scholarly journals Adenovirus RIDαβ Complex Inhibits Lipopolysaccharide Signaling without Altering TLR4 Cell Surface Expression

2006 ◽  
Vol 80 (13) ◽  
pp. 6378-6386 ◽  
Author(s):  
Fernando Delgado-Lopez ◽  
Marshall S. Horwitz
Keyword(s):  
Signal Transduction ◽  
Cell Surface ◽  
Signaling Pathways ◽  
Interleukin 1 ◽  
Mechanistic Explanation ◽  
Surface Expression ◽  
Receptor Internalization ◽  
Cell Surface Receptor ◽  
Cell Surface Expression ◽  
Down Regulation

ABSTRACT The transmembrane heterotrimer complex 10.4K/14.5K, also known as RID (for “receptor internalization and degradation”), is encoded by the adenovirus E3 region, and it down-regulates the cell surface expression of several unrelated receptors. We recently showed that RID expression correlates with down-regulation of the cell surface expression of the tumor necrosis factor (TNF) receptor 1 in several human cells. This observation provided the first mechanistic explanation for the inhibition of TNF alpha-induced chemokines by RID. Here we analyze the immunoregulatory activities of RID on lipopolysaccharide (LPS) and interleukin-1 beta (IL-1β)-mediated responses. Although both signaling pathways are strongly inhibited by RID, the chemokines up-regulated by IL-1β stimulation are only marginally inhibited. In addition, RID inhibits signaling induced by LPS without affecting the expression of the LPS receptor Toll-like receptor 4, demonstrating that RID need not target degradation of the receptor to alter signal transduction. Taken together, our data demonstrate the inhibitory effect of RID on two additional cell surface receptor-mediated signaling pathways involved in inflammatory processes. The data suggest that RID has intracellular targets that impair signal transduction and chemokine expression without evidence of receptor down-regulation.

Effect of aging on CD11b and CD69 surface expression by vesicular insertion in human polymorphonuclear leucocytes

1999 ◽  
Vol 97 (3) ◽  
pp. 323-329 ◽  
Author(s):  
J. M. NOBLE ◽  
G. A. FORD ◽  
T. H. THOMAS
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Surface Expression ◽  
Cell Surface Receptor ◽  
Cell Surface Expression ◽  
Elderly Subjects ◽  
Polymorphonuclear Leucocytes ◽  
Cd11b Expression ◽  
Cd69 Expression ◽  
Vesicle Exocytosis

The exocytosis of intracellular vesicles is an important function of the plasma membrane, which is responsible for hormone secretion, cell surface expression of antigens, ion transporters and receptors, and intracellular and intercellular signalling. Human aging is associated with many physiological and cellular changes, many of which are due to alterations in plasma membrane functioning. Alterations in vesicle externalization with age could account for many of these changes. We investigated whether alterations in vesicle exocytosis occur with increasing age by flow-cytometric determination of CD11b and CD69 expression on the surface of human polymorphonuclear leucocytes (PMN) stimulated with phorbol myristate acetate (PMA), a tumour promoter which binds to and activates protein kinase C (PKC) directly, or with formyl-Met-Leu-Phe (fMLP), which activates PKC indirectly via interactions with a cell surface receptor and G-protein, and subsequent inositol phosphate hydrolysis. Following stimulation with PMA, a decrease in the proportion of PMN expressing CD69 at high levels was observed in elderly compared with young subjects (young, 55.3%; elderly, 43.9%; P = 0.01). No aging-related differences in the proportion of PMN expressing CD11b (young, 73.7%; elderly, 68.4%; P = 0.15), or in the number of molecules of CD69 or CD11b expressed per cell, were observed. Stimulation with fMLP or low PMA concentrations resulted in full CD11b expression but minimal CD69 expression in both young and elderly subjects. Cells which expressed CD69 had no CD11b expression, while those cells expressing CD11b had minimal CD69 expression. Thus the PMA-induced expression of CD11b and CD69 in human PMN represents two separate processes, only one of which is affected in aging. CD11b expression appears to require a lesser degree of PKC stimulation compared with that required for CD69 expression. The age-associated reduction in PMA-stimulated CD69 expression may occur either at or distal to PKC activation. Such a decrease may contribute to the age-associated impairments in PMN function that contribute, in turn, to immunosenescence.

Assessing factors for predictive response to ADC targets in clinical tissue for companion diagnostic development.

