}3 complex except with a 10{sup o} rotation in T{beta}RI docking orientation. Quantitative binding studies showed distinct kinetics between the receptors and the isoforms of TGF-{beta}. T{beta}RI showed significant binding to TGF-{beta}2 and TGF-{beta}3 but not TGF-{beta}1, and the binding to all three isoforms of TGF-{beta} was enhanced considerably in the presence of T{beta}RII. The preference of TGF-{beta}2 to T{beta}RI suggests a variation in its receptor recruitment in vivo. Although TGF-{beta}1 and TGF-{beta}3 bind and assemble their ternary complexes in a similar manner, their structural differences together with differences in the affinities and kinetics of their receptor binding may underlie their unique biological activities. Structural comparisons revealed that the receptor-ligand pairing in the TGF-{beta} superfamily is dictated by unique insertions, deletions, and disulfide bonds rather than amino acid conservation at the interface. The binding mode of T{beta}RII on TGF-{beta} is unique to TGF-{beta}s, whereas that of type II receptor for bone morphogenetic protein on bone morphogenetic protein appears common to all other cytokines in the superfamily. Further, extensive hydrogen bonds and salt bridges are present at the high affinity cytokine-receptor interfaces, whereas hydrophobic interactions dominate the low affinity receptor-ligand interfaces.

  4. Novel salicylic acid-oriented thiourea-type receptors as colorimetric chemosensor: Synthesis, characterizations and selective naked-eye recognition properties

    NASA Astrophysics Data System (ADS)

    Li, Shaowei; Cao, Xiufang; Chen, Changshui; Ke, Shaoyong

    2012-10-01

    Based on the salicylic acid backbone, three highly sensitive and selective colorimetric chemosensors with an acylthiourea binding unit have been designed, synthesized and characterized. These chemosensors have been utilized for selective recognition of fluoride anions in dry DMSO solution by typical spectroscopic titration techniques. Furthermore, the obtained chemosensors AR1-3 have shown naked-eye sensitivity for detection of biologically important fluoride ion over other anions in solution.

  5. Non-equivalence of Key Positively Charged Residues of the Free Fatty Acid 2 Receptor in the Recognition and Function of Agonist Versus Antagonist Ligands*

    PubMed Central

    Sergeev, Eugenia; Hansen, Anders Højgaard; Pandey, Sunil K.; MacKenzie, Amanda E.; Hudson, Brian D.; Ulven, Trond; Milligan, Graeme

    2016-01-01

    Short chain fatty acids (SCFAs) are produced in the gut by bacterial fermentation of poorly digested carbohydrates. A key mediator of their actions is the G protein-coupled free fatty acid 2 (FFA2) receptor, and this has been suggested as a therapeutic target for the treatment of both metabolic and inflammatory diseases. However, a lack of understanding of the molecular determinants dictating how ligands bind to this receptor has hindered development. We have developed a novel radiolabeled FFA2 antagonist to probe ligand binding to FFA2, and in combination with mutagenesis and molecular modeling studies, we define how agonist and antagonist ligands interact with the receptor. Although both agonist and antagonist ligands contain negatively charged carboxylates that interact with two key positively charged arginine residues in transmembrane domains V and VII of FFA2, there are clear differences in how these interactions occur. Specifically, although agonists require interaction with both arginine residues to bind the receptor, antagonists require an interaction with only one of the two. Moreover, different chemical series of antagonist interact preferentially with different arginine residues. A homology model capable of rationalizing these observations was developed and provides a tool that will be invaluable for identifying improved FFA2 agonists and antagonists to further define function and therapeutic opportunities of this receptor. PMID:26518871

  6. Cation-pi interactions in ligand recognition by serotonergic (5-HT3A) and nicotinic acetylcholine receptors: the anomalous binding properties of nicotine.

    PubMed

    Beene, Darren L; Brandt, Gabriel S; Zhong, Wenge; Zacharias, Niki M; Lester, Henry A; Dougherty, Dennis A

    2002-08-13

    A series of tryptophan analogues has been introduced into the binding site regions of two ion channels, the ligand-gated nicotinic acetylcholine and serotonin 5-HT(3A) receptors, using unnatural amino acid mutagenesis and heterologous expression in Xenopus oocytes. A cation-pi interaction between serotonin and Trp183 of the serotonin channel 5-HT(3A)R is identified for the first time, precisely locating the ligand-binding site of this receptor. The energetic contribution of the observed cation-pi interaction between a tryptophan and the primary ammonium ion of serotonin is estimated to be approximately 4 kcal/mol, while the comparable interaction with the quaternary ammonium of acetylcholine is approximately 2 kcal/mol. The binding mode of nicotine to the nicotinic receptor of mouse muscle is examined by the same technique and found to differ significantly from that of the natural agonist, acetylcholine.

  7. Proestrous compared to diestrous wildtype, but not estrogen receptor beta knockout, mice have better performance in the spontaneous alternation and object recognition tasks and reduced anxiety-like behavior in the elevated plus and mirror maze.

    PubMed

    Walf, Alicia A; Koonce, Carolyn; Manley, Kevin; Frye, Cheryl A

    2009-01-23

    17beta-Estradiol (E(2)) may influence cognitive and/or affective behavior in part via the beta isoform of the estrogen receptor (ERbeta). Endocrine status and behavior in cognitive (object recognition, T-maze), anxiety (open field, elevated plus maze, mirror maze, emergence), and motor/coordination (rotarod, activity chamber) tasks of proestrous and diestrous wildtype (WT) and ERbeta knockout (betaERKO) mice was examined. Proestrous (WT or betaERKO), versus diestrous, mice had higher E(2) and progestin levels in plasma, hippocampus, and cortex. The only effect of genotype on hormone levels was for corticosterone, such that betaERKO mice had higher concentrations of corticosterone than did WT mice. Proestrous WT, but not betaERKO, mice had improved performance in the object recognition (greater percentage of time with novel object) and T-maze tasks (greater percentage of spontaneous alternations) and less anxiety-like behavior in the plus maze (increased duration on open arms) and mirror chamber task (increased duration in mirror) than did diestrous mice. This pattern was not seen in the rotarod, open field, or activity monitor, suggesting effects may be specific to affective and cognitive behavior, rather than motor behavior/coordination. Thus, enhanced performance in cognitive tasks and anti-anxiety-like behavior of proestrous mice may require actions of ERbeta in the hippocampus and/or cortex.

  8. Severe Cross-Modal Object Recognition Deficits in Rats Treated Sub-Chronically with NMDA Receptor Antagonists are Reversed by Systemic Nicotine: Implications for Abnormal Multisensory Integration in Schizophrenia

    PubMed Central

    Jacklin, Derek L; Goel, Amit; Clementino, Kyle J; Hall, Alexander W M; Talpos, John C; Winters, Boyer D

    2012-01-01

    Schizophrenia is a complex and debilitating disorder, characterized by positive, negative, and cognitive symptoms. Among the cognitive deficits observed in patients with schizophrenia, recent work has indicated abnormalities in multisensory integration, a process that is important for the formation of comprehensive environmental percepts and for the appropriate guidance of behavior. Very little is known about the neural bases of such multisensory integration deficits, partly because of the lack of viable behavioral tasks to assess this process in animal models. In this study, we used our recently developed rodent cross-modal object recognition (CMOR) task to investigate multisensory integration functions in rats treated sub-chronically with one of two N-methyl-D-aspartate receptor (NMDAR) antagonists, MK-801, or ketamine; such treatment is known to produce schizophrenia-like symptoms. Rats treated with the NMDAR antagonists were impaired on the standard spontaneous object recognition (SOR) task, unimodal (tactile or visual only) versions of SOR, and the CMOR task with intermediate to long retention delays between acquisition and testing phases, but they displayed a selective CMOR task deficit when mnemonic demand was minimized. This selective impairment in multisensory information processing was dose-dependently reversed by acute systemic administration of nicotine. These findings suggest that persistent NMDAR hypofunction may contribute to the multisensory integration deficits observed in patients with schizophrenia and highlight the valuable potential of the CMOR task to facilitate further systematic investigation of the neural bases of, and potential treatments for, this hitherto overlooked aspect of cognitive dysfunction in schizophrenia. PMID:22669170

  9. Lipopolysaccharide- and β-1,3-glucan-binding protein from Fenneropenaeus merguiensis functions as a pattern recognition receptor with a broad specificity for diverse pathogens in the defense against microorganisms.

    PubMed

    Chaosomboon, Areerat; Phupet, Benjaporn; Rattanaporn, Onnicha; Runsaeng, Phanthipha; Utarabhand, Prapaporn

    2017-02-01

    In crustaceans, lipopolysaccharide- and β-1,3-glucan-binding protein (LGBP) plays an important role in innate immunity by mediating the recognition of pathogens to host cells. Hereby, LGBP was cloned from Fenneropenaeus merguiensis hepatopancreas. Its full-length cDNA (1280 bp) had an open reading frame of 1101 bp, encoding a peptide of 366 amino acids. The LGBP primary structure comprises a recognition motif for β-1,3-linkage of polysaccharides, two integrin binding motifs, a kinase C phosphorylation site and a bacterial glucanase motif. The LGBP mRNA was strongly expressed in hepatopancreas and significantly up-regulated to get the maximum at 12 h upon Vibrio harveyi challenge. Recombinant LGBP (rLGBP) could agglutinate Gram-negative and Gram-positive bacteria including yeast with Ca(2+)-dependence. V. harveyi agglutination induced by rLGBP was intensively inhibited by lipoteichoic acid, less in order were lipopolysaccharide, β-1,3-glucan and N-acetyl neuraminic acid. Western blotting revealed that rLGBP bound widely to Gram-negative and Gram-positive bacteria and also yeast. By ELISA quantification, rLGBP could bind to β-1,3-glucan better than to lipopolysaccharide and lipoteichoic acid. These findings suggest that LGBP may function as a receptor which recognizes invading diverse pathogens and contribute in F. merguiensis immune response.

  10. Positive modulation of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors reverses sub-chronic PCP-induced deficits in the novel object recognition task in rats.

    PubMed

    Damgaard, Trine; Larsen, Dorrit Bjerg; Hansen, Suzanne L; Grayson, Ben; Neill, Jo C; Plath, Niels

    2010-02-11

    Cognitive deficits are a major clinical unmet need in schizophrenia. The psychotomimetic drug phencyclidine (PCP) is widely applied in rodents to mimic symptoms of schizophrenia, including cognitive deficits. Previous studies have shown that sub-chronic PCP induces an enduring episodic memory deficit in female Lister Hooded rats in the novel object recognition (NOR) task. Here we show that positive modulation of AMPA receptor (AMPAR) mediated glutamate transmission alleviates cognitive deficits induced by sub-chronic PCP treatment. Female Lister hooded rats were treated sub-chronically with either vehicle (0.9% saline) or PCP (2mg/kg two doses per day for 7 days), followed by a 7 days washout period. 30 min prior to the acquisition trial of the NOR task animals were dosed with either vehicle, CX546 (10, 40 or 80 mg/kg) or CX516 (0.5, 2.5, 10, 40 or 80 mg/kg). Our results show that sub-chronic PCP treatment induced a significant decrease in the discrimination index (DI) and both ampakines CX546 and CX516 were able to reverse this disruption of object memory in rats in the novel object recognition task. These data suggest that positive AMPAR modulation may represent a mechanism for treatment of cognitive deficits in schizophrenia.

  11. APP, APOE, complement receptor 1, clusterin and PICALM and their involvement in the herpes simplex life cycle.

    PubMed

    Carter, C J

    2010-10-11

    The major Alzheimer's disease susceptibility genes (APOE, clusterin, complement receptor 1 (CR1) and phosphatidylinositol binding clathrin assembly protein, PICALM) can be implicated directly (APOE, CR1) or indirectly (clusterin and PICALM) in the herpes simplex life cycle. The virus binds to proteoliposomes containing APOE or APOA1 and also to CR1, and both clusterin and PICALM are related to a mannose-6-phosphate receptor used by the virus for cellular entry and intracellular transport. PICALM also binds to a nuclear exportin used by the virus for nuclear egress. Clusterin and complement receptor 1 are both related to the complement pathways and play a general role in pathogen defence. In addition, the amyloid precursor protein APP is involved in herpes viral transport and gamma-secretase cleaves a number of receptors used by the virus for cellular entry. APOE, APOA1 and clusterin, or alpha 2-macroglobulin, insulysin and caspase 3, which also bind to the virus, are involved in beta-amyloid clearance or degradation, as are the viral binding complement components, C3 and CR1. There are multiple ways in which the products of key susceptibility genes might be able to modify the viral life cycle and in turn the virus interacts with key proteins involved in APP and beta-amyloid processing. These interactions support a role for the herpes simplex virus in Alzheimer's disease pathology and suggest that antiviral agents or vaccination might be considered as viable therapeutic strategies in Alzheimer's disease.

