Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: UNIPROT:P43026 (lipopolysaccharide)
62,215 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The receptor complex resulting from association of MD-2 and the ectodomain of Toll-like receptor 4 (TLR4) mediates lipopolysaccharide (LPS) signal transduction across the cell membrane. We prepared a tertiary structure model of MD-2, based on the known structures of homologous lipid-binding proteins. Analysis of circular dichroic spectra of purified bacterially expressed MD-2 indicates high content of beta-type secondary structure, in agreement with the structural model. Bacterially expressed MD-2 was able to confer LPS responsiveness to cells expressing TLR4 despite lacking glycosylation. We identified several clusters of basic residues on the surface of MD-2. Mutation of each of two clusters encompassing the residues Lys(89)-Arg(90)-Lys(91) and Lys(125)-Lys(125) significantly decreased the signal transduction of the respective MD-2 mutants either upon co-expression with TLR4 or upon addition as soluble protein into the supernatant of cells overexpressing TLR4. These basic clusters lie at the edge of the beta-sheet sandwich, which in cholesterol-binding protein connected to Niemann-Pick disease C2 (NPC2), dust mite allergen Der p2, and ganglioside GM2-activator protein form a hydrophobic pocket. In contrast, mutation of another basic cluster composed of Arg(69)-Lys(72), which according to the model lies further apart from the hydrophobic pocket only weakly decreased MD-2 activity. Furthermore, addition of the peptide, comprising the surface loop between Cys(95) and Cys(105), predicted by model, particularly in oxidized form, decreased LPS-induced production of tumor necrosis factor alpha and interleukin-8 upon application to monocytic cells and fibroblasts, respectively, supporting its involvement in LPS signaling. Our structural model of MD-2 is corroborated by biochemical analysis and contributes to the unraveling of molecular interactions in LPS recognition.
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PMID:Structural model of MD-2 and functional role of its basic amino acid clusters involved in cellular lipopolysaccharide recognition. 1511 23

The phenomenon of endotoxin tolerance has been widely investigated, but to date, the molecular mechanisms of endotoxin tolerance remain to be resolved clearly. The discovery of the Toll-like receptor (TLR) family as the major receptors for lipopolysaccharide (LPS) and other bacterial products has prompted a resurgence of interest in endotoxin tolerance mechanisms. Changes of cell surface molecules, signaling proteins, pro-inflammatory and anti-inflammatory cytokines and other mediators have been examined. During tolerance expression of LPS-binding protein (LBP), CD14, myeloid differentiation protein-2 (MD-2) and TLR2 are unchanged or up-regulated, whereas TLR4 is transiently suppressed or unchanged. Proximal post-receptor signaling proteins that are altered in tolerance include augmented degradation of interleukin-1 receptor-associated kinase (IRAK), and decreased TLR4-myeloid differentiation factor 88 (MyD88) and IRAK-MyD88 association. Tolerance has also been shown to be associated with decreased Gi protein content and activity, decreased protein kinase C (PKC) activity, reduction in mitogen-activated protein kinase (MAP kinase) activity, and reduced activator protein-1 (AP-1) and nuclear factor kappa B (NF-kappaB) induced gene transactivation. However, not all signaling proteins and pathways are suppressed in tolerance and induction of specific anti-inflammatory proteins and signaling pathways may serve important counter inflammatory functions. The latter include induction of IRAK-M and suppressor of cytokine-signaling-1 (SOCS-1), phosphoinositide-3-kinase (PI3K) signaling, and increased or maintained expression of inhibitor-kappaB (IkappaB) isoforms. Also at the nuclear level, increase in the NF-kappaB subunit p50 homodimer expression and increased activation of peroxisome-proliferator-activated receptors-gamma (PPARgamma) have been linked to tolerance phenotype. Although there are species and cellular variations in manifestation of the LPS tolerant phenotype, it is clear that the tolerance phenomena have evolved as a complex orchestrated counter regulatory response to inflammation.
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PMID:Molecular mechanisms of endotoxin tolerance. 1511 98