2013 ◽  
Vol 31 (15_suppl) ◽  
pp. e22187-e22187
Author(s):  
Steven James Potts ◽  
Joseph Krueger ◽  
Holger Lange ◽  
David Eberhard ◽  
George David Young
Keyword(s):  
Tumor Microenvironment ◽  
Cell Surface ◽  
Surface Expression ◽  
Receptor Internalization ◽  
Critical Parameters ◽  
Cell Surface Expression ◽  
Cell Surface Antigens ◽  
Drug Conjugates ◽  
Coefficients Of Variation

e22187 Background: One premise of antibody-drug conjugates (ADC) is that the bound mAb-antigen complex on the cell surface will internalize and be metabolized by lysosomal proteases to release the free drug. Thus, the efficacy of an ADC is dependent not only on the presence of cell surface antigens, but also intact delivery of the conjugated drug. In most cases, the biological mechanisms behind these processes have not been elucidated. Furthermore, the extracellular stability of the ADC may be affected by the activity of the target proteases in the tumor microenvironment outside of the lysozome. These concepts are especially important for CDx approaches, which would ideally account not only for the degree of cell surface expression of the target, but also receptor internalization and potential effects of tumor microenvironment. Although in vitro approaches exist to measure these attributes, there are no clinically amenable approaches to measure these critical parameters. Methods: Flagship Biosciences has invented several proprietary approaches for measuring critical properties of the therapeutic target on the cell surface or inside the cell, as well as properties of the TME which could be used to predict efficacy to an ADC using FFPE biopsies. These image analysis approaches have been designed specifically in context of ADC CDx programs to: 1) Accurately define cell surface target expression independent of cytoplasmic expression; 2) Assess critical factors in the TME which may affect delivery of the drug to the intracellular target; and 3) Multiplex these evaluations for an integrative answer to be derived from a typical clinical biopsy. Results: The measurement of cytoplasm/membrane localization was evaluated on several hundred tissue sections, and the methodology was found to be highly reproducible, with coefficients of variation lower than that observed by manual pathologist accessment. Conclusions: Our approach discretely measures endpoints of cell surface biomarker prevalence, biomarker membrane/cytoplasmic ratio, heterogeneity, stromal contribution, inflammatory environment, and other important cell-by-cell outputs from a single FFPE slide in the context of typical drug discovery.

Down-Regulation of HLA-G1 Cell Surface Expression in Human Cytomegalovirus Infected Cells

2003 ◽  
Vol 50 (4) ◽  
pp. 328-333 ◽  
Author(s):  
Nathalie Pizzato ◽  
Barbara Garmy-Susini ◽  
Philippe Le Bouteiller ◽  
Francoise Lenfant
Keyword(s):  
Cell Surface ◽  
Human Cytomegalovirus ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Down Regulation ◽  
Infected Cells

scholarly journals Granulocyte macrophage colony stimulating factor produced in lesioned peripheral nerves induces the up-regulation of cell surface expression of MAC-2 by macrophages and Schwann cells.

1996 ◽  
Vol 133 (1) ◽  
pp. 159-167 ◽  
Author(s):  
A Saada ◽  
F Reichert ◽  
S Rotshenker
Keyword(s):  
Cell Surface ◽  
Schwann Cells ◽  
Interleukin 1 ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Gm Csf ◽  
Molecular Events ◽  
Macrophage Colony

Peripheral nerve injury is followed by Wallerian degeneration which is characterized by cellular and molecular events that turn the degenerating nerve into a tissue that supports nerve regeneration. One of these is the removal, by phagocytosis, of myelin that contains molecules which inhibit regeneration. We have recently documented that the scavenger macrophage and Schwann cells express the galactose-specific lectin MAC-2 which is significant to myelin phagocytosis. In the present study we provide evidence for a mechanism leading to the augmented expression of cell surface MAC-2. Nerve lesion causes noneuronal cells, primarily fibroblasts, to produce the cytokine granulocyte macrophage-colony stimulating factor (GM-CSF). In turn, GM-CSF induces Schwann cells and macrophages to up-regulate surface expression of MAC-2. The proposed mechanism is based on the following novel observations. GM-CSF mRNA was detected by PCR in in vitro and in vivo degenerating nerves, but not in intact nerves. The GM-CSF molecule was detected by ELISA in medium conditioned by in vitro and in vivo degenerating peripheral nerves as of the 4th h after injury. GM-CSF activity was demonstrated by two independent bioassays, and repressed by activity blocking antibodies. Significant levels of GM-CSF were produced by nerve derived fibroblasts, but neither by Schwann cells nor by nerve derived macrophages. Mouse rGM-CSF enhanced MAC-2 production in nerve explants, and up-regulated cell surface expression of MAC-2 by Schwann cells and macrophages. Interleukin-1 beta up-regulated GM-CSF production thus suggesting that injury induced GM-CSF production may be mediated by interleukin-1 beta. Our findings highlight the fact that fibroblasts, by producing GM-CSF and thereby affecting macrophage and Schwann function, play a significant role in the cascade of molecular events and cellular interactions of Wallerian degeneration.

scholarly journals The PAR2 signal peptide prevents premature receptor cleavage and activation

