Industrial constructions of publics and public knowledge: a qualitative investigation of practice in the UK chemicals industry

This is a post print version of the article. The official published version can be obtained from the link below - © 2007 by SAGE PublicationsWhile the rhetoric of public engagement is increasingly commonplace within industry, there has been little research that examines how lay knowledge is conceptualized and whether it is really used within companies. Using the chemicals sector as an example, this paper explores how companies conceive of publics and "public knowledge," and how this relates to modes of engagement/communication with them. Drawing on qualitative empirical research in four companies, we demonstrate that the public for industry are primarily conceived as "consumers" and "neighbours," having concerns that should be allayed rather than as groups with knowledge meriting engagement. We conclude by highlighting the dissonance between current advocacy of engagement and the discourses and practices prevalent within industry, and highlight the need for more realistic strategies for industry/public engagement.Funding was received from the ESRC Science in Society Programme

Contribution of NADPH Oxidase to Membrane CD38 Internalization and Activation in Coronary Arterial Myocytes

The CD38-ADP-ribosylcyclase-mediated Ca2+ signaling pathway importantly contributes to the vasomotor response in different arteries. Although there is evidence indicating that the activation of CD38-ADP-ribosylcyclase is associated with CD38 internalization, the molecular mechanism mediating CD38 internalization and consequent activation in response to a variety of physiological and pathological stimuli remains poorly understood. Recent studies have shown that CD38 may sense redox signals and is thereby activated to produce cellular response and that the NADPH oxidase isoform, NOX1, is a major resource to produce superoxide (O2·−) in coronary arterial myocytes (CAMs) in response to muscarinic receptor agonist, which uses CD38-ADP-ribosylcyclase signaling pathway to exert its action in these CAMs. These findings led us hypothesize that NOX1-derived O2·− serves in an autocrine fashion to enhance CD38 internalization, leading to redox activation of CD38-ADP-ribosylcyclase activity in mouse CAMs. To test this hypothesis, confocal microscopy, flow cytometry and a membrane protein biotinylation assay were used in the present study. We first demonstrated that CD38 internalization induced by endothelin-1 (ET-1) was inhibited by silencing of NOX1 gene, but not NOX4 gene. Correspondingly, NOX1 gene silencing abolished ET-1-induced O2·− production and increased CD38-ADP-ribosylcyclase activity in CAMs, while activation of NOX1 by overexpression of Rac1 or Vav2 or administration of exogenous O2·−significantly increased CD38 internalization in CAMs. Lastly, ET-1 was found to markedly increase membrane raft clustering as shown by increased colocalization of cholera toxin-B with CD38 and NOX1. Taken together, these results provide direct evidence that Rac1-NOX1-dependent O2·− production mediates CD38 internalization in CAMs, which may represent an important mechanism linking receptor activation with CD38 activity in these cells

Molecular Characterization of a Novel Intracellular ADP-Ribosyl Cyclase

Background. ADP-ribosyl cyclases are remarkable enzymes capable of catalyzing multiple reactions including the synthesis of the novel and potent intracellular calcium mobilizing messengers, cyclic ADP-ribose and NAADP. Not all ADP-ribosyl cyclases however have been characterized at the molecular level. Moreover, those that have are located predominately at the outer cell surface and thus away from their cytosolic substrates. Methodology/Principal Findings. Here we report the molecular cloning of a novel expanded family of ADP-ribosyl cyclases from the sea urchin, an extensively used model organism for the study of inositol trisphosphate-independent calcium mobilization. We provide evidence that one of the isoforms (SpARC1) is a soluble protein that is targeted exclusively to the endoplasmic reticulum lumen when heterologously expressed. Catalytic activity of the recombinant protein was readily demonstrable in crude cell homogenates, even under conditions where luminal continuity was maintained. Conclusions/Significance. Our data reveal a new intracellular location for ADP-ribosyl cyclases and suggest that production of calcium mobilizing messengers may be compartmentalized