Endothelial cells are activated by microbial agonists through Toll-like receptors (TLRs) to express inflammatory mediators; this is of significance in acute as well as chronic inflammatory states such as septic shock and atherosclerosis, respectively. We investigated mechanisms of lipopolysaccharide (LPS)-induced cell activation in human coronary artery endothelial cells (HCAEC) using a combination of FACS, confocal microscopy, RT-PCR, and functional assays. We found that TLR4, in contrast to TLR2, is not only located intracellularly but also functions intracellularly. That being the case, internalization of LPS is required for activation. We further characterized the HCAEC LPS uptake system and found that HCAEC exhibit an effective LPS uptake only in the presence of LPS binding protein (LBP). In addition to its function as a catalyst for LPS-CD14 complex formation, LBP enables HCAEC activation at low LPS concentrations by facilitating the uptake, and therefore delivery, of LPS-CD14 complexes to intracellular TLR4-MD-2. LBP-dependent uptake involves a scavenger receptor pathway. Our findings may be of pathophysiological relevance in the initial response of the organism to infection. Results further suggest that LBP levels, which vary as LBP is an acute phase reactant, could be relevant to initiating inflammatory responses in the vasculature in response to chronic or recurring low LPS.
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PMID:Toll-like receptor 4 functions intracellularly in human coronary artery endothelial cells: roles of LBP and sCD14 in mediating LPS responses. 1513 88

The zinc finger protein A20 is encoded by an immediate early response gene and acts as an inhibitor of nuclear factor (NF)-kappaB-dependent gene expression induced by different stimuli, including tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta). Toll-like receptor 2 (TLR2) and TLR4 have been found to transduce, respectively, peptidoglycan (PGN) and lipopolysaccharide (LPS) signals for the activation of NF-kappaB and the production of inflammatory cytokines. Here, we have examined the role of A20 in TLR-mediated NF-kappaB-dependent gene expression in human airway epithelial cells (AECs). Stimulation with LPS and PGN resulted in a significant increase in the level of A20 mRNA in primary cultured AECs and in NCI-H292 AECs. LPS and PGN induced activation of the IL-8 promoter both in NCI-H292 AECs and in HEK293 cells expressing either TLR2 or TLR4 plus MD-2. Dominant-negative myeloid differentiation protein and a mutant form of IkappaBalpha attenuated this PGN- or LPS-induced activation of the IL-8 promoter. Furthermore, overexpression of A20 inhibited activation of both NF-kappaB and the IL-8 promoter by PGN or LPS in these cells. Taken together, our results suggest that A20 may function as a negative regulator of TLR-mediated inflammatory responses in the airway, thereby protecting the host against harmful overresponses to pathogens.
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PMID:A20 inhibits toll-like receptor 2- and 4-mediated interleukin-8 synthesis in airway epithelial cells. 1514 65

