Ubiquitin-dependent down-regulation of the neurokinin-1 receptor

Transient stimulation with substance P (SP) induces endocytosis and recycling of the neurokinin-1 receptor (NK(1)R). The effects of sustained stimulation by high concentrations of SP on NK(1)R trafficking and Ca(2+) signaling, as may occur during chronic inflammation and pain, are unknown. Chronic exposure to SP (100 nm, 3 h) completely desensitized Ca(2+) signaling by wild-type NK(1)R (NK(1)Rwt). Resensitization occurred after 16 h, and cycloheximide prevented resensitization, implicating new receptor synthesis. Lysine ubiquitination of G-protein-coupled receptors is a signal for their trafficking and degradation. Lysine-deficient mutant receptors (NK(1)RDelta5K/R, C-terminal tail lysines; and NK(1)RDelta10K/R, all intracellular lysines) were expressed at the plasma membrane and were functional because they responded to SP by endocytosis and by mobilization of Ca(2+) ions. SP desensitized NK(1)Rwt, NK(1)RDelta5K/R, and NK(1)RDelta10K/R. However, NK(1)RDelta5K/R and NK(1)RDelta10K/R resensitized 4-8-fold faster than NK(1)Rwt by cycloheximide-independent mechanisms. NK(1)RDelta325 (a naturally occurring truncated variant) showed incomplete desensitization, followed by a marked sensitization of signaling. Upon labeling receptors in living cells using antibodies to extracellular epitopes, we observed that SP induced endocytosis of NK(1)Rwt, NK(1)RDelta5K/R, and NK(1)RDelta10K/R. After 4 h in SP-free medium, NK(1)RDelta5K/R and NK(1)RDelta10K/R recycled to the plasma membrane, whereas NK(1)Rwt remained internalized. SP induced ubiquitination of NK(1)Rwt and NK(1)RDelta5K/R as determined by immunoprecipitation under nondenaturing and denaturing conditions and detected with antibodies for mono- and polyubiquitin. NK(1)RDelta10K/R was not ubiquitinated. Whereas SP induced degradation of NK(1)Rwt, NK(1)RDelta5K/R and NK(1)RDelta10K/R showed approximately 50% diminished degradation. Thus, chronic stimulation with SP induces ubiquitination of the NK(1)R, which mediates its degradation and down-regulation.

c-Cbl mediates ubiquitination, degradation, and down-regulation of human protease-activated receptor 2

Mechanisms that arrest G-protein-coupled receptor (GPCR) signaling prevent uncontrolled stimulation that could cause disease. Although uncoupling from heterotrimeric G-proteins, which transiently arrests signaling, is well described, little is known about the mechanisms that permanently arrest signaling. Here we reported on the mechanisms that terminate signaling by protease-activated receptor 2 (PAR(2)), which mediated the proinflammatory and nociceptive actions of proteases. Given its irreversible mechanism of proteolytic activation, PAR(2) is a model to study the permanent arrest of GPCR signaling. By immunoprecipitation and immunoblotting, we observed that activated PAR(2) was mono-ubiquitinated. Immunofluorescence indicated that activated PAR(2) translocated from the plasma membrane to early endosomes and lysosomes where it was degraded, as determined by immunoblotting. Mutant PAR(2) lacking intracellular lysine residues (PAR(2)Delta14K/R) was expressed at the plasma membrane and signaled normally but was not ubiquitinated. Activated PAR(2) Delta14K/R internalized but was retained in early endosomes and avoided lysosomal degradation. Activation of wild type PAR(2) stimulated tyrosine phosphorylation of the ubiquitin-protein isopeptide ligase c-Cbl and promoted its interaction with PAR(2) at the plasma membrane and in endosomes in an Src-dependent manner. Dominant negative c-Cbl lacking the ring finger domain inhibited PAR(2) ubiquitination and induced retention in early endosomes, thereby impeding lysosomal degradation. Although wild type PAR(2) was degraded, and recovery of agonist responses required synthesis of new receptors, lysine mutation and dominant negative c-Cbl impeded receptor ubiquitination and degradation and allowed PAR(2) to recycle and continue to signal. Thus, c-Cbl mediated ubiquitination and lysosomal degradation of PAR(2) to irrevocably terminate signaling by this and perhaps other GPCRs.