  12. Conjoint Recognition.

    ERIC Educational Resources Information Center

    Brainerd, C. J.; Reyna, V. F.; Mojardin, A. H.

    1999-01-01

    Reviews some limiting properties of the process-dissociation model as it applies to the study of dual-process conceptions of memory. A second-generation model (conjoint recognition) is proposed to address these limitations and supply additional capabilities. Worked applications to data are provided. (Author/GCP)

  13. Molecular Basis for the Recognition of Structurally Distinct Autoinducer Mimics by the Pseudomonas aeruginosa LasR Quorum-Sensing Signaling Receptor

    SciTech Connect

    Zou, Yaozhong; Nair, Satish K.

    2010-01-12

    The human pathogen Pseudomonas aeruginosa coordinates the expression of virulence factors using quorum sensing, a signaling cascade triggered by the activation of signal receptors by small-molecule autoinducers. These homoserine lactone autoinducers stabilize their cognate receptors and activate their functions as transcription factors. Because quorum sensing regulates the progression of infection and host immune resistance, significant efforts have been devoted toward the identification of small molecules that disrupt this process. Screening efforts have identified a class of triphenyl compounds that are structurally distinct from the homoserine lactone autoinducer, yet interact specifically and potently with LasR receptor to modulate quorum sensing (Muh et al., 2006a). Here we present the high-resolution crystal structures of the ligand binding domain of LasR in complex with the autoinducer N-3-oxo-dodecanoyl homoserine lactone (1.4 {angstrom} resolution), and with the triphenyl mimics TP-1, TP-3, and TP-4 (to between 1.8 {angstrom} and 2.3 {angstrom} resolution). These crystal structures provide a molecular rationale for understanding how chemically distinct compounds can be accommodated by a highly selective receptor, and provide the framework for the development of novel quorum-sensing regulators, utilizing the triphenyl scaffold.

  14. Overlap between the ligand recognition properties of the anandamide transporter and the VR1 vanilloid receptor: inhibitors of anandamide uptake with negligible capsaicin-like activity.

    PubMed

    De Petrocellis, L; Bisogno, T; Davis, J B; Pertwee, R G; Di Marzo, V

    2000-10-13

    Some synthetic agonists of the VR1 vanilloid (capsaicin) receptor also inhibit the facilitated transport into cells of the endogenous cannabinoid anandamide (arachidonoylethanolamide, AEA). Here we tested several AEA derivatives containing various derivatized phenyl groups or different alkyl chains as either inhibitors of the AEA membrane transporter (AMT) in intact cells or functional agonists of the VR1 vanilloid receptor in HEK cells transfected with the human VR1. We found that four known AMT inhibitors, AM404, arvanil, olvanil and linvanil, activate VR1 receptors at concentrations 400-10000-fold lower than those necessary to inhibit the AMT. However, we also found three novel AEA derivatives, named VDM11, VDM12 and VDM13, which inhibit the AMT as potently as AM404 but exhibit little or no agonist activity at hVR1. These compounds are weak inhibitors of AEA enzymatic hydrolysis and poor CB(1)/CB(2) receptor ligands. We show for the first time that, despite the overlap between the chemical moieties of AMT inhibitors and VR1 agonists, selective inhibitors of AEA uptake that do not activate VR1 (e.g. VDM11) can be developed.

  15. Pattern recognition: A basis for remote sensing data analysis

    NASA Technical Reports Server (NTRS)

    Swain, P. H.

    1973-01-01

    The theoretical basis for the pattern-recognition-oriented algorithms used in the multispectral data analysis software system is discussed. A model of a general pattern recognition system is presented. The receptor or sensor is usually a multispectral scanner. For each ground resolution element the receptor produces n numbers or measurements corresponding to the n channels of the scanner.

  16. Phytophthora infestans RXLR-WY Effector AVR3a Associates with Dynamin-Related Protein 2 Required for Endocytosis of the Plant Pattern Recognition Receptor FLS2

    PubMed Central

    Chaparro-Garcia, Angela; Schwizer, Simon; Sklenar, Jan; Yoshida, Kentaro; Petre, Benjamin; Bos, Jorunn I. B.; Schornack, Sebastian; Jones, Alexandra M. E.; Bozkurt, Tolga O.; Kamoun, Sophien

    2015-01-01

    Pathogens utilize effectors to suppress basal plant defense known as PTI (Pathogen-associated molecular pattern-triggered immunity). However, our knowledge of PTI suppression by filamentous plant pathogens, i.e. fungi and oomycetes, remains fragmentary. Previous work revealed that the co-receptor BAK1/SERK3 contributes to basal immunity against the potato pathogen Phytophthora infestans. Moreover BAK1/SERK3 is required for the cell death induced by P. infestans elicitin INF1, a protein with characteristics of PAMPs. The P. infestans host-translocated RXLR-WY effector AVR3a is known to supress INF1-mediated cell death by binding the plant E3 ligase CMPG1. In contrast, AVR3aKI-Y147del, a deletion mutant of the C-terminal tyrosine of AVR3a, fails to bind CMPG1 and does not suppress INF1-mediated cell death. Here, we studied the extent to which AVR3a and its variants perturb additional BAK1/SERK3-dependent PTI responses in N. benthamiana using the elicitor/receptor pair flg22/FLS2 as a model. We found that all tested variants of AVR3a suppress defense responses triggered by flg22 and reduce internalization of activated FLS2. Moreover, we discovered that AVR3a associates with the Dynamin-Related Protein 2 (DRP2), a plant GTPase implicated in receptor-mediated endocytosis. Interestingly, silencing of DRP2 impaired ligand-induced FLS2 internalization but did not affect internalization of the growth receptor BRI1. Our results suggest that AVR3a associates with a key cellular trafficking and membrane-remodeling complex involved in immune receptor-mediated endocytosis. We conclude that AVR3a is a multifunctional effector that can suppress BAK1/SERK3-mediated immunity through at least two different pathways. PMID:26348328

  17. Collagen binding specificity of the discoidin domain receptors: binding sites on collagens II and III and molecular determinants for collagen IV recognition by DDR1.

    PubMed

    Xu, Huifang; Raynal, Nicolas; Stathopoulos, Stavros; Myllyharju, Johanna; Farndale, Richard W; Leitinger, Birgit

    2011-01-01

    The discoidin domain receptors, DDR1 and DDR2 are cell surface receptor tyrosine kinases that are activated by triple-helical collagen. While normal DDR signalling regulates fundamental cellular processes, aberrant DDR signalling is associated with several human diseases. We previously identified GVMGFO (O is hydroxyproline) as a major DDR2 binding site in collagens I-III, and located two additional DDR2 binding sites in collagen II. Here we extend these studies to the homologous DDR1 and the identification of DDR binding sites on collagen III. Using sets of overlapping triple-helical peptides, the Collagen II and Collagen III Toolkits, we located several DDR2 binding sites on both collagens. The interaction of DDR1 with Toolkit peptides was more restricted, with DDR1 mainly binding to peptides containing the GVMGFO motif. Triple-helical peptides containing the GVMGFO motif induced DDR1 transmembrane signalling, and DDR1 binding and receptor activation occurred with the same amino acid requirements as previously defined for DDR2. While both DDRs exhibit the same specificity for binding the GVMGFO motif, which is present only in fibrillar collagens, the two receptors display distinct preferences for certain non-fibrillar collagens, with the basement membrane collagen IV being exclusively recognised by DDR1. Based on our recent crystal structure of a DDR2-collagen complex, we designed mutations to identify the molecular determinants for DDR1 binding to collagen IV. By replacing five amino acids in DDR2 with the corresponding DDR1 residues we were able to create a DDR2 construct that could function as a collagen IV receptor.

  18. The N-terminal portion of domain E of retinoic acid receptors alpha and beta is essential for the recognition of retinoic acid and various analogs.

    PubMed

    Ostrowski, J; Hammer, L; Roalsvig, T; Pokornowski, K; Reczek, P R

    1995-03-14

    Utilizing a strategy involving domain exchange between retinoic acid receptors alpha and beta (RAR alpha and RAR beta) and monitoring the transcriptional activity of the resulting chimeric receptors with receptor-selective retinoids, we identified a 70-aa region within the N-terminal portion of the RAR alpha and -beta domain E which is important for an RAR alpha- or RAR beta-specific response. Two amino acid residues within this region, serine-232 (S232) and threonine-239 (T239) in RAR alpha and the corresponding alanine-225 (A225) and isoleucine-232 (I232) in RAR beta, were found to be essential for this effect. In addition, binding studies using the chimeric receptors expressed in Escherichia coli showed that the N-terminal portion of domain E was also important for the characteristic binding profile of t-RA and various retinoids with RAR alpha or RAR beta. Structural predictions of the primary amino acid sequence in this region indicate the presence of an amphipathic helix-turn-helix structure with five hydrophobic amino acids that resemble a leucine zipper motif. The amino acid residues identified by domain swapping, S232 and T239 in RAR alpha and A225 and I232 in RAR beta, were found within the hydrophobic face of an alpha-helix in close proximity to this zipper motif, suggesting that the ligand may interact with the receptor in the region adjacent to a surface involved in protein-protein interactions. This finding may link ligand binding to other processes important for transcriptional activation.

  19. Phytophthora infestans RXLR-WY Effector AVR3a Associates with Dynamin-Related Protein 2 Required for Endocytosis of the Plant Pattern Recognition Receptor FLS2.

    PubMed

    Chaparro-Garcia, Angela; Schwizer, Simon; Sklenar, Jan; Yoshida, Kentaro; Petre, Benjamin; Bos, Jorunn I B; Schornack, Sebastian; Jones, Alexandra M E; Bozkurt, Tolga O; Kamoun, Sophien

    2015-01-01

    Pathogens utilize effectors to suppress basal plant defense known as PTI (Pathogen-associated molecular pattern-triggered immunity). However, our knowledge of PTI suppression by filamentous plant pathogens, i.e. fungi and oomycetes, remains fragmentary. Previous work revealed that the co-receptor BAK1/SERK3 contributes to basal immunity against the potato pathogen Phytophthora infestans. Moreover BAK1/SERK3 is required for the cell death induced by P. infestans elicitin INF1, a protein with characteristics of PAMPs. The P. infestans host-translocated RXLR-WY effector AVR3a is known to supress INF1-mediated cell death by binding the plant E3 ligase CMPG1. In contrast, AVR3aKI-Y147del, a deletion mutant of the C-terminal tyrosine of AVR3a, fails to bind CMPG1 and does not suppress INF1-mediated cell death. Here, we studied the extent to which AVR3a and its variants perturb additional BAK1/SERK3-dependent PTI responses in N. benthamiana using the elicitor/receptor pair flg22/FLS2 as a model. We found that all tested variants of AVR3a suppress defense responses triggered by flg22 and reduce internalization of activated FLS2. Moreover, we discovered that AVR3a associates with the Dynamin-Related Protein 2 (DRP2), a plant GTPase implicated in receptor-mediated endocytosis. Interestingly, silencing of DRP2 impaired ligand-induced FLS2 internalization but did not affect internalization of the growth receptor BRI1. Our results suggest that AVR3a associates with a key cellular trafficking and membrane-remodeling complex involved in immune receptor-mediated endocytosis. We conclude that AVR3a is a multifunctional effector that can suppress BAK1/SERK3-mediated immunity through at least two different pathways.

  20. The N-terminal portion of domain E of retinoic acid receptors alpha and beta is essential for the recognition of retinoic acid and various analogs.