We have previously shown that non-pathogenic Gram negative bacteria induce RelA phosphorylation, nuclear factor (NF)-kappaB transcriptional activity and pro-inflammatory gene expression in intestinal epithelial cells (IEC) in vivo and in vitro. In this study, we investigated the molecular mechanism of immune-epithelial cell cross-talk on Gram-negative enteric bacteria-induced NF-kappaB signalling and pro-inflammatory gene expression in IEC using HT-29/MTX as well as CaCO-2 transwell cultures Interestingly, while differentiated HT-29/MTX cells are unresponsive to non-pathogenic Gram negative bacterial stimulation, interleukin-8 (IL-8) mRNA accumulation is strongly induced in Escherichia coli- but not Bacteroides vulgatus-stimulated IEC cocultured with peripheral blood (PBMC) and lamina propria mononuclear cells (LPMC). The presence of PBMC triggered both E. coli- and B. vulgatus-induced mRNA expression of the Toll-like receptor-4 accessory protein MD-2 as well as endogenous IkappaBalpha phosphorylation, demonstrating similar capabilities of these bacteria to induce proximal NF-kappaB signalling. However, B. vulgatus failed to trigger IkappaBalpha degradation and NF-kappaB transcriptional activity in the presence of PBMC. Interestingly, B. vulgatus- and E. coli-derived lipopolysaccharide-induced similar IL-8 mRNA expression in epithelial cells after basolateral stimulation of HT-29/PBMC cocultures. Although luminal enteric bacteria have adjuvant and antigenic properties in chronic intestinal inflammation, PBMC from patients with active ulcerative colitis and Crohn's disease differentially trigger epithelial cell activation in response to E. coli and E. coli-derived LPS. In conclusion, this study provides evidence for a differential regulation of non-pathogenic Gram-negative bacteria-induced NF-kappaB signalling and IL-8 gene expression in IEC cocultured with immune cells and suggests the presence of mechanisms that assure hyporesponsiveness of the intestinal epithelium to certain commensally enteric bacteria.
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PMID:Differential effect of immune cells on non-pathogenic Gram-negative bacteria-induced nuclear factor-kappaB activation and pro-inflammatory gene expression in intestinal epithelial cells. 1514 74

MD-2, a glycoprotein that is essential for the innate response to lipopolysaccharide (LPS), binds to both LPS and the extracellular domain of Toll-like receptor 4 (TLR4). Following synthesis, MD-2 is either secreted directly into the medium as a soluble, active protein, or binds directly to TLR4 in the endoplasmic reticulum before migrating to the cell surface. Here we investigate the function of the secreted form of MD-2. We show that secreted MD-2 irreversibly loses activity over a 24-h period at physiological temperature. LPS, but not lipid A, prevents this loss in activity by forming a stable complex with MD-2, in a CD14-dependent process. Once formed, the stable MD-2.LPS complex activates TLR4 in the absence of CD14 or free LPS indicating that the activating ligand of TLR4 is the MD-2.LPS complex. Finally we show that the MD-2.LPS complex, but not LPS alone, induces epithelial cells, which express TLR4 but not MD-2, to secrete interleukin-6 and interleukin-8. We propose that the soluble MD-2.LPS complex plays a crucial role in the LPS response by activating epithelial and other TLR4(+)/MD-2(-) cells in the inflammatory microenvironment.
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PMID:A complex of soluble MD-2 and lipopolysaccharide serves as an activating ligand for Toll-like receptor 4. 1517 34

Toll-like receptor 4 (TLR4) and MD-2 recognizes lipid A, the active moiety of microbial lipopolysaccharide (LPS). Little is known about mechanisms for LPS recognition by TLR4-MD-2. Here we show ligand-induced TLR4 oligomerization, homotypic interaction of TLR4, which directly leads to TLR4 signaling. Since TLR4 oligomerization normally occurred in the absence of the cytoplasmic portion of TLR4, TLR4 oligomerization works upstream of TLR4 signaling. Lipid IVa, a lipid A precursor, is agonistic on mouse TLR4-MD-2 but turns antagonistic on chimeric mouse TLR4-human MD-2, demonstrating that the antagonistic activity of lipid IVa is determined by human MD-2. Binding studies with radioactive lipid A and lipid IVa revealed that lipid IVa is similar to lipid A in dose-dependent and saturable binding to mouse TLR4-human MD-2. Lipid IVa, however, did not induce TLR4 oligomerization, and inhibited lipid A-dependent oligomerization of mouse TLR4-human MD-2. Thus, lipid IVa binds mouse TLR4-human MD-2 but does not trigger TLR4 oligomerization. Binding study further revealed that the antagonistic activity of lipid IVa correlates with augmented maximal binding to mouse TLR4-human MD-2, which was approximately 2-fold higher than lipid A. Taken together, lipid A antagonist lipid IVa is distinct from lipid A in binding to TLR4-MD-2 and in subsequent triggering of TLR4 oligomerization. Given that the antagonistic activity of lipid IVa is determined by MD-2, MD-2 has an important role in a link between ligand interaction and TLR4 oligomerization.
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PMID:Lipid A antagonist, lipid IVa, is distinct from lipid A in interaction with Toll-like receptor 4 (TLR4)-MD-2 and ligand-induced TLR4 oligomerization. 1518 44