Identification of critical residues in the active site of porcine membrane-bound aminopeptidase P

The membrane-bound form of mammalian aminopeptidase P (AP-P; EC 3.4. 11.9) is a mono-zinc-containing enzyme that lacks any of the typical metal binding motifs found in other zinc metalloproteases. To identify residues involved in metal binding and catalysis, sequence and structural information was used to align the sequence of porcine membrane-bound AP-P with other members of the peptidase clan MG, including Escherichia coli AP-P and methionyl aminopeptidases. Residues predicted to be critical for activity were mutated and the resultant proteins were expressed in COS-1 cells. Immunoelectrophoretic blot analysis was used to compare the levels of expression of the mutant proteins, and their ability to hydrolyze bradykinin and Gly-Pro-hydroxyPro was assessed. Asp449, Asp460, His523, Glu554, and Glu568 are predicted to serve as metal ion ligands in the active site, and mutagenesis of these residues resulted in fully glycosylated proteins that were catalytically inactive. Mutation of His429 and His532 also resulted in catalytically inactive proteins, and these residues, by analogy with E. coli AP-P, are likely to play a role in shuttling protons during catalysis. These studies indicate that mammalian membrane-bound AP-P has an active-site configuration similar to that of other members of the peptidase clan MG, which is compatible with either a dual metal ion model or a single metal ion in the active site. The latter model is consistent, however, with the known metal stoichiometry of both the membrane-bound and cytosolic forms of AP-P and with a recently proposed model for methionyl aminopeptidase.

Effect of anionic co-ligands on structure and magnetic coupling of bis(μ-phenoxo-bridged dinuclear copper(II) complexes

Two phenoxo bridged dinuclear Cu(II) complexes, [Cu2L2(NO2)(2)] (1) and [Cu2L2(NO3)(2)] (2) have been synthesized using the tridentate reduced Schiff-base ligand 2-[(2-dimethylamino-ethylamino)-methyl]-phenol (HL). The complexes have been characterized by X-ray structural analyses and variable-temperature magnetic susceptibility measurements. The structures of the two compounds are very similar having the same tridentate chelating ligand (L) and mono-dentate anionic ligand nitrite for 1 and nitrate for 2. In both complexes Cu(II) is penta-coordinated but the square pyramidal geometry of the copper ions is severely distorted (Addison parameter (tau) = 0.33) in 1 while the distortion is quite small (average tau = 0.11) in 2. These differences have marked effect on the magnetic properties of two compounds. Although both are antiferromagnetically coupled, the coupling constants (J = -140.8 and -614.7 cm (1) for 1 and 2, respectively) show that the coupling is much stronger in 2.

Complexes of palladium using di-2-pyridyl ketone as ligand: synthesis, structure and, spectral and catalytic properties

During the reaction of di-2-pyridyl ketone (dpk) with Na(2)[PdCl(4)] in alcoholic media, the C=O fragment of dpk undergoes facile solvolysis and the transformed di-2-pyridyl ketone (dpk(ROH), R = Me or H) binds to palladium as NN-donor. When the reaction is carried out in refluxing methanol, a mono-complex of type [Pd(dpk(MeOH))Cl(2)] is obtained. A similar reaction in ethanol affords a bis-complex of type [Pd(dpk(ROH))(2)]Cl(2). Structure of both the complexes have been determined by X-ray crystallography. In acetonitrile solution the [pd(dpk(MeOH))Cl(2)] and [pd(dpk(ROH))(2)]Cl(2) complexes show intense absorptions in the visible and ultraviolet region, origin of which has been probed through uvr calculations. These two palladium complexes are found to be efficient catalysts for Suzuki cross-coupling reactions.