Extracellular NAD and ATP: Partners in immune cell modulation

Extracellular NAD and ATP exert multiple, partially overlapping effects on immune cells. Catabolism of both nucleotides by extracellular enzymes keeps extracellular concentrations low under steady-state conditions and generates metabolites that are themselves signal transducers. ATP and its metabolites signal through purinergic P2 and P1 receptors, whereas extracellular NAD exerts its effects by serving as a substrate for ADP-ribosyltransferases (ARTs) and NAD glycohydrolases/ADPR cyclases like CD38 and CD157. Both nucleotides activate the P2X7 purinoceptor, although by different mechanisms and with different characteristics. While ATP activates P2X7 directly as a soluble ligand, activation via NAD occurs by ART-dependent ADP-ribosylation of cell surface proteins, providing an immobilised ligand. P2X7 activation by either route leads to phosphatidylserine exposure, shedding of CD62L, and ultimately to cell death. Activation by ATP requires high micromolar concentrations of nucleotide and is readily reversible, whereas NAD-dependent stimulation begins at low micromolar concentrations and is more stable. Under conditions of cell stress or inflammation, ATP and NAD are released into the extracellular space from intracellular stores by lytic and non-lytic mechanisms, and may serve as ‘danger signals–to alert the immune response to tissue damage. Since ART expression is limited to naïve/resting T cells, P2X7-mediated NAD-induced cell death (NICD) specifically targets this cell population. In inflamed tissue, NICD may inhibit bystander activation of unprimed T cells, reducing the risk of autoimmunity. In draining lymph nodes, NICD may eliminate regulatory T cells or provide space for the preferential expansion of primed cells, and thus help to augment an immune response

Functional visualization of the separate but interacting calcium stores sensitive to NAADP and cyclic ADP-ribose

Cells possess multiple Ca2+ stores and their selective mobilization provides the spatial-temporal Ca2+ signals crucial in regulating diverse cellular functions. Except for the inositol trisphosphate (IP3)-sensitive Ca2+ stores, the identities and the mechanisms of how these internal stores are mobilized are largely unknown. In this study, we describe two Ca2+ stores, one of which is regulated by cyclic ADP-ribose (cADPR) and the other by nicotinic acid adenine dinucleotide phosphate (NAADP). We took advantage of the large size of the sea urchin egg and stratified its organelles by centrifugation. Using photolysis to produce either uniform or localized increases of cADPR and NAADP from their respective caged analogs, the two separate stores could be visually identified by Ca2+ imaging and shown to be segregated to the opposite poles of the eggs. The cADPR-pole also contained the IP3-sensitive Ca2+ stores, the egg nucleus and the endoplasmic reticulum (ER); the latter was visualized using Bodipy-thapsigargin. On the other hand, the mitochondria, as visualized by rhodamine 123, were segregated to the opposite pole together with the NAADP-sensitive calcium stores. Fertilization of the stratified eggs elicited a Ca2+ wave starting at the cADPR-pole and propagating toward the NAADP-pole. These results provide the first direct and visual evidence that the NAADP-sensitive Ca2+ stores are novel and distinct from the ER. During fertilization, communicating signals appear to be transmitted from the ER to NAADP-sensitive Ca2+ stores, leading to their activation.link_to_subscribed_fulltex

Structural determinants of nicotinic acid adenine dinucleotide phosphate important for its calcium-mobilizing activity

Nicotinic acid adenine dinucleotide phosphate (NAADP) mobilizes Ca2+ through a mechanism totally independent of cyclic ADP-ribose or inositol trisphosphate. The structural determinants important for its Ca2+ release activity were investigated using a series of analogs. It is shown that changing the 3-carboxyl group of the nicotinic acid (NA) moiety in NAADP to either an uncharged carbinol or from the 3-position to the 4-position of the pyridine ring totally eliminates the Ca2+ release activity. Conversion of the 3-carboxyl to other negatively charged groups, either 3-sulfonate, 3- acetate, or 3-quinoline carboxylate, retains the Ca2+ release activity, although their half-maximal effective concentrations (EC50) are 100-200- fold higher. Changing the 6-amino group of the adenine to a hydroxyl group results in more than a 1000-fold decrease in the Ca2+ release activity. Conversion of the 2'-phosphate to 2',3'-cyclic phosphate or 3'-phosphate likewise increases the EC50 by about 5- and 20-fold, respectively. Similar to NAADP, all of the active analogs can also desensitize the Ca2+ release mechanism at subthreshold concentrations, suggesting that this novel property is intrinsic to the release mechanism. The series of analogs used was produced by using ADP-ribosyl cyclase to catalyze the exchange of the nicotinamide group of various analogs of NADP with various analogs of NA. An important determinant in NA that is crucial to the base exchange reaction was shown to be the 2-position of the pyridine ring. Neither pyridine-2- carboxylate nor 2-methyl-NA support the exchange reaction. The negative charge and the position of the 3-carboxyl group are nonessential since both pyridine-3-carbinol and pyridine-4-carboxylate support the base exchange reaction. In addition to the information on the structure-activity relationships of NAADP and NA, this study also demonstrates the utility of the base exchange reaction as a general approach for synthesizing NAADP analogs.link_to_subscribed_fulltex