    PubMed Central

    Ostrowski, J; Hammer, L; Roalsvig, T; Pokornowski, K; Reczek, P R

    1995-01-01

    Utilizing a strategy involving domain exchange between retinoic acid receptors alpha and beta (RAR alpha and RAR beta) and monitoring the transcriptional activity of the resulting chimeric receptors with receptor-selective retinoids, we identified a 70-aa region within the N-terminal portion of the RAR alpha and -beta domain E which is important for an RAR alpha- or RAR beta-specific response. Two amino acid residues within this region, serine-232 (S232) and threonine-239 (T239) in RAR alpha and the corresponding alanine-225 (A225) and isoleucine-232 (I232) in RAR beta, were found to be essential for this effect. In addition, binding studies using the chimeric receptors expressed in Escherichia coli showed that the N-terminal portion of domain E was also important for the characteristic binding profile of t-RA and various retinoids with RAR alpha or RAR beta. Structural predictions of the primary amino acid sequence in this region indicate the presence of an amphipathic helix-turn-helix structure with five hydrophobic amino acids that resemble a leucine zipper motif. The amino acid residues identified by domain swapping, S232 and T239 in RAR alpha and A225 and I232 in RAR beta, were found within the hydrophobic face of an alpha-helix in close proximity to this zipper motif, suggesting that the ligand may interact with the receptor in the region adjacent to a surface involved in protein-protein interactions. This finding may link ligand binding to other processes important for transcriptional activation. PMID:7892182

  1. The Molecular Chaperone HSPA2 Plays a Key Role in Regulating the Expression of Sperm Surface Receptors That Mediate Sperm-Egg Recognition

    PubMed Central

    Redgrove, Kate A.; Nixon, Brett; Baker, Mark A.; Hetherington, Louise; Baker, Gordon; Liu, De-Yi; Aitken, R. John

    2012-01-01

    A common defect encountered in the spermatozoa of male infertility patients is an idiopathic failure of sperm–egg recognition. In order to resolve the molecular basis of this condition we have compared the proteomic profiles of spermatozoa exhibiting an impaired capacity for sperm-egg recognition with normal cells using label free mass spectrometry (MS)-based quantification. This analysis indicated that impaired sperm–zona binding was associated with reduced expression of the molecular chaperone, heat shock 70 kDa protein 2 (HSPA2), from the sperm proteome. Western blot analysis confirmed this observation in independent patients and demonstrated that the defect did not extend to other members of the HSP70 family. HSPA2 was present in the acrosomal domain of human spermatozoa as a major component of 5 large molecular mass complexes, the most dominant of which was found to contain HSPA2 in close association with just two other proteins, sperm adhesion molecule 1 (SPAM1) and arylsulfatase A (ARSA), both of which that have previously been implicated in sperm-egg interaction. The interaction between SPAM1, ARSA and HSPA2 in a multimeric complex mediating sperm-egg interaction, coupled with the complete failure of this process when HSPA2 is depleted in infertile patients, provides new insights into the mechanisms by which sperm function is impaired in cases of male infertility. PMID:23209833

  2. The molecular chaperone HSPA2 plays a key role in regulating the expression of sperm surface receptors that mediate sperm-egg recognition.

    PubMed

    Redgrove, Kate A; Nixon, Brett; Baker, Mark A; Hetherington, Louise; Baker, Gordon; Liu, De-Yi; Aitken, R John

    2012-01-01

    A common defect encountered in the spermatozoa of male infertility patients is an idiopathic failure of sperm-egg recognition. In order to resolve the molecular basis of this condition we have compared the proteomic profiles of spermatozoa exhibiting an impaired capacity for sperm-egg recognition with normal cells using label free mass spectrometry (MS)-based quantification. This analysis indicated that impaired sperm-zona binding was associated with reduced expression of the molecular chaperone, heat shock 70 kDa protein 2 (HSPA2), from the sperm proteome. Western blot analysis confirmed this observation in independent patients and demonstrated that the defect did not extend to other members of the HSP70 family. HSPA2 was present in the acrosomal domain of human spermatozoa as a major component of 5 large molecular mass complexes, the most dominant of which was found to contain HSPA2 in close association with just two other proteins, sperm adhesion molecule 1 (SPAM1) and arylsulfatase A (ARSA), both of which that have previously been implicated in sperm-egg interaction. The interaction between SPAM1, ARSA and HSPA2 in a multimeric complex mediating sperm-egg interaction, coupled with the complete failure of this process when HSPA2 is depleted in infertile patients, provides new insights into the mechanisms by which sperm function is impaired in cases of male infertility.

  3. Design, synthesis and 1H NMR study of C3v-symmetric anion receptors with urethane-NH as recognition group

    NASA Astrophysics Data System (ADS)

    Park, Jin-Oh; Sahoo, Suban K.; Choi, Heung-Jin

    2016-01-01

    C3v-Symmetric anion receptors 3 and 4 with urethane groups were synthesized by using trindane triol as tripodal molecular framework. In 1H NMR titration study, the receptors showed noticeable downfield shift/disappearance of the urethane-NH peak in presence of H2PO4- and F- due to the host-guest complexation occurred through multiple hydrogen bonding and/or the deprotonation of urethane-NH groups. Other tested anions such as Cl-, Br-, HSO4-, and NO3- showed either no or negligible chemical shift of the urethane groups. The deprotonation event in 4 allowed selective detection of F- by perceptible color change from colorless to yellowish-red with the appearance of a new charge transfer absorption band at 450 nm.

  4. Conjoint recognition.

    PubMed

    Brainerd, C J; Reyna, V F; Mojardin, A H

    1999-01-01

    The process-dissociation model has stimulated important advances in the study of dual-process conceptions of memory. The authors review some limiting properties of that model and consider the degree of support for its parent theory (the recollection-familiarity distinction). A 2nd-generation model (conjoint recognition) is proposed that addresses these limitations and supplies additional capabilities, such as goodness-of-fit tests, the ability to measure dual processes for false-memory responses, and statistical procedures for testing within- and between-conditions hypotheses about its parameters. The conjoint-recognition model also implements an alternative theoretical interpretation (the identity-similarity distinction of fuzzy-trace theory). Worked applications to data are provided.

  5. Dynamic chemistry of anion recognition

    SciTech Connect

    Custelcean, Radu

    2012-01-01

    In the past 40 years, anion recognition by synthetic receptors has grown into a rich and vibrant research topic, developing into a distinct branch of Supramolecular Chemistry. Traditional anion receptors comprise organic scaffolds functionalized with complementary binding groups that are assembled by multistep organic synthesis. Recently, a new approach to anion receptors has emerged, in which the host is dynamically self-assembled in the presence of the anionic guest, via reversible bond formation between functional building units. While coordination bonds were initially employed for the self-assembly of the anion hosts, more recent studies demonstrated that reversible covalent bonds can serve the same purpose. In both cases, due to their labile connections, the molecular constituents have the ability to assemble, dissociate, and recombine continuously, thereby creating a dynamic combinatorial library (DCL) of receptors. The anionic guests, through specific molecular recognition, may then amplify (express) the formation of a particular structure among all possible combinations (real or virtual) by shifting the equilibria involved towards the most optimal receptor. This approach is not limited to solution self-assembly, but is equally applicable to crystallization, where the fittest anion-binding crystal may be selected. Finally, the pros and cons of employing dynamic combinatorial chemistry (DCC) vs molecular design for developing anion receptors, and the implications of both approaches to selective anion separations, will be discussed.

  6. Magnesium modification up-regulates the bioactivity of bone morphogenetic protein-2 upon calcium phosphate cement via enhanced BMP receptor recognition and Smad signaling pathway.

    PubMed

    Ding, Sai; Zhang, Jing; Tian, Yu; Huang, Baolin; Yuan, Yuan; Liu, Changsheng

    2016-09-01

    Efficient presentation of growth factors is one of the great challenges in tissue engineering. In living systems, bioactive factors exist in soluble as well as in matrix-bound forms, both of which play an integral role in regulating cell behaviors. Herein, effect of magnesium on osteogenic bioactivity of recombinant human bone morphogenetic protein-2 (rhBMP-2) was investigated systematically with a series of Mg modified calcium phosphate cements (xMCPCs, x means the content of magnesium phosphate cement wt%) as matrix model. The results indicated that the MCPC, especially 5MCPC, could promote the rhBMP-2-induced in vitro osteogenic differentiation via Smad signaling of C2C12 cells. Further studies demonstrated that all MCPC substrates exhibited similar rhBMP-2 release rate and preserved comparable conformation and biological activity of the released rhBMP-2. Also, the ionic extracts of MCPC made little difference to the bioactivity of rhBMP-2, either in soluble or in matrix-bound forms. However, with the quartz crystal microbalance (QCM), we observed a noticeable enhancement of rhBMP-2 mass-uptake on 5MCPC as well as a better recognition of the bound rhBMP-2 to BMPR IA and BMPR II. In vivo results demonstrated a better bone regeneration capacity of 5MCPC/rhBMP-2. From the above, our results demonstrated that it was the Mg anchored on the underlying substrates that tailored the way of rhBMP-2 bound on MCPC, and thus facilitated the recognition of BMPRs to stimulate osteogenic differentiation. The study will guide the development of Mg-doped bioactive bone implants for tissue regeneration.

  7. The pattern recognition receptor CD36 is a chondrocyte hypertrophy marker associated with suppression of catabolic responses and promotion of repair responses to inflammatory stimuli.

    PubMed

    Cecil, Denise L; Appleton, C Thomas G; Polewski, Monika D; Mort, John S; Schmidt, Ann Marie; Bendele, Alison; Beier, Frank; Terkeltaub, Robert

    2009-04-15

    Multiple inflammatory mediators in osteoarthritis (OA) cartilage, including S100/calgranulin ligands of receptor for advanced glycation end products (RAGE), promote chondrocyte hypertrophy, a differentiation state associated with matrix catabolism. In this study, we observed that RAGE knockout was not chondroprotective in instability-induced knee OA in 8-wk-old mice. Hence, we tested the hypothesis that expression of the alternative S100/calgranulin and patterning receptor CD36, identified here as a marker of growth plate chondrocyte hypertrophy, mediates chondrocyte inflammatory and differentiation responses that promote OA. In rat knee joint destabilization-induced OA, RAGE expression was initially sparse throughout cartilage but increased diffusely by 4 wk after surgery. In contrast, CD36 expression focally increased at sites of cartilage injury and colocalized with developing chondrocyte hypertrophy and aggrecan cleavage NITEGE neoepitope formation. However, CD36 transfection in normal human knee-immortalized chondrocytes (CH-8 cells) was associated with decreased capacity of S100A11 and TNF-alpha to induce chondrocyte hypertrophy and ADAMTS-4 and matrix metalloproteinase 13 expression. S100A11 lost the capacity to inhibit proteoglycans synthesis and gained the capacity to induce proteoglycan synthesis in CD36-transfected CH-8 cells. Moreover, S100A11 required the p38 MAPK pathway kinase MKK3 to induce NITEGE development in mouse articular cartilage explants. However, CH-8 cells transfected with CD36 demonstrated decreased S100A11-induced MKK3 and p38 phosphorylation. Therefore, RAGE and CD36 patterning receptor expression were linked with opposing effects on inflammatory, procatabolic responses to S100A11 and TNF-alpha in chondrocytes.

  8. Histone recognition and nuclear receptor co-activator functions of Drosophila Cara Mitad, a homolog of the N-terminal portion of mammalian MLL2 and MLL3

    PubMed Central

    Chauhan, Chhavi; Zraly, Claudia B.; Parilla, Megan; Diaz, Manuel O.; Dingwall, Andrew K.

    2012-01-01

    MLL2 and MLL3 histone lysine methyltransferases are conserved components of COMPASS-like co-activator complexes. In vertebrates, the paralogous MLL2 and MLL3 contain multiple domains required for epigenetic reading and writing of the histone code involved in hormone-stimulated gene programming, including receptor-binding motifs, SET methyltransferase, HMG and PHD domains. The genes encoding MLL2 and MLL3 arose from a common ancestor. Phylogenetic analyses reveal that the ancestral gene underwent a fission event in some Brachycera dipterans, including Drosophila species, creating two independent genes corresponding to the N- and C-terminal portions. In Drosophila, the C-terminal SET domain is encoded by trithorax-related (trr), which is required for hormone-dependent gene activation. We identified the cara mitad (cmi) gene, which encodes the previously undiscovered N-terminal region consisting of PHD and HMG domains and receptor-binding motifs. The cmi gene is essential and its functions are dosage sensitive. CMI associates with TRR, as well as the EcR-USP receptor, and is required for hormone-dependent transcription. Unexpectedly, although the CMI and MLL2 PHDf3 domains could bind histone H3, neither showed preference for trimethylated lysine 4. Genetic tests reveal that cmi is required for proper global trimethylation of H3K4 and that hormone-stimulated transcription requires chromatin binding by CMI, methylation of H3K4 by TRR and demethylation of H3K27 by the demethylase UTX. The evolutionary split of MLL2 into two distinct genes in Drosophila provides important insight into distinct epigenetic functions of conserved readers and writers of the histone code. PMID:22569554

  9. Histone recognition and nuclear receptor co-activator functions of Drosophila cara mitad, a homolog of the N-terminal portion of mammalian MLL2 and MLL3.