2019 ◽  
Author(s):  
Belinda Liu ◽  
Grace Lee ◽  
Jiejun Wu ◽  
Janise Deming ◽  
Chester Kuei ◽  
...  
Keyword(s):  
Cell Surface ◽  
Signal Peptide ◽  
Synthetic Peptide ◽  
Surface Expression ◽  
Receptor Expression ◽  
Cell Surface Receptor ◽  
Cell Surface Expression ◽  
Peptide Ligand ◽  
Protease Activated Receptors ◽  
N Terminus

AbstractUnlike closely related GPCRs, protease-activated receptors (PAR1, PAR2, PAR3, and PAR4) have a predicted signal peptide at their N-terminus, which is encoded by a separate exon, suggesting that the signal peptides of PARs may serve an important and unique function, specific for PARs. In this report, we show that the PAR2 signal peptide, when fused to the N-terminus of IgG-Fc, effectively induced IgG-Fc secretion into culture medium, thus behaving like a classical signal peptide. The presence of PAR2 signal peptide has a strong effect on PAR2 cell surface expression, as deletion of the signal peptide (PAR2ΔSP) led to dramatic reduction of the cell surface expression and decreased responses to trypsin or the synthetic peptide ligand (SLIGKV). However, further deletion of the tethered ligand region (SLIGKV) at the N-terminus rescued the cell surface receptor expression and the response to the synthetic peptide ligand, suggesting that the signal peptide of PAR2 may be involved in preventing PAR2 from intracellular protease activation before reaching the cell surface. Supporting this hypothesis, an Arg36Ala mutation on PAR2ΔSP, which disabled the trypsin activation site, increased the receptor cell surface expression and the response to ligand stimulation. Similar effects were observed when PAR2ΔSP expressing cells were treated with protease inhibitors. Our findings indicated that these is a role of the PAR2 signal peptide in preventing the premature activation of PAR2 from intracellular protease cleavage before reaching the cells surface. The same mechanism may also apply to PAR1, PAR3, and PAR4.

scholarly journals Aminooxypentane-RANTES Induces CCR5 Internalization but Inhibits Recycling: A Novel Inhibitory Mechanism of HIV Infectivity

1998 ◽  
Vol 187 (8) ◽  
pp. 1215-1224 ◽  
Author(s):  
Matthias Mack ◽  
Bruno Luckow ◽  
Peter J. Nelson ◽  
Josef Cihak ◽  
Graham Simmons ◽  
...  
Keyword(s):  
Cell Surface ◽  
Hiv Infection ◽  
Mononuclear Cells ◽  
Chinese Hamster ◽  
Surface Expression ◽  
Receptor Internalization ◽  
Cell Surface Expression ◽  
Macrophage Infection ◽  
Potent Inhibition ◽  
Hiv 1

CCR5, a chemokine receptor expressed on T cells and macrophages, is the principal coreceptor for M-tropic HIV-1 strains. Recently, we described an NH2-terminal modification of the CCR5 ligand regulated on activation, normal T cell expressed and secreted (RANTES), aminooxypentane-RANTES (AOP-RANTES), that showed potent inhibition of macrophage infection by HIV-1 under conditions where RANTES was barely effective. To investigate the mechanism of AOP-RANTES inhibition of HIV infectivity we examined the surface expression of CCR5 using a monoclonal anti-CCR5 antibody, MC-1. We demonstrate that AOP-RANTES rapidly caused >90% decrease in cell surface expression of CCR5 on lymphocytes, monocytes/ macrophages, and CCR5 transfected Chinese hamster ovary (CHO) cells. RANTES also caused a loss of cell surface CCR5, although its effect was less than with AOP-RANTES. Significantly, AOP-RANTES inhibited recycling of internalized CCR5 to the cell surface, whereas RANTES did not. When peripheral blood mononuclear cells are cultured for prolonged periods of time in the presence of RANTES, CCR5 expression is comparable to that seen on cells treated with control medium, whereas there is no CCR5 surface expression on cells cultured in the presence of AOP-RANTES. Immunofluorescence indicated that both AOP-RANTES and RANTES induced downmodulation of cell surface CCR5, and that the receptor was redistributed into endocytic organelles containing the transferrin receptor. When RANTES was removed, the internalized receptor was recycled to the cell surface; however, the receptor internalized in the presence of AOP-RANTES was retained in endosomes. Using human osteosarcoma (GHOST) 34/CCR5 cells, the potency of AOP-RANTES and RANTES to inhibit infection by the M-tropic HIV-1 strain, SF 162, correlated with the degree of downregulation of CCR5 induced by the two chemokines. These differences between AOP-RANTES and RANTES in their effect on receptor downregulation and recycling suggest a mechanism for the potent inhibition of HIV infection by AOP-RANTES. Moreover, these results support the notion that receptor internalization and inhibition of receptor recycling present new targets for therapeutic agents to prevent HIV infection.

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