Synthesis and spectroscopic properties of cobalt(III) complexes of some aroyl hydrazones: x-ray crystal structures of one cobalt(III) complex and two aroyl hydrazone ligands

Cobalt(III) complexes of diacetyl monooxime benzoyl hydrazone (dmoBH(2)) and diacetyl monooxime isonicotinoyl hydrazone (dmoInH(2)) have been synthesized and characterized by elemental analyses and spectroscopic methods. The X-ray crystal structures of the two hydrazone ligands, as well as that of the cobalt(III) complex [Co(III)(dmoInH)(2)]Cl center dot 2H(2)O, are also reported. It is found that in the cobalt(III) complexes the Co(III) ion is hexa-coordinated, the hydrazone ligands behaving as mono-anionic tridentate O,N,N donors. In the [Co(III)(dmoInH) (2)]Cl center dot 2H(2)O complex, the amide and the oxime hydrogens are deprotonated for both the ligands, while the isonicotine nitrogens are protonated. In the [Co(III)(d-moBH)(2)] Cl complex, only the amide nitrogens are deprotonated. It is shown that the additional hydrogen bonding capability of the isonicotine nitrogen results in different conformation and supramolecular structure for dmoInH(2), compared to dmoBH(2), in the solid state. Comparing the structure of the [CoIII(dmoInH)(2)]Cl center dot 2H(2)O with that of the Zn(II) complex of the same ligand, reported earlier, it is seen that the metal ion has a profound influence on the supramolecular structure, due to change in geometrical dispositions of the chelate rings.

Is fatty acid intake a predictor of arterial stiffness and blood pressure in men? Evidence from the Caerphilly Prospective Study

Background and aims: Arterial stiffness is an independent predictor of cardiovascular disease (CVD) events and all-cause mortality and may be differentially affected by dietary fatty acid (FA) intake. The aim of this study was to investigate the relationship between FA consumption and arterial stiffness and blood pressure in a community-based population. Methods and results: The Caerphilly Prospective Study recruited 2398 men, aged 45-59 years, who were followed up at 5-year intervals for a mean of 17.8-years (n 787). A semi-quantitative food frequency questionnaire estimated intakes of total, saturated, mono- and poly-unsaturated fatty acids (SFA, MUFA, PUFA). Multiple regression models investigated associations between intakes of FA at baseline with aortic pulse wave velocity (aPWV), augmentation index (AIx), systolic and diastolic blood pressure (SBP, DBP) and pulse pressure after a 17.8-year follow-up - as well as cross-sectional relationships with metabolic markers. After adjustment, higher SFA consumption at baseline was associated with higher SBP (P = 0.043) and DBP (P = 0.002) and after a 17.8-year follow-up was associated with a 0.51 m/s higher aPWV (P = 0.006). After adjustment, higher PUFA consumption at baseline was associated with lower SBP (P = 0.022) and DBP (P = 0.036) and after a 17.8-year follow-up was associated with a 0.63 m/s lower aPWV (P = 0.007). Conclusion: This study suggests that consumption of SFA and PUFA have opposing effects on arterial stiffness and blood pressure. Importantly, this study suggests that consumption of FA is an important risk factor for arterial stiffness and CVD.

A global assessment of the effects of climate policy on the impacts of climate change

This study presents the first global-scale multi-sectoral regional assessment of the magnitude and uncertainty in the impacts of climate change avoided by emissions policies. The analysis suggests that the most stringent emissions policy considered here – which gives a 50% chance of remaining below a 2oC temperature rise target - reduces impacts by 20-65% by 2100 relative to a ‘business-as-usual’ pathway (A1B) which reaches 4oC, and can delay impacts by several decades. Effects vary between sector and region, and there are few noticeable effects of mitigation policy by 2030. The impacts avoided by 2100 are more strongly influenced by the date and level at which emissions peak than the rate of decline of emissions, with an earlier and lower emissions peak avoiding more impacts. The estimated proportion of impacts avoided at the global scale is relatively robust despite uncertainty in the spatial pattern of climate change, but the absolute amount of avoided impacts is considerably more variable and therefore uncertain.

Synthesis and characterization of nickel(II) and copper(II) complexes with tetradentate Schiff base ligands

The 1:1 condensation of 1,2-diaminopropane and 1-phenylbutane-1,3-dione at high dilution gives a mixture of two positional isomers of terdentate mono-condensed Schiff bases 6-amino-3-methyl-1-phenyl-4-aza-2-hepten-1-one (HAMPAH) and 6-amino-3,5-dimethyl-1-phenyl-4-aza-2-hexen-1-one (HADPAH). The mixture of the terdentate ligands has been used for further condensation with pyridine-2-carboxaldehyde or 2-acetylpyridine to obtain the unsymmetrical tetradentate Schiff base ligands. The tetradentate Schiff bases are then allowed to react with the methanol solution of copper(II) and nickel(II) perchlorate separately. The X-ray diffraction confirms the structures of two of the complexes and shows that the condensation site of the diamine with 1-phenylbutane-1,3-dione is the same.