    PubMed

    Chauhan, Chhavi; Zraly, Claudia B; Parilla, Megan; Diaz, Manuel O; Dingwall, Andrew K

    2012-06-01

    MLL2 and MLL3 histone lysine methyltransferases are conserved components of COMPASS-like co-activator complexes. In vertebrates, the paralogous MLL2 and MLL3 contain multiple domains required for epigenetic reading and writing of the histone code involved in hormone-stimulated gene programming, including receptor-binding motifs, SET methyltransferase, HMG and PHD domains. The genes encoding MLL2 and MLL3 arose from a common ancestor. Phylogenetic analyses reveal that the ancestral gene underwent a fission event in some Brachycera dipterans, including Drosophila species, creating two independent genes corresponding to the N- and C-terminal portions. In Drosophila, the C-terminal SET domain is encoded by trithorax-related (trr), which is required for hormone-dependent gene activation. We identified the cara mitad (cmi) gene, which encodes the previously undiscovered N-terminal region consisting of PHD and HMG domains and receptor-binding motifs. The cmi gene is essential and its functions are dosage sensitive. CMI associates with TRR, as well as the EcR-USP receptor, and is required for hormone-dependent transcription. Unexpectedly, although the CMI and MLL2 PHDf3 domains could bind histone H3, neither showed preference for trimethylated lysine 4. Genetic tests reveal that cmi is required for proper global trimethylation of H3K4 and that hormone-stimulated transcription requires chromatin binding by CMI, methylation of H3K4 by TRR and demethylation of H3K27 by the demethylase UTX. The evolutionary split of MLL2 into two distinct genes in Drosophila provides important insight into distinct epigenetic functions of conserved readers and writers of the histone code.

  10. Receptors for oxidized low-density lipoprotein on elicited mouse peritoneal macrophages can recognize both the modified lipid moieties and the modified protein moieties: Implications with respect to macrophage recognition of apoptotic cells

    PubMed Central

    Bird, David A.; Gillotte, Kristin L.; Hörkkö, Sohvi; Friedman, Peter; Dennis, Edward A.; Witztum, Joseph L.; Steinberg, Daniel

    1999-01-01

    It has been shown previously that the binding of oxidized low-density lipoprotein (OxLDL) to resident mouse peritoneal macrophages can be inhibited (up to 70%) by the apoprotein B (apoB) isolated from OxLDL, suggesting that macrophage recognition of OxLDL is primarily dependent on its modified protein moiety. However, recent experiments have demonstrated that the lipids isolated from OxLDL and reconstituted into a microemulsion can also strongly inhibit uptake of OxLDL (up to 80%). The present studies show that lipid microemulsions prepared from OxLDL bind to thioglycollate-elicited macrophages at 4°C in a saturable fashion and inhibit the binding of intact OxLDL and also of the apoB from OxLDL. Reciprocally, the binding of the OxLDL-lipid microemulsions was strongly inhibited by intact OxLDL. A conjugate of synthetic 1-palmitoyl 2(5-oxovaleroyl) phosphatidylcholine (an oxidation product of 1-palmitoyl 2-arachidonoyl phosphatidylcholine) with serum albumin, shown previously to inhibit macrophage binding of intact OxLDL, also inhibited the binding of both the apoprotein and the lipid microemulsions prepared from OxLDL. Finally, a monoclonal antibody against oxidized phospholipids, one that inhibits binding of intact OxLDL to macrophages, also inhibited the binding of both the resolubilized apoB and the lipid microemulsions prepared from OxLDL. These studies support the conclusions that: (i) at least some of the macrophage receptors for oxidized LDL can recognize both the lipid and the protein moieties; and (ii) oxidized phospholipids, in the lipid phase of the lipoprotein and/or covalently linked to the apoB of OxLDL, likely play a role in that recognition. PMID:10339590

  11. LDL receptor/lipoprotein recognition: endosomal weakening of ApoB and ApoE binding to the convex face of the LR5 repeat.

    PubMed

    Martínez-Oliván, Juan; Arias-Moreno, Xabier; Velazquez-Campoy, Adrián; Millet, Oscar; Sancho, Javier

    2014-03-01

    The molecular mechanism of lipoprotein binding by the low-density lipoprotein (LDL) receptor (LDLR) is poorly understood, one reason being that structures of lipoprotein-receptor complexes are not available. LDLR uses calcium-binding repeats (LRs) to interact with apolipoprotein B and apolipoprotein E (ApoB and ApoE). We have used NMR and SPR to characterize the complexes formed by LR5 and three peptides encompassing the putative binding regions of ApoB (site A and site B) and ApoE. The three peptides bind at the hydrophilic convex face of LR5, forming complexes that are weakened at low [Ca(2+) ] and low pH. Thus, endosomal conditions favour dissociation of LDLR/lipoprotein complexes regardless of whether active displacement of bound lipoproteins by the β-propeller in LDLR takes place. The multiple ApoE copies in β very low density lipoproteins (β-VLDLs), and the presence of two competent binding sites (A and B) in LDLs, suggest that LDLR chelates lipoproteins and enhances complex affinity by using more than one LR.

  12. The Inorganic Side of NGF: Copper(II) and Zinc(II) Affect the NGF Mimicking Signaling of the N-Terminus Peptides Encompassing the Recognition Domain of TrkA Receptor

    PubMed Central

    Pandini, Giuseppe; Satriano, Cristina; Pietropaolo, Adriana; Gianì, Fiorenza; Travaglia, Alessio; La Mendola, Diego; Nicoletti, Vincenzo G.; Rizzarelli, Enrico

    2016-01-01

    The nerve growth factor (NGF) N-terminus peptide, NGF(1–14), and its acetylated form, Ac-NGF(1–14), were investigated to scrutinize the ability of this neurotrophin domain to mimic the whole protein. Theoretical calculations demonstrated that non-covalent forces assist the molecular recognition of TrkA receptor by both peptides. Combined parallel tempering/docking simulations discriminated the effect of the N-terminal acetylation on the recognition of NGF(1–14) by the domain 5 of TrkA (TrkA-D5). Experimental findings demonstrated that both NGF(1–14) and Ac-NGF(1–14) activate TrkA signaling pathways essential for neuronal survival. The NGF-induced TrkA internalization was slightly inhibited in the presence of Cu2+ and Zn2+ ions, whereas the metal ions elicited the NGF(1–14)-induced internalization of TrkA and no significant differences were found in the weak Ac-NGF(1–14)-induced receptor internalization. The crucial role of the metals was confirmed by experiments with the metal-chelator bathocuproine disulfonic acid, which showed different inhibitory effects in the signaling cascade, due to different metal affinity of NGF, NGF(1–14) and Ac-NGF(1–14). The NGF signaling cascade, activated by the two peptides, induced CREB phosphorylation, but the copper addition further stimulated the Akt, ERK and CREB phosphorylation in the presence of NGF and NGF(1–14) only. A dynamic and quick influx of both peptides into PC12 cells was tracked by live cell imaging with confocal microscopy. A significant role of copper ions was found in the modulation of peptide sub-cellular localization, especially at the nuclear level. Furthermore, a strong copper ionophoric ability of NGF(1–14) was measured. The Ac-NGF(1–14) peptide, which binds copper ions with a lower stability constant than NGF(1–14), exhibited a lower nuclear localization with respect to the total cellular uptake. These findings were correlated to the metal-induced increase of CREB and BDNF

  13. Structural Basis for the Recognition of Mutant Self by a Tumor-Specific, MHC Class II-Restricted T Cell Receptor

    SciTech Connect

    Deng,L.; Langley, R.; Brown, P.; Xu, G.; Teng, L.; Wang, Q.; Gonzales, M.; Callender, G.; Nishimura, M.; et al.

    2007-01-01

    Structural studies of complexes of T cell receptor (TCR) and peptide-major histocompatibility complex (MHC) have focused on TCRs specific for foreign antigens or native self. An unexplored category of TCRs includes those specific for self determinants bearing alterations resulting from disease, notably cancer. We determined here the structure of a human melanoma-specific TCR (E8) bound to the MHC molecule HLA-DR1 and an epitope from mutant triosephosphate isomerase. The structure had features intermediate between 'anti-foreign' and autoimmune TCR-peptide-MHC class II complexes that may reflect the hybrid nature of altered self. E8 manifested very low affinity for mutant triosephosphate isomerase-HLA-DR1 despite the highly tumor-reactive properties of E8 cells. A second TCR (G4) had even lower affinity but underwent peptide-specific formation of dimers, suggesting this as a mechanism for enhancing low-affinity TCR-peptide-MHC interactions for T cell activation.

  14. Inhibition of the prostaglandin E2 receptor EP2 prevents status epilepticus-induced deficits in the novel object recognition task in rats.

    PubMed

    Rojas, Asheebo; Ganesh, Thota; Manji, Zahra; O'neill, Theon; Dingledine, Raymond

    2016-11-01

    Survivors of exposure to an organophosphorus nerve agent may develop a number of complications including long-term cognitive deficits (Miyaki et al., 2005; Nishiwaki et al., 2001). We recently demonstrated that inhibition of the prostaglandin E2 receptor, EP2, attenuates neuroinflammation and neurodegeneration caused by status epilepticus (SE) induced by the soman analog, diisopropylfluorophosphate (DFP), which manifest within hours to days of the initial insult. Here, we tested the hypothesis that DFP exposure leads to a loss of cognitive function in rats that is blocked by early, transient EP2 inhibition. Adult male Sprague-Dawley rats were administered vehicle or the competitive EP2 antagonist, TG6-10-1, (ip) at various times relative to DFP-induced SE. DFP administration resulted in prolonged seizure activity as demonstrated by cortical electroencephalography (EEG). A single intraperitoneal injection of TG6-10-1 or vehicle 1 h prior to DFP did not alter the development of seizures, the latency to SE or the duration of SE. Rats administered six injections of TG6-10-1 starting 90 min after the onset of DFP-induced SE could discriminate between a novel and familiar object 6-12 weeks after SE, unlike vehicle treated rats which showed no preference for the novel object. By contrast, behavioral changes in the light-dark box and open field assays were not affected by TG6-10-1. Delayed mortality after DFP was also unaffected by TG6-10-1. Thus, selective inhibition of the EP2 receptor may prevent SE-induced memory impairment in rats caused by exposure to a high dose of DFP.

  15. Association of genetic variants of membrane receptors related to recognition and induction of immune response with Helicobacter pylori infection in Ecuadorian individuals.

    PubMed

    Cabrera-Andrade, A; López-Cortés, A; Muñoz, M J; Jaramillo-Koupermann, G; Rodriguez, O; Leone, P E; Paz-y-Miño, C

    2014-08-01

    Helicobacter pylori (Hp) has a worldwide distribution showing its higher prevalence of infection in developing countries. Toll-like receptors (TLRs) and C-type lectin receptors (CLRs) are proteins that recognize pathogen-associated molecular patterns (PAMPs) and initiate an innate immune response by promoting growth and differentiation of specialized hematopoietic cells for host defense. Gastric infections led by Hp induce a Th-1 cellular immune response, regulated mainly by the expression of IFN-γ. In this retrospective case-control study, we evaluated the TLR1 1805T/G, TLR2 2029C/T, TLR4 896A/G, CD209 -336A/G and IFNGR1 -56C/T polymorphisms and their relationship with susceptibility to Hp infection. TLR1 1805T/G showed statistical differences when the control (Hp-) and infected (Hp+) groups (P = 0.041*) were compared; the TLR1 1805G allele had a protective effect towards infection (OR = 0.1; 95% CI = 0.01-0.88, P = 0.033*). Similarly, the IFNGR1 -56C/T polymorphism showed statistical differences between Hp+ and Hp- (P = 0.018*), and the IFNGR1 -56TT genotype exhibited significant risk to Hp infection (OR = 2.9, 95% CI = 1.27-6.54, P = 0.018*). In conclusion, the pro-inflammatory TLR1 1805T and IFNGR1 -56T alleles are related with susceptibility to Hp infection in Ecuadorian individuals. The presence of these polymorphisms in individuals with chronic infection increases the risk of cellular damage and diminishes the cellular immune response efficiency towards colonizing agents.

  16. 3-(1H-indol-3-yl)-2-[3-(4-nitrophenyl)ureido]propanamide enantiomers with human formyl-peptide receptor agonist activity: molecular modeling of chiral recognition by FPR2.