Mammalian responses to bacterial LPS (lipopolysaccharide) from the outer membrane of Gram-negative bacteria can lead to an uncontrolled inflammatory response that can be deadly for the host. It has been shown that the innate immune system employs at least three cell surface receptors, CD14, TLR4 (Toll-like receptor 4) and MD-2, in order to recognize bacterial LPS. In our previous work we have found that Hsps (heat-shock proteins) are also involved in the innate recognition of bacterial products. Their presence on the cell surface, as well as their involvement in the innate recognition process, are poorly understood. In the present study we have investigated the association of TLR4 with Hsp70 and Hsp90 following LPS stimulation, both on the cell surface and intracellularly. Our results show that Hsp70 and Hsp90 form a cluster with TLR4 within lipid microdomains following LPS stimulation. In addition, Hsp70 and Hsp90 seem to be involved in TLR4/LPS trafficking and targeting to the Golgi apparatus, since upon LPS stimulation we found that both Hsps are targeted to the Golgi along with TLR4. The present study sheds new light into the involvement of Hsps in the innate immune response.
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PMID:Heat-shock protein 70 and heat-shock protein 90 associate with Toll-like receptor 4 in response to bacterial lipopolysaccharide. 1527 Jun 95

Lipopolysaccharides are potent inflammatory mediators considered to contribute to destruction of periodontal tissues. Here, we hypothesized that Porphyromonas gingivalis lipopolysaccharide (P-LPS) treatment would regulate gene expression in murine cementoblasts through Toll-like receptor 4. Real-time (RT)-PCR and Northern blot analysis indicated that P-LPS decreased expression of transcripts for osteocalcin (OCN) and receptor activator of nuclear factor kappaB ligand (RANKL). In contrast, a dose-dependent up-regulation in mRNA levels for osteopontin (OPN) and osteoprotegerin (OPG) was observed. Similarly, ELISA demonstrated decreased RANKL and increased OPG levels. A monoclonal antibody specific for mouse TLR-4/MD-2 partially neutralized the P-LPS effect on cementoblasts. These results indicate that exposure of cementoblasts to P-LPS can alter cell function by regulating markers of osteoclastic activity (e.g., RANKL/OPG), thereby potentially affecting the inflammation-associated resorption of mineralized tissues.
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PMID:Cementoblast gene expression is regulated by Porphyromonas gingivalis lipopolysaccharide partially via toll-like receptor-4/MD-2. 1527 67

As typical PRRs (pattern-recognition receptors), TLRs (Toll-like receptors) play an important role during innate immunity recognition. MD-2 (myeloid differential protein-2) may contain distinct functional domains that can separately and simultaneously bind TLRs (TLR4 or TLR2) and TLR ligands, such as lipopolysaccharide (LPS). The special structure of MD-2 may result in its three main functions: (1) An association with TLR4 that amplifies TLR4 responsiveness to ligands, especially LPS. (2) Enabling TLR2-mediated responses to LPS and enhancing TLR2-mediated responses to bacteria and their cell wall components. (3) Increasing the expression of TLR2 and TLR4 and possibly influencing the correct intracellular distribution of TLR4. Importantly, while MD-2 regulation of TLR expression and distribution is well established, determining whether the interaction is direct or not will require further study. Thus, MD-2 is not only an assistant molecule of TLR4 but is also a key regulatory molecule in innate immunity, and may play important roles during infection, inflammation, immune responses and many other pathologic and physiologic processes.
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PMID:[The role of myeloid differential protein-2 in innate immunity]. 1528 20


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