Proceedings of the inaugural construction management and economics ‘Past, Present and Future’ conference CME25, 16-18 July 2007, University of Reading, UK

This conference was an unusual and interesting event. Celebrating 25 years of Construction Management and Economics provides us with an opportunity to reflect on the research that has been reported over the years, to consider where we are now, and to think about the future of academic research in this area. Hence the sub-title of this conference: “past, present and future”. Looking through these papers, some things are clear. First, the range of topics considered interesting has expanded hugely since the journal was first published. Second, the research methods are also more diverse. Third, the involvement of wider groups of stakeholder is evident. There is a danger that this might lead to dilution of the field. But my instinct has always been to argue against the notion that Construction Management and Economics represents a discipline, as such. Granted, there are plenty of university departments around the world that would justify the idea of a discipline. But the vast majority of academic departments who contribute to the life of this journal carry different names to this. Indeed, the range and breadth of methodological approaches to the research reported in Construction Management and Economics indicates that there are several different academic disciplines being brought to bear on the construction sector. Some papers are based on economics, some on psychology and others on operational research, sociology, law, statistics, information technology, and so on. This is why I maintain that construction management is not an academic discipline, but a field of study to which a range of academic disciplines are applied. This may be why it is so interesting to be involved in this journal. The problems to which the papers are applied develop and grow. But the broad topics of the earliest papers in the journal are still relevant today. What has changed a lot is our interpretation of the problems that confront the construction sector all over the world, and the methodological approaches to resolving them. There is a constant difficulty in dealing with topics as inherently practical as these. While the demands of the academic world are driven by the need for the rigorous application of sound methods, the demands of the practical world are quite different. It can be difficult to meet the needs of both sets of stakeholders at the same time. However, increasing numbers of postgraduate courses in our area result in larger numbers of practitioners with a deeper appreciation of what research is all about, and how to interpret and apply the lessons from research. It also seems that there are contributions coming not just from construction-related university departments, but also from departments with identifiable methodological traditions of their own. I like to think that our authors can publish in journals beyond the construction-related areas, to disseminate their theoretical insights into other disciplines, and to contribute to the strength of this journal by citing our articles in more mono-disciplinary journals. This would contribute to the future of the journal in a very strong and developmental way. The greatest danger we face is in excessive self-citation, i.e. referring only to sources within the CM&E literature or, worse, referring only to other articles in the same journal. The only way to ensure a strong and influential position for journals and university departments like ours is to be sure that our work is informing other academic disciplines. This is what I would see as the future, our logical next step. If, as a community of researchers, we are not producing papers that challenge and inform the fundamentals of research methods and analytical processes, then no matter how practically relevant our output is to the industry, it will remain derivative and secondary, based on the methodological insights of others. The balancing act between methodological rigour and practical relevance is a difficult one, but not, of course, a balance that has to be struck in every single paper.

Becoming poverty focused: implications for health actors

The article explores the relationship between health sector interventions and poverty analysis. It is suggested that a dynamic asset approach to poverty and health provides a framework for intervention that recognizes the complex strategies adopted by poor individuals, households and communities. The linked nature of the asset approach leads to an inter-sectoral focus and provides extra stimulation to engage with diverse partners who may be slow to own poverty reduction and health policies.

Economic transformation in Ghana: where will the path lead?

In the context of the Ghanaian government’s objective of structural transformation with an emphasis on manufacturing, this paper provides a case study of economic transformation in Ghana, exploring patterns of growth, sectoral transformation, and agglomeration. We document and examine why, despite impressive growth and poverty reduction figures, Ghana’s economy has exhibited less transformation than might be expected for a country that has recently achieved middle-income status. Ghana’s reduced share of agriculture in the economy, unlike many successfully transformed countries in Asia and Latin America, has been filled by services, while manufacturing has stagnated and even declined. Likely causes include weak transformation of the agricultural sector and therefore little development of agroprocessing, the emergence of consumption cities and consumption-driven growth, upward pressure on the exchange rate, weak production linkages, and a poor environment for private-sector-led manufacturing.