    PubMed

    Schepetkin, Igor A; Kirpotina, Liliya N; Khlebnikov, Andrei I; Leopoldo, Marcello; Lucente, Ermelinda; Lacivita, Enza; De Giorgio, Paola; Quinn, Mark T

    2013-02-01

    N-formyl peptide receptors (FPRs) are G protein-coupled receptors (GPCRs) that play critical roles in inflammatory reactions, and FPR-specific interactions can possibly be used to facilitate the resolution of pathological inflammatory reactions. Recent studies indicated that FPRs have stereo-selective preference for chiral ligands. Here, we investigated the structure-activity relationship of 24 chiral ureidopropanamides, including previously reported compounds PD168368/PD176252 and their close analogs, and used molecular modeling to define chiral recognition by FPR2. Unlike previously reported 6-methyl-2,4-disubstituted pyridazin-3(2H)-ones, whose R-forms preferentially activated FPR1/FPR2, we found that four S-enantiomers in the seven ureidopropanamide pairs tested preferentially activated intracellular Ca(2+) flux in FPR2-transfected cells, while the R-counterpart was more active in two enantiomer pairs. Thus, active enantiomers of FPR2 agonists can be in either R- or S-configurations, depending on the molecular scaffold and specific substituents at the chiral center. Using molecular modeling approaches, including field point methodology, homology modeling, and docking studies, we propose a model that can explain stereoselective activity of chiral FPR2 agonists. Importantly, our docking studies of FPR2 chiral agonists correlated well with the FPR2 pharmacophore model derived previously. We conclude that the ability of FPR2 to discriminate between the enantiomers is the consequence of the arrangement of the three asymmetric hydrophobic subpockets at the main orthosteric FPR2 binding site with specific orientation of charged regions in the subpockets.

  17. 3-(1H-Indol-3-yl)-2-[3-(4-nitrophenyl)ureido]propanamide Enantiomers With Human Formyl-Peptide Receptor Agonist Activity: Molecular Modeling of Chiral Recognition by FPR2

    PubMed Central

    Schepetkin, Igor A.; Kirpotina, Liliya N.; Khlebnikov, Andrei I.; Leopoldo, Marcello; Lucente, Ermelinda; Lacivita, Enza; De Giorgio, Paola; Quinn, Mark T.

    2012-01-01

    N-formyl peptide receptors (FPRs) are G protein-coupled receptors (GPCRs) that play critical roles in inflammatory reactions, and FPR-specific interactions can possibly be used to facilitate the resolution of pathological inflammatory reactions. Recent studies indicated that FPRs have stereo-selective preference for chiral ligands. Here, we investigated the structure-activity relationship of 24 chiral ureidopropanamides, including previously reported compounds PD168368/PD176252 and their close analogs, and used molecular modeling to define chiral recognition by FPR2. Unlike previously reported 6-methyl-2,4-disubstituted pyridazin-3(2H)-ones, whose R-forms preferentially activated FPR1/FPR2, we found that four S-enantiomers in the seven ureidopropanamide pairs tested preferentially activated intracellular Ca2+ flux in FPR2-transfected cells, while the R-counterpart was more active in two enantiomer pairs. Thus, active enantiomers of FPR2 agonists can be in either R- or S- configurations, depending on the molecular scaffold and specific substituents at the chiral center. Using molecular modeling approaches, including field point methodology, homology modeling, and docking studies, we propose a model that can explain stereoselective activity of chiral FPR2 agonists. Importantly, our docking studies of FPR2 chiral agonists correlated well with the FPR2 pharmacophore model derived previously. We conclude that the ability of FPR2 to discriminate between the enantiomers is the consequence of the arrangement of the three asymmetric hydrophobic subpockets at the main orthosteric FPR2 binding site with specific orientation of charged regions in the subpockets. PMID:23219934

  18. Recognition Tunneling

    PubMed Central

    Lindsay, Stuart; He, Jin; Sankey, Otto; Hapala, Prokop; Jelinek, Pavel; Zhang, Peiming; Chang, Shuai; Huang, Shuo

    2010-01-01

    Single molecules in a tunnel junction can now be interrogated reliably using chemically-functionalized electrodes. Monitoring stochastic bonding fluctuations between a ligand bound to one electrode and its target bound to a second electrode (“tethered molecule-pair” configuration) gives insight into the nature of the intermolecular bonding at a single molecule-pair level, and defines the requirements for reproducible tunneling data. Simulations show that there is an instability in the tunnel gap at large currents, and this results in a multiplicity of contacts with a corresponding spread in the measured currents. At small currents (i.e. large gaps) the gap is stable, and functionalizing a pair of electrodes with recognition reagents (the “free analyte” configuration) can generate a distinct tunneling signal when an analyte molecule is trapped in the gap. This opens up a new interface between chemistry and electronics with immediate implications for rapid sequencing of single DNA molecules. PMID:20522930

  19. Beta-glucan recognition by the innate immune system.

    PubMed

    Goodridge, Helen S; Wolf, Andrea J; Underhill, David M

    2009-07-01

    Beta-glucans are recognized by the innate immune system. This recognition plays important roles in host defense and presents specific opportunities for clinical modulation of the host immune response. Neutrophils, macrophages, and dendritic cells among others express several receptors capable of recognizing beta-glucan in its various forms. This review explores what is currently known about beta-glucan recognition and how this recognition stimulates immune responses. Special emphasis is placed on Dectin-1, as we know the most about how this key beta-glucan receptor translates recognition into intracellular signaling, stimulates cellular responses, and participates in orchestrating the adaptive immune response.

  20. Differential Recognition of CD1d-[alpha]-Galactosyl Ceramide by the V[beta]8.2 and V[beta]7 Semi-invariant NKT T Cell Receptors

    SciTech Connect

    Pellicci, Daniel G.; Patel, Onisha; Kjer-Nielsen, Lars; Pang, Siew Siew; Sullivan, Lucy C.; Kyparissoudis, Konstantinos; Brooks, Andrew G.; Reid, Hugh H.; Gras, Stephanie; Lucet, Isabelle S.; Koh, Ruide; Smyth, Mark J.; Mallevaey, Thierry; Matsuda, Jennifer L.; Gapin, Laurent; McCluskey, James; Godfrey, Dale I.; Rossjohn, Jamie; PMCI-A; Monash; UCHSC; Melbourne

    2009-09-02

    The semi-invariant natural killer T cell receptor (NKT TCR) recognizes CD1d-lipid antigens. Although the TCR{alpha} chain is typically invariant, the {beta} chain expression is more diverse, where three V{beta} chains are commonly expressed in mice. We report the structures of V{alpha}14-V{beta}8.2 and V{alpha}14-V{beta}7 NKT TCRs in complex with CD1d-{alpha}-galactosylceramide ({alpha}-GalCer) and the 2.5 {angstrom} structure of the human NKT TCR-CD1d-{alpha}-GalCer complex. Both V{beta}8.2 and V{beta}7 NKT TCRs and the human NKT TCR ligated CD1d-{alpha}-GalCer in a similar manner, highlighting the evolutionarily conserved interaction. However, differences within the V{beta} domains of the V{beta}8.2 and V{beta}7 NKT TCR-CD1d complexes resulted in altered TCR{beta}-CD1d-mediated contacts and modulated recognition mediated by the invariant {alpha} chain. Mutagenesis studies revealed the differing contributions of V{beta}8.2 and V{beta}7 residues within the CDR2{beta} loop in mediating contacts with CD1d. Collectively we provide a structural basis for the differential NKT TCR V{beta} usage in NKT cells.

  1. High molecular recognition: design of "Keys".

    PubMed

    Chen, Beining; Piletsky, Sergey; Turner, Anthony P F

    2002-09-01

    Molecular recognition between molecules is one of the most fundamental processes in biology and chemistry. The recognition process is largely driven by non-covalent forces such as hydrogen bonding, electrostatics, van der Waals forces, pi-pi interactions, and conformational energy. The complementarity between the receptor and substrate is very similar to the "lock and key" function, first described by Emil Fischer over 100 years ago, - the lock being the molecular receptor such as a protein or enzyme and the key being the substrate such as a drug, that is recognized to give a defined receptor-substrate complex. This review focuses on the design of specific ligand systems as "Keys" to enable the induced fit of these keys into the target macromolecules, protein/enzyme (Locks) with particular emphasis on protein recognition.

  2. A C1q Domain Containing Protein from Scallop Chlamys farreri Serving as Pattern Recognition Receptor with Heat-Aggregated IgG Binding Activity

    PubMed Central

    Wang, Leilei; Wang, Lingling; Zhang, Huan; Zhou, Zhi; Siva, Vinu S.; Song, Linsheng

    2012-01-01

    Background The C1q domain containing (C1qDC) proteins refer to a family of all proteins that contain the globular C1q (gC1q) domain, and participate in a series of immune responses depending on their gC1q domains to bind a variety of self and non-self binding ligands. Methodology In the present study, the mRNA expression patterns, localization, and activities of a C1qDC protein from scallop Chlamys farreri (CfC1qDC) were investigated to understand its possible functions in innate immunity. The relative expression levels of CfC1qDC mRNA in hemocytes were all significantly up-regulated after four typical PAMPs (LPS, PGN, β-glucan and polyI:C) stimulation. During the embryonic development of scallop, the mRNA transcripts of CfC1qDC were detected in all the stages, and the expression level was up-regulated from D-hinged larva and reached the highest at eye-spot larva. The endogenous CfC1qDC was dominantly located in the hepatopancreas, gill, kidney and gonad of adult scallop through immunofluorescence. The recombinant protein of CfC1qDC (rCfC1qDC) could not only bind various PAMPs, such as LPS, PGN, β-glucan as well as polyI:C, but also enhance the phagocytic activity of scallop hemocytes towards Escherichia coli. Meanwhile, rCfC1qDC could interact with human heat-aggregated IgG, and this interaction could be inhibited by LPS. Conclusions All these results indicated that CfC1qDC in C. farreri not only served as a PRR involved in the PAMPs recognition, but also an opsonin participating in the clearance of invaders in innate immunity. Moreover, the ability of CfC1qDC to interact with immunoglobulins provided a clue to understand the evolution of classical pathway in complement system. PMID:22905248

  3. Tick receptor for outer surface protein A from Ixodes ricinus — the first intrinsically disordered protein involved in vector-microbe recognition

    NASA Astrophysics Data System (ADS)

    Urbanowicz, Anna; Lewandowski, Dominik; Szpotkowski, Kamil; Figlerowicz, Marek

    2016-04-01

    The tick receptor for outer surface protein A (TROSPA) is the only identified factor involved in tick gut colonization by various Borrelia species. TROSPA is localized in the gut epithelium and can recognize and bind the outer surface bacterial protein OspA via an unknown mechanism. Based on earlier reports and our latest observations, we considered that TROSPA would be the first identified intrinsically disordered protein (IDP) involved in the interaction between a vector and a pathogenic microbe. To verify this hypothesis, we performed structural studies of a TROSPA mutant from Ixodes ricinus using both computational and experimental approaches. Irrespective of the method used, we observed that the secondary structure content of the TROSPA polypeptide chain is low. In addition, the collected SAXS data indicated that this protein is highly extended and exists in solution as a set of numerous conformers. These features are all commonly considered hallmarks of IDPs. Taking advantage of our SAXS data, we created structural models of TROSPA and proposed a putative mechanism for the TROSPA-OspA interaction. The disordered nature of TROSPA may explain the ability of a wide spectrum of Borrelia species to colonize the tick gut.

  4. Tick receptor for outer surface protein A from Ixodes ricinus — the first intrinsically disordered protein involved in vector-microbe recognition

    PubMed Central

    Urbanowicz, Anna; Lewandowski, Dominik; Szpotkowski, Kamil; Figlerowicz, Marek

    2016-01-01

    The tick receptor for outer surface protein A (TROSPA) is the only identified factor involved in tick gut colonization by various Borrelia species. TROSPA is localized in the gut epithelium and can recognize and bind the outer surface bacterial protein OspA via an unknown mechanism. Based on earlier reports and our latest observations, we considered that TROSPA would be the first identified intrinsically disordered protein (IDP) involved in the interaction between a vector and a pathogenic microbe. To verify this hypothesis, we performed structural studies of a TROSPA mutant from Ixodes ricinus using both computational and experimental approaches. Irrespective of the method used, we observed that the secondary structure content of the TROSPA polypeptide chain is low. In addition, the collected SAXS data indicated that this protein is highly extended and exists in solution as a set of numerous conformers. These features are all commonly considered hallmarks of IDPs. Taking advantage of our SAXS data, we created structural models of TROSPA and proposed a putative mechanism for the TROSPA-OspA interaction. The disordered nature of TROSPA may explain the ability of a wide spectrum of Borrelia species to colonize the tick gut. PMID:27112540

  5. Unusual Recognition and Separation of Hydrated Metal Sulfates [M2(μ-SO4)2(H2O)n, M = Zn(II), Cd(II), Co(II), Mn(II)] by a Ditopic Receptor.