A new scenario framework for climate change research: background, process, and future directions

The scientific community is developing new global, regional, and sectoral scenarios to facilitate interdisciplinary research and assessment to explore the range of possible future climates and related physical changes that could pose risks to human and natural systems; how these changes could interact with social, economic, and environmental development pathways; the degree to which mitigation and adaptation policies can avoid and reduce risks; the costs and benefits of various policy mixes; residual impacts under alternative pathways; and the relationship of future climate change and adaptation and mitigation policy responses with sustainable development. This paper provides the background to and process of developing the conceptual framework for these scenarios, as described in the three subsequent papers in this Special Issue (Van Vuuren et al.; O’Neill et al.; Kriegler et al.). The paper also discusses research needs to further develop and apply this framework. A key goal of the current framework design and its future development is to facilitate the collaboration of climate change researchers from a broad range of perspectives and disciplines to develop policy- and decision-relevant scenarios and explore the challenges and opportunities human and natural systems could face with additional climate change.

Solvent-Dependent Formation of Os(0) Complexes by Electrochemical Reduction of [Os(CO)(2,2′-bipyridine)(L)Cl2]; L = Cl−,PrCN

Cyclic voltammetry and ultraviolet−visible/infrared (UV−vis/IR) spectroelectrochemistry were used to study the cathodic electrochemical behavior of the osmium complexes mer-[OsIII(CO) (bpy)Cl3] (bpy = 2,2′-bipyridine) and trans(Cl)-[OsII(CO) (PrCN)(bpy)Cl2] at variable temperature in different solvents (tetrahydrofuran (THF), butyronitrile (PrCN), acetonitrile (MeCN)) and electrolytes (Bu4NPF6, Bu4NCl). The precursors can be reduced to mer-[OsII(CO) (bpy•−)Cl3]2− and trans(Cl)-[OsII(CO)(PrCN) (bpy•−)Cl2]−, respectively, which react rapidly at room temperature, losing the chloride ligands and forming Os(0) species. mer-[OsIII(CO) (bpy)Cl3] is reduced in THF to give ultimately an Os−Os-bonded polymer, probably [Os0(CO) (THF)-(bpy)]n, whereas in PrCN the well-soluble, probably mononuclear [Os0(CO) (PrCN)(bpy)], species is formed. The same products were observed for the 2 electron reduction of trans(Cl)-[OsII(CO)(PrCN) (bpy)Cl2] in both solvents. In MeCN, similar to THF, the[Os0(CO) (MeCN)(bpy)]n polymer is produced. It is noteworthy that the bpy ligand in mononuclear [Os0(CO) (PrCN)(bpy)] is reduced to the corresponding radical anion at a significantly less negative potential than it is in polymeric [Os0(CO) (THF)(bpy)]n: ΔE1/2 = 0.67 V. Major differences also exist in the IR spectra of the Os(0) species: the polymer shows a broad ν(CO) band at much smaller wavenumbers compared to the soluble Os(0) monomer that exhibits a characteristic ν(Pr-CN) band below 2200 cm−1 in addition to the intense and narrow ν(CO) absorption band. For the first time, in this work the M0-bpy(M = Ru, Os) mono- and dicarbonyl species soluble in PrCN have been formulated as a mononuclear complex. Density functional theory (DFT) and time-dependent-DFT calculations confirm the Os(0) oxidation state and suggest that [Os0(CO)(PrCN)(bpy)] is a square planar moiety. The reversible bpy-based reduction of [Os0(CO) (PrCN)(bpy)] triggers catalytic reduction of CO2 to CO and HCOO−.

From farmers’ perspective : pesticide use and pest control

Many studies have shown that farmers in developing countries often overuse pesticides and do not adopt safety practices. Policies and interventions to promote a safer use of pesticides are often based on a limited understanding of the farmers’ own perspective of pesticide use. This often results in ineffective policies and the persistence of significant pesticide-related health and environmental problems, especially in developing countries. This chapter explores potentials and limitations of different approaches to study pesticide use in agriculture from the farmers’ perspective. In contrast to the reductionist and mono-disciplinary approaches often adopted, this chapter calls for integrative methodological approaches to provide a realistic and thorough understanding of the farmers’ perspective on pesticide use and illustrates the added value of such an approach with three case studies of pesticide use in Iran, India, and Colombia.