    PubMed

    Ghosh, Tamal Kanti; Dutta, Ranjan; Ghosh, Pradyut

    2016-04-04

    A ditopic receptor L1, having metal binding bis(2-picolyl) donor and anion binding urea group, is synthesized and explored toward metal sulfate recognition via formation of dinuclear assembly, (L1)2M2(SO4)2. Mass spectrometric analysis, (1)H-DOSY NMR, and crystal structure analysis reveal the existence of a dinuclear assembly of MSO4 with two units of L1. (1)H NMR study reveals significant downfield chemical shift of -NH protons of urea moiety of L1 selectively with metal sulfates (e.g., ZnSO4, CdSO4) due to second-sphere interactions of sulfate with the urea moiety. Variable-temperature (1)H NMR studies suggest the presence of intramolecular hydrogen bonding interaction toward metal sulfate recognition in solution state, whereas intermolecular H-bonding interactions are observed in solid state. In contrast, anions in their tetrabutylammonium salts fail to interact with the urea -NH probably due to poor acidity of the tertiary butyl urea group of L1. Metal sulfate binding selectivity in solution is further supported by isothermal titration calorimetric studies of L1 with different Zn salts in dimethyl sulfoxide (DMSO), where a binding affinity is observed for ZnSO4 (Ka = 1.23 × 10(6)), which is 30- to 50-fold higher than other Zn salts having other counteranions in DMSO. Sulfate salts of Cd(II)/Co(II) also exhibit binding constants in the order of ∼1 × 10(6) as in the case of ZnSO4. Positive role of the urea unit in the selectivity is confirmed by studying a model ligand L2, which is devoid of anion recognition urea unit. Structural characterization of four MSO4 [M = Zn(II), Cd(II), Co(II), Mn(II)] complexes of L1, that is, complex 1, [(L1)2(Zn)2(μ-SO4)2]; complex 2, [(L1)2(H2O)2(Cd)2(μ-SO4)2]; complex 3, [(L1)2(H2O)2(Co)2(μ-SO4)2]; and complex 4, [(L1)2(H2O)2(Mn)2(μ-SO4)2], reveal the formation of sulfate-bridged eight-membered crownlike binuclear complexes, similar to one of the concentration-dependent dimeric forms of MSO4 as observed in solid state

  6. Crystallization and preliminary X-ray diffraction studies of the carbohydrate-recognition domain of SIGN-R1, a receptor for microbial polysaccharides and sialylated antibody on splenic marginal zone macrophages

    PubMed Central

    Silva-Martin, Noella; Schauer, Joseph D.; Park, Chae Gyu; Hermoso, Juan A.

    2009-01-01

    SIGN-R1, or CD209b, is a mouse C-type lectin receptor that is expressed at high levels on macrophages in lymphoid tissues, especially within the marginal zone of the spleen. SIGN-R1 can bind and mediate the uptake of various microbial polysaccharides, including dextrans, lipopolysaccharides and pneumococcal capsular polysaccharides. It has been shown that SIGN-R1 mediates the clearance of encapsulated pneumococcus, complement fixation via binding C1q independent of antibody and innate resistance to pneumococcal infection. Recently, SIGN-R1 has also been demonstrated to bind sialylated antibody and mediate its activity to suppress autoimmunity. The carbohydrate-recognition domain (CRD) of SIGN-R1 has been cloned and overexpressed in a soluble secretory form in mammalian Chinese hamster ovary (CHO) cells. The CRD protein of SIGN-R1 was purified from CHO cell-culture supernatant and concentrated for crystallization using the hanging-drop vapour-diffusion method at 291 K. Crystals grew from a mixture of 2 M ammonium sulfate in 0.1 M bis-tris pH 5.5. Single crystals, which belonged to the monoclinic space group C2 with unit-cell parameters a = 146.72, b = 92.77, c = 77.06 Å, β = 121.66°, allowed the collection of a full X-ray data set to a maximum resolution of 1.87 Å. PMID:20054124

  7. The Potential Repertoire of the Innate Immune System in the Bladder: Expression of Pattern Recognition Receptors in the Rat Bladder and a Rat Urothelial Cell Line (MYP3 cells)

    PubMed Central

    Hughes, Francis M.; Turner, David P.; Purves, J. Todd

    2015-01-01

    Purpose The urothelium is a frontline sensor of the lower urinary tract, sampling the bladder lumen and stimulating an immune response to infectious and noxious agents. Pattern recognition receptors (PRRs) recognize such agents and coordinate the innate response, often by forming inflammasomes that activate caspase-1 and the release of Interleukin-1β. We have shown the presence of one PRR (NLRP3) in the urothelia and its central role in the inflammatory response to cyclophosphamide. The purpose of this study was to 1) assess the likely range of the PPR response by assessing the repertior present in the rat bladder and 2) determine the utility of the MYP3 rat urothelia cell line for in vitro studies by assessing it’s PPR repertior and functional responsiveness. Methods Immunohistochemistry was performed for seven PPRs (NLRP1, NLRP3, NLRP6, NLRP7, NLRP12, NLRC4 and AIM2) on bladder sections and MYP3 cells. For functionality, MYP3 cells were challanged with the quinessential NLRP3 activator ATP and assessed for caspase-1 activation. Results All PPRs examined were expressed in the bladder and localized to the urothelial layer with several also in the detrusor (none in the interstitia). MYP3 cells also expressed all PRRs with a variable intracellular location. ATP stimulated caspase-1 activity in MYP3 cells in a dose-dependent manner that was reduced by knockdown of NLRP3 expression. Conclusion The results suggest that the bladder possesses the capacity to initiate an innate immune response to a wide array of uropathological agents and the MYP3 cells will provide an excellent investigational tool for this field. PMID:26490556

  8. Fungal Surface and Innate Immune Recognition of Filamentous Fungi

    PubMed Central

    Figueiredo, Rodrigo T.; Carneiro, Leticia A. M.; Bozza, Marcelo T.

    2011-01-01

    The innate immune system performs specific detection of molecules from infectious agents through pattern recognition receptors. This recognition triggers inflammatory responses and activation of microbicidal mechanisms by leukocytes. Infections caused by filamentous fungi have increased in incidence and represent an important cause of mortality and morbidity especially in individuals with immunosuppression. This review will discuss the innate immune recognition of filamentous fungi molecules and its importance to infection control and disease. PMID:22194732

  9. Recognition and rejection of self in plant reproduction.

    PubMed

    Nasrallah, June B

    2002-04-12

    Plant self-incompatibility (SI) systems are unique among self/nonself recognition systems in being based on the recognition of self rather than nonself. SI in crucifer species is controlled by highly polymorphic and co-evolving genes linked in a complex. Self recognition is based on allele-specific interactions between stigma receptors and pollen ligands that result in the arrest of pollen tube development. Commonalities and differences between SI and other self/nonself discrimination systems are discussed.

  10. The recognition of biomaterials: pattern recognition of medical polymers and their adsorbed biomolecules.

    PubMed

    Love, Ryan J; Jones, Kim S

    2013-09-01

    All biomedical materials are recognized as foreign entities by the host immune system despite the substantial range of different materials that have been developed by material scientists and engineers. Hydrophobic biomaterials, hydrogels, biomaterials with low protein binding surfaces, and those that readily adsorb a protein layer all seem to incite similar host responses in vivo that may differ in magnitude, but ultimately result in encapsulation by fibrotic tissue. The recognition of medical materials by the host is explained by the very intricate pattern recognition system made up of integrins, toll-like receptors, scavenger receptors, and other surface proteins that enable leukocytes to perceive almost any foreign body. In this review, we describe the various pattern recognition receptors and processes that occur on biomedical material surfaces that permit detection of a range of materials within the host.

  11. Normal sorting but defective endocytosis of the low density lipoprotein receptor in mice with autosomal recessive hypercholesterolemia.

    PubMed

    Jones, Christopher; Hammer, Robert E; Li, Wei-Ping; Cohen, Jonathan C; Hobbs, Helen H; Herz, Joachim

    2003-08-01

    Autosomal recessive hypercholesterolemia (ARH) is a genetic form of hypercholesterolemia that clinically resembles familial hypercholesterolemia (FH). As in FH, the rate of clearance of circulating low density lipoprotein (LDL) by the LDL receptor (LDLR) in the liver is markedly reduced in ARH. Unlike FH, LDL uptake in cultured fibroblasts from ARH patients is normal or only slightly impaired. The gene defective in ARH encodes a putative adaptor protein that has been implicated in linking the LDLR to the endocytic machinery. To determine the role of ARH in the liver, ARH-deficient mice were developed. Plasma levels of LDL-cholesterol were elevated in the chow-fed Arh-/- mice (83 +/- 8 mg/dl versus 68 +/- 8 mg/dl) but were lower than those of mice expressing no LDLR (Ldlr-/-) (197 +/- 8 mg/dl). Cholesterol feeding elevated plasma cholesterol levels in both strains. The fractional clearance rate of radiolabeled LDL was reduced to similar levels in the Arh-/- and Ldlr-/- mice, whereas the rate of removal of alpha2-macroglobulin by the LDLR-related protein, which also interacts with ARH, was unchanged. Immunolocalization studies revealed that a much greater proportion of immunodetectable LDLR, but not LDLR-related protein, was present on the sinusoidal surface of hepatocytes in the Arh-/- mice. Taken together, these results are consistent with ARH playing a critical and specific role in LDLR endocytosis in the liver.

  12. Cloning of a cDNA encoding a putative human very low density lipoprotein/Apolipoprotein E receptor and assignment of the gene to chromosome 9pter-p23[sup 6

    SciTech Connect

    Gafvels, M.E.; Strauss, J.F. III ); Caird, M.; Patterson, D. ); Britt, D.; Jackson, C.L. )

    1993-11-01

    The authors report the cloning of a 3656-bp cDNA encoding a putative human very low density lipoprotein (VLDL)/apolipoprotein E (ApoE) receptor. The gene encoding this protein was mapped to chromosome 9pter-p23. Northern analysis of human RNA identified cognate mRNAs of 6.0 and 3.8 kb with most abundant expression in heart and skeletal muscle, followed by kidney, placenta, pancreas, and brain. The pattern of expression generally paralleled that of lipoprotein lipase mRNA but differed from that of the low density lipoprotein (LDL) receptor and the low density lipoprotein receptor-related protein/[alpha][sub 2]-macroglobulin receptor (LRP), which are members of the same gene family. VLDL/ApoE receptor message was not detected in liver, whereas mRNAs for both LDL receptor and LRP were found in hepatic tissue. In mouse 3T3-L1 cells, VLDL/ApoE receptor mRNA was induced during the transformation of the cells into adipocytes. Expression was also detected in human choriocarcinoma cells, suggesting that at least part of the expression observed in placenta may be in trophoblasts, cells which would be exposed to maternal blood. Expression in brain may be related to high levels of ApoE expression in that organ, an observation of potential relevance to the recently hypothesized role for ApoE in late onset Alzheimer disease. The results suggest that the putative VLDL/ApoE receptor could play a role in the uptake of triglyceride-rich lipoprotein particles by specific organs including striated and cardiac muscle and adipose tissue and in the transport of maternal lipids across the placenta. The findings presented here, together with recent observations from other laboratories, bring up the possibility that a single gene, the VLDL/ApoE receptor, may play a role in the pathogenesis of certain forms of atherosclerosis, Alzheimer disease, and obesity.

  13. Innate immune recognition of microbial cell wall components and microbial strategies to evade such recognitions.

    PubMed

    Sukhithasri, V; Nisha, N; Biswas, Lalitha; Anil Kumar, V; Biswas, Raja

    2013-08-25

    The innate immune system constitutes the first line of defence against invading microbes. The basis of this defence resides in the recognition of defined structural motifs of the microbes called "Microbial associated molecular patterns" that are absent in the host. Cell wall, the outer layer of both bacterial and fungal cells, a unique structure that is absent in the host and is recognized by the germ line encoded host receptors. Nucleotide oligomerization domain proteins, peptidoglycan recognition proteins and C-type lectins are host receptors that are involved in the recognition of bacterial cell wall (usually called peptidoglycan), whereas fungal cell wall components (N- and O-linked mannans, β-glucans etc.) are recognized by host receptors like C-type lectins (Dectin-1, Dectin-2, mannose receptor, DC-SIGN), Toll like receptors-2 and -4 (TLR-2 and TLR-4). These recognitions lead to activation of a variety of host signaling cascades and ultimate production of anti-microbial compounds including phospholipase A2, antimicrobial peptides, lysozyme, reactive oxygen and nitrogen species. These molecules act in cohort against the invading microbes to eradicate infections. Additionally pathogen recognition leads to the production of cytokines, which further activate the adaptive immune system. Both pathogenic and commensal bacteria and fungus use numerous strategies to subvert the host defence. These strategies include bacterial peptidoglycan glycan backbone modifications by O-acetylation, N-deacetylation, N-glycolylation and stem peptide modifications by amidation of meso-Diaminopimelic acid; fungal cell wall modifications by shielding the β-glucan layer with mannoproteins and α-1,3 glucan. This review focuses on the recent advances in understanding the role of bacterial and fungal cell wall in their innate immune recognition and evasion strategies.

  14. Low-density Lipoprotein Receptor-related Proteins in a Novel Mechanism of Axon Guidance and Peripheral Nerve Regeneration.

    PubMed

    Landowski, Lila M; Pavez, Macarena; Brown, Lachlan S; Gasperini, Robert; Taylor, Bruce V; West, Adrian K; Foa, Lisa

    2016-01-15

    The low-density lipoprotein receptor-related protein receptors 1 and 2 (LRP1 and LRP2) are emerging as important cell signaling mediators in modulating neuronal growth and repair. We examined whether LRP1 and LRP2 are able to mediate a specific aspect of neuronal growth: axon guidance. We sought to identify LRP1 and LRP2 ligands that could induce axonal chemoattraction, which might have therapeutic potential. Using embryonic sensory neurons (rat dorsal root ganglia) in a growth cone turning assay, we tested a range of LRP1 and LRP2 ligands for the ability to guide growth cone navigation. Three ligands were chemorepulsive: α-2-macroglobulin, tissue plasminogen activator, and metallothionein III. Conversely, only one LRP ligand, metallothionein II, was found to be chemoattractive. Chemoattraction toward a gradient of metallothionein II was calcium-dependent, required the expression of both LRP1 and LRP2, and likely involves further co-receptors such as the tropomyosin-related kinase A (TrkA) receptor. The potential for LRP-mediated chemoattraction to mediate axonal regeneration was examined in vivo in a model of chemical denervation in adult rats. In these in vivo studies, metallothionein II was shown to enhance epidermal nerve fiber regeneration so that it was complete within 7 days compared with 14 days in saline-treated animals. Our data demonstrate that both LRP1 and LRP2 are necessary for metallothionein II-mediated chemotactic signal transduction and that they may form part of a signaling complex. Furthermore, the data suggest that LRP-mediated chemoattraction represents a novel, non-classical signaling system that has therapeutic potential as a disease-modifying agent for the injured peripheral nervous system.

  15. Viral cell recognition and entry.

    PubMed Central

    Rossmann, M. G.

    1994-01-01

    Rhinovirus infection is initiated by the recognition of a specific cell-surface receptor. The major group of rhinovirus serotypes attach to intercellular adhesion molecule-1 (ICAM-1). The attachment process initiates a series of conformational changes resulting in the loss of genomic RNA from the virion. X-ray crystallography and sequence comparisons suggested that a deep crevice or canyon is the site on the virus recognized by the cellular receptor molecule. This has now been verified by electron microscopy of human rhinovirus 14 (HRV14) and HRV16 complexed with a soluble component of ICAM-1. A hydrophobic pocket underneath the canyon is the site of binding of various hydrophobic drug compounds that can inhibit attachment and uncoating. This pocket is also associated with an unidentified, possibly cellular in origin, "pocket factor." The pocket factor binding site overlaps the binding site of the receptor. It is suggested that competition between the pocket factor and receptor regulates the conformational changes required for the initiation of the entry of the genomic RNA into the cell. PMID:7849588

  16. Image Recognition Based on Biometric Pattern Recognition

    NASA Astrophysics Data System (ADS)

    Sun, Shuliang; Chen, Zhong; Liu, Chenglian; Guo, Yongning; Lin, Xueyun

    2011-09-01

    A new method, biomimetric pattern recognition, is mentioned to recognize images. At first, the image is pretreatment and feature extraction, then a high vector is got. A biomimetric pattern recognition model is designed. The judgment function is used to discriminate the classification of the samples. It is showed that the method is effective for little samples by experiment. It would be useful in many fields in future.

  17. Optical Pattern Recognition

    NASA Astrophysics Data System (ADS)

    Yu, Francis T. S.; Jutamulia, Suganda

    2008-10-01

    Contributors; Preface; 1. Pattern recognition with optics Francis T. S. Yu and Don A. Gregory; 2. Hybrid neural networks for nonlinear pattern recognition Taiwei Lu; 3. Wavelets, optics, and pattern recognition Yao Li and Yunglong Sheng; 4. Applications of the fractional Fourier transform to optical pattern recognition David Mendlovic, Zeev Zalesky and Haldum M. Oxaktas; 5. Optical implementation of mathematical morphology Tien-Hsin Chao; 6. Nonlinear optical correlators with improved discrimination capability for object location and recognition Leonid P. Yaroslavsky; 7. Distortion-invariant quadratic filters Gregory Gheen; 8. Composite filter synthesis as applied to pattern recognition Shizhou Yin and Guowen Lu; 9. Iterative procedures in electro-optical pattern recognition Joseph Shamir; 10. Optoelectronic hybrid system for three-dimensional object pattern recognition Guoguang Mu, Mingzhe Lu and Ying Sun; 11. Applications of photrefractive devices in optical pattern recognition Ziangyang Yang; 12. Optical pattern recognition with microlasers Eung-Gi Paek; 13. Optical properties and applications of bacteriorhodopsin Q. Wang Song and Yu-He Zhang; 14. Liquid-crystal spatial light modulators Aris Tanone and Suganda Jutamulia; 15. Representations of fully complex functions on real-time spatial light modulators Robert W. Cohn and Laurence G. Hassbrook; Index.

  18. Multimodal eye recognition

    NASA Astrophysics Data System (ADS)

    Zhou, Zhi; Du, Yingzi; Thomas, N. L.; Delp, Edward J., III

    2010-04-01

    Multimodal biometrics use more than one means of biometric identification to achieve higher recognition accuracy, since sometimes a unimodal biometric is not good enough used to do identification and classification. In this paper, we proposed a multimodal eye recognition system, which can obtain both iris and sclera patterns from one color eye image. Gabor filter and 1-D Log-Gabor filter algorithms have been applied as the iris recognition algorithms. In sclera recognition, we introduced automatic sclera segmentation, sclera pattern enhancement, sclera pattern template generation, and sclera pattern matching. We applied kernelbased matching score fusion to improve the performance of the eye recognition system. The experimental results show that the proposed eye recognition method can achieve better performance compared to unimodal biometric identification, and the accuracy of our proposed kernel-based matching score fusion method is higher than two classic linear matching score fusion methods: Principal Component Analysis (PCA) and Linear Discriminant Analysis (LDA).

  19. Low-density lipoprotein receptor-related protein-1 (LRP-1) expression in a rat model of oxygen-induced retinal neovascularization.

    PubMed

    Sánchez, María C; Barcelona, Pablo F; Luna, Jose D; Ortiz, Susana G; Juarez, Patricio C; Riera, Clelia M; Chiabrando, Gustavo A

    2006-12-01

    The low-density lipoprotein receptor-related protein-1 (LRP-1) is a high-molecular weight receptor of the LDL receptor gene family. Its ability to bind and internalize both proteinases and proteinase-inhibitor complexes from the extracellular space suggests that it has a major role in modulating uncontrolled retinal cell proliferation. In order to test this assumption, we investigated the expression of LRP-1 and receptor-associated ligands in a rat model of oxygen-induced retinal neovascularization. Wistar albino rats were placed into incubators at birth and exposed to an atmosphere alternating between 50% and 10% of oxygen every 24 h. After 14 days, the animals were allowed to recover in room air and sacrificed at postnatal day 20 (P20). The protein expression of LRP-1 and alpha2-macroglobulin (alpha2M) in the retina from unexposed and hyperoxia-exposed rats was investigated by Western blot. The localization of LRP-1 after neovascularization was assessed by immunohistochemical staining. The activity of metalloproteinases (MMPs) was determined by zymography. Histological analysis was done to quantitate the neovascular response in these animals. Western blot analysis showed that LRP-1 was expressed, along with alpha2M, in the retina of rats with oxygen-induced neovascularization at P20. By immunohistochemical analysis, positive staining for LRP-1 appeared in cells extending from the inner limiting membrane (ILM) to the outer limiting membrane (OLM). The cells of the retina that expressed LRP-1 were identified by immunofluorescence as Müller cells. Zymographic analysis demonstrated increased activity of MMP-2 and MMP-9 under neovascular conditions. This is the first demonstration of the involvement of LRP-1 in retinal neovascularization. In retinas of rats with oxygen-induced neovascularization, the expression of LRP-1 and alpha2M was increased along with an enhanced activity of MMPs, suggesting that LRP-1 expression may play a role in modulating retinal

  20. Moreland Recognition Program.

    ERIC Educational Resources Information Center

    Moreland Elementary School District, San Jose, CA.

    THE FOLLOWING IS THE FULL TEXT OF THIS DOCUMENT: Recognition for special effort and achievement has been noted as a component of effective schools. Schools in the Moreland School District have effectively improved standards of discipline and achievement by providing forty-six different ways for children to receive positive recognition. Good…

  1. Biological chiral recognition: the substrate's perspective.

    PubMed

    Sundaresan, Vidyasankar; Abrol, Ravinder

    2005-01-01

    A novel stereocenter-recognition (SR) model has been proposed recently for describing the stereoselectivity of biological and other macromolecules towards substrates that have multiple stereocenters, based on the topology of substrate stereocenters (Sundaresan and Abrol, Prot Sci 11:1330-1339, 2002). The SR model provides the minimum number of substrate locations interacting with receptor sites that need to be considered for understanding stereoselectivity characteristics. Interactions between substrate locations and receptor sites may be binding, nonbinding or repulsive in nature and may occur in a many-to-one or one-to-many fashion, but for a receptor to be stereoselective, its interactions with substrate stereoisomers have to involve a minimum number of locations, in the correct geometry. The SR model is topologically rigorous, explains several previous experimental observations, and is predictive in nature. It predicts that stereoselectivity towards a substrate with N stereocenters in a linear structure involves a minimum of N + 2 substrate locations, distributed over all stereocenters in the substrate, such that effectively at least three locations per stereocenter interact with one or more receptor sites. This article uses the SR model to provide an insight into the chiral recognition process from a substrate's perspective that is intuitive and simple, furnishing a rigorous stereochemical basis for explaining stereoselectivity characteristics of many biological systems.

  2. Recognition of a signal peptide by the signal recognition particle

    PubMed Central

    Janda, Claudia Y.; Li, Jade; Oubridge, Chris; Hernández, Helena; Robinson, Carol V.; Nagai, Kiyoshi

    2010-01-01

    Targeting of proteins to appropriate sub-cellular compartments is a crucial process in all living cells. Secretory and membrane proteins usually contain an N-terminal signal peptide, which is recognised by the signal recognition particle (SRP) when nascent polypeptide chains emerge from the ribosome. The SRP-ribosome nascent chain complex is then targeted through its GTP-dependent interaction with SRP-receptor to the protein-conducting channel on endoplasmic reticulum membrane in eukaryotes or plasma membrane in bacteria. A universally conserved component of SRP1, 2, SRP54 or its bacterial homolog, fifty-four homolog (Ffh), binds the signal peptides which have a highly divergent sequence divisible into a positively charged n-region, an h-region commonly containing 8-20 hydrophobic residues and a polar c-region 3-5. No structure has been reported that exemplified SRP54 binding of any signal sequence. We have produced a fusion protein between Sulfolobus solfataricus SRP54 and a signal peptide connected via a flexible linker. This fusion protein oligomerises in solution, through interaction between the SRP54 and signal peptide moieties belonging to different chains, and it is functional, able to bind SRP RNA and SRP-receptor FtsY. Here we present the crystal structure at 3.5 Å resolution of an SRP54-signal peptide complex in the dimer, which reveals how a signal sequence is recognised by SRP54. PMID:20364120

  3. Biomimetic Receptors and Sensors

    PubMed Central

    Dickert, Franz L.

    2014-01-01

    In biomimetics, living systems are imitated to develop receptors for ions, molecules and bioparticles. The most pertinent idea is self-organization in analogy to evolution in nature, which created the key-lock principle. Today, modern science has been developing host-guest chemistry, a strategy of supramolecular chemistry for designing interactions of analytes with synthetic receptors. This can be realized, e.g., by self-assembled monolayers (SAMs) or molecular imprinting. The strategies are used for solid phase extraction (SPE), but preferably in developing recognition layers of chemical sensors. PMID:25436653

  4. Biomimetic receptors and sensors.

    PubMed

    Dickert, Franz L

    2014-11-27

    In biomimetics, living systems are imitated to develop receptors for ions, molecules and bioparticles. The most pertinent idea is self-organization in analogy to evolution in nature, which created the key-lock principle. Today, modern science has been developing host-guest chemistry, a strategy of supramolecular chemistry for designing interactions of analytes with synthetic receptors. This can be realized, e.g., by self-assembled monolayers (SAMs) or molecular imprinting. The strategies are used for solid phase extraction (SPE), but preferably in developing recognition layers of chemical sensors.

  5. Pattern recognition technique

    NASA Technical Reports Server (NTRS)

    Hong, J. P.

    1971-01-01

    Technique operates regardless of pattern rotation, translation or magnification and successfully detects out-of-register patterns. It improves accuracy and reduces cost of various optical character recognition devices and page readers and provides data input to computer.

  6. Macrophage recognition of ICAM-3 on apoptotic leukocytes.

    PubMed

    Moffatt, O D; Devitt, A; Bell, E D; Simmons, D L; Gregory, C D

    1999-06-01

    Cells undergoing apoptosis are cleared rapidly by phagocytes, thus preventing tissue damage caused by loss of plasma membrane integrity. In this study, we show that the surface of leukocytes is altered during apoptosis such that the first Ig-like domain of ICAM-3 (CD50) can participate in the recognition and phagocytosis of the apoptotic cells by macrophages. Macrophage recognition of apoptotic cell-associated ICAM-3 was demonstrated both on leukocytes and, following transfection of exogenous ICAM-3, on nonleukocytes. The change in ICAM-3 was a consistent consequence of apoptosis triggered by various stimuli, suggesting that it occurs as part of a final common pathway of apoptosis. Alteration of ICAM-3 on apoptotic cells permitting recognition by macrophages resulted in a switch in ICAM-3-binding preference from the prototypic ICAM-3 counterreceptor, LFA-1, to an alternative macrophage receptor. Using mAbs to block macrophage/apoptotic cell interactions, we were unable to obtain evidence that either the alternative ICAM-3 counterreceptor alpha d beta 2 or the apoptotic cell receptor alpha v beta 3 was involved in the recognition of ICAM-3. By contrast, mAb blockade of macrophage CD14 inhibited ICAM-3-dependent recognition of apoptotic cells. These results show that ICAM-3 can function as a phagocytic marker of apoptotic leukocytes on which it acquires altered macrophage receptor-binding activity.

  7. Molecular imprinting of proteins emerging as a tool for protein recognition.

    PubMed

    Takeuchi, Toshifumi; Hishiya, Takayuki

    2008-07-21

    This article gives the recent developments in molecular imprinting for proteins. Currently bio-macromolecules such as antibodies and enzymes are mainly employed for protein recognition purposes. However, such bio-macromolecules are sometimes difficult to find and/or produce, therefore, receptor-like synthetic materials such as protein-imprinted polymers have been intensively studied as substitutes for natural receptors. Recent advances in protein imprinting shown here demonstrate the possibility of this technique as a future technology of protein recognition.

  8. Human transforming growth factor. beta. -. cap alpha. /sub 2/-macroglobulin complex is a latent form of transforming growth factor. beta

    SciTech Connect

    Huang, S.S.; O'Grady, P.; Huang, J.S.

    1987-05-01

    Human platelet-derived transforming growth factor ..beta.. (TGF..beta..) has been shown to be present as a high molecular weight latent form in human serum. Appearance of transforming growth factor activity, along with the change from high molecular weight form to low molecular weight form, was observed following treatment of the latent form of TGF..beta.. with acid or urea, suggesting that the latent form of TGF..beta.. is a complex of TGF..beta.. and a high molecular weight binding protein. Human ..cap alpha../sub 2/-M has been found to be a plasma binding protein for platelet-derived growth factor (PDGF) in serum or plasma. TGF..beta.. and PDGF share similar properties. They, therefore, investigated the interaction between /sup 125/I-TGF..beta.. and ..cap alpha../sub 2/M. /sup 125/I-TGF..beta.. and purified human ..cap alpha../sub 2/M formed a complex as demonstrated by polyacrylamide gel electrophoresis. Most of the /sup 125/I-TGF..beta..-..cap alpha../sub 2/M complex could be dissociated by acid or urea treatment. These results suggest that ..cap alpha../sub 2/M is a binding protein for TGF..beta.. and that TGF..beta..-..cap alpha../sub 2/M complex may be the latent form of TGF..beta.. in serum.

  9. Evolutionary grass roots for odor recognition.

    PubMed

    Olender, Tsviya; Lancet, Doron

    2012-09-01

    Considerable evidence supports the idea that odorant recognition depends on specific sequence variations in olfactory receptor (OR) proteins. Much of this emerges from in vitro screens in heterogenous expression systems. However, the ultimate proof should arise from measurements of odorant thresholds in human individuals harboring different OR genetic variants, a research vein that has so far been only scantly explored. The study of McRae et al., published in this issue of Chemical Senses, shows how the recognition of a grassy odorant depends on specific OR interindividual sequence changes. It provides a clear relevant example for the impact of genetics on olfaction and is an excellent portrayal of the power of human genomics to decipher olfactory perception.

  10. Quantitative isoform-profiling of highly diversified recognition molecules

    PubMed Central

    Schreiner, Dietmar; Simicevic, Jovan; Ahrné, Erik; Schmidt, Alexander; Scheiffele, Peter

    2015-01-01

    Complex biological systems rely on cell surface cues that govern cellular self-recognition and selective interactions with appropriate partners. Molecular diversification of cell surface recognition molecules through DNA recombination and complex alternative splicing has emerged as an important principle for encoding such interactions. However, the lack of tools to specifically detect and quantify receptor protein isoforms is a major impediment to functional studies. We here developed a workflow for targeted mass spectrometry by selected reaction monitoring that permits quantitative assessment of highly diversified protein families. We apply this workflow to dissecting the molecular diversity of the neuronal neurexin receptors and uncover an alternative splicing-dependent recognition code for synaptic ligands. DOI: http://dx.doi.org/10.7554/eLife.07794.001 PMID:25985086

  11. Estrogenic involvement in social learning, social recognition and pathogen avoidance.

    PubMed

    Choleris, Elena; Clipperton-Allen, Amy E; Phan, Anna; Valsecchi, Paola; Kavaliers, Martin

    2012-04-01

    Sociality comes with specific cognitive skills that allow the proper processing of information about others (social recognition), as well as of information originating from others (social learning). Because sociality and social interactions can also facilitate the spread of infection among individuals the ability to recognize and avoid pathogen threat is also essential. We review here various studies primarily from the rodent literature supporting estrogenic involvement in the regulation of social recognition, social learning (socially acquired food preferences and mate choice copying) and the recognition and avoidance of infected and potentially infected individuals. We consider both genomic and rapid estrogenic effects involving estrogen receptors α and β, and G-protein coupled estrogen receptor 1, along with their interactions with neuropeptide systems in the processing of social stimuli and the regulation and expression of these various socially relevant behaviors.

  12. Small Peptide Recognition Sequence for Intracellular Sorting

    PubMed Central

    Pandey, Kailash N.

    2010-01-01

    Increasing evidence indicate that complex arrays of short signals and recognition peptide sequence ensure accurate trafficking and distribution of transmembrane receptors and/or proteins and their ligands into intracellular compartments. Internalization and subsequent trafficking of cell-surface receptors into the cell interior is mediated by specific short-sequence peptide signals within the cytoplasmic domains of these receptor proteins. The short signals usually consist of small linear amino acid sequences, which are recognized by adaptor coat proteins along the endocytic and sorting pathways. In recent years, much has been learned about the function and mechanisms of endocytic pathways responsible for the trafficking and molecular sorting of membrane receptors and their ligands into intracellular compartments, however, the significance and scope of the short sequence motifs in these cellular events is not well understood. Here a particular emphasis has been given to the functions of short-sequence signal motifs responsible for the itinerary and destination of membrane receptors and proteins moving into subcellular compartments. PMID:20817434

  13. [Prosopagnosia and facial expression recognition].

    PubMed

    Koyama, Shinichi

    2014-04-01

    This paper reviews clinical neuropsychological studies that have indicated that the recognition of a person's identity and the recognition of facial expressions are processed by different cortical and subcortical areas of the brain. The fusiform gyrus, especially the right fusiform gyrus, plays an important role in the recognition of identity. The superior temporal sulcus, amygdala, and medial frontal cortex play important roles in facial-expression recognition. Both facial recognition and facial-expression recognition are highly intellectual processes that involve several regions of the brain.

  14. Nicotine Administration Attenuates Methamphetamine-Induced Novel Object Recognition Deficits

    PubMed Central

    Vieira-Brock, Paula L.; McFadden, Lisa M.; Nielsen, Shannon M.; Smith, Misty D.; Hanson, Glen R.

    2015-01-01

    Background: Previous studies have demonstrated that methamphetamine abuse leads to memory deficits and these are associated with relapse. Furthermore, extensive evidence indicates that nicotine prevents and/or improves memory deficits in different models of cognitive dysfunction and these nicotinic effects might be mediated by hippocampal or cortical nicotinic acetylcholine receptors. The present study investigated whether nicotine attenuates methamphetamine-induced novel object recognition deficits in rats and explored potential underlying mechanisms. Methods: Adolescent or adult male Sprague-Dawley rats received either nicotine water (10–75 μg/mL) or tap water for several weeks. Methamphetamine (4×7.5mg/kg/injection) or saline was administered either before or after chronic nicotine exposure. Novel object recognition was evaluated 6 days after methamphetamine or saline. Serotonin transporter function and density and α4β2 nicotinic acetylcholine receptor density were assessed on the following day. Results: Chronic nicotine intake via drinking water beginning during either adolescence or adulthood attenuated the novel object recognition deficits caused by a high-dose methamphetamine administration. Similarly, nicotine attenuated methamphetamine-induced deficits in novel object recognition when administered after methamphetamine treatment. However, nicotine did not attenuate the serotonergic deficits caused by methamphetamine in adults. Conversely, nicotine attenuated methamphetamine-induced deficits in α4β2 nicotinic acetylcholine receptor density in the hippocampal CA1 region. Furthermore, nicotine increased α4β2 nicotinic acetylcholine receptor density in the hippocampal CA3, dentate gyrus and perirhinal cortex in both saline- and methamphetamine-treated rats. Conclusions: Overall, these findings suggest that nicotine-induced increases in α4β2 nicotinic acetylcholine receptors in the hippocampus and perirhinal cortex might be one mechanism by which

  15. Recognition of Streptococcus pneumoniae by the innate immune system.

    PubMed

    Koppe, Uwe; Suttorp, Norbert; Opitz, Bastian

    2012-04-01

    Streptococcus pneumoniae is both a frequent colonizer of the upper respiratory tract and a leading cause of life-threatening infections such as pneumonia, meningitis and sepsis. The innate immune system is critical for the control of colonization and for defence during invasive disease. Initially, pneumococci are recognized by different sensors of the innate immune system called pattern recognition receptors (PRRs), which control most subsequent host defence pathways. These PRRs include the transmembrane Toll-like receptors (TLRs) as well as the cytosolic NOD-like receptors (NLRs) and DNA sensors. Recognition of S. pneumoniae by members of these PRR families regulates the production of inflammatory mediators that orchestrate the following immune response of infected as well as neighbouring non-infected cells, stimulates the recruitment of immune cells such as neutrophils and macrophages, and shapes the adaptive immunity. This review summarizes the current knowledge of the function of different PRRs in S. pneumoniae infection.

  16. Automatic object recognition

    NASA Technical Reports Server (NTRS)

    Ranganath, H. S.; Mcingvale, Pat; Sage, Heinz

    1988-01-01

    Geometric and intensity features are very useful in object recognition. An intensity feature is a measure of contrast between object pixels and background pixels. Geometric features provide shape and size information. A model based approach is presented for computing geometric features. Knowledge about objects and imaging system is used to estimate orientation of objects with respect to the line of sight.

  17. Units of Word Recognition.

    ERIC Educational Resources Information Center

    Santa, Carol M.; And Others

    Both psychologists and reading specialists have been interested in whether words are processed letter by letter or in larger units. A reaction time paradigm was used to evaluate these options with interest focused on potential units of word recognition which might be functional within single syllable words. The basic paradigm involved presenting…

  18. Optical Character Recognition.

    ERIC Educational Resources Information Center

    Converso, L.; Hocek, S.

    1990-01-01

    This paper describes computer-based optical character recognition (OCR) systems, focusing on their components (the computer, the scanner, the OCR, and the output device); how the systems work; and features to consider in selecting a system. A list of 26 questions to ask to evaluate systems for potential purchase is included. (JDD)

  19. Automat