Working Group 1: Future developments in deterministic and stochastic parametrizations

Our group considered the desirability of including representations of uncertainty in the development of parameterizations. (By ‘uncertainty’ here we mean the deviation of sub-grid scale fluxes or tendencies in any given model grid box from truth.) We unanimously agreed that the ECWMF should attempt to provide a more physical basis for uncertainty estimates than the very effective but ad hoc methods being used at present. Our discussions identified several issues that will arise.

Modulation of endothelial cell KCa3.1 Channels during endothelium-derived hyperpolarizing factor signaling in mesenteric resistance arteries

Arterial hyperpolarization to acetylcholine (ACh) reflects coactivation of KCa3.1 (IKCa) channels and KCa2.3 (SKCa) channels in the endothelium that transfers through myoendothelial gap junctions and diffusible factor(s) to affect smooth muscle relaxation (endothelium-derived hyperpolarizing factor [EDHF] response). However, ACh can differentially activate KCa3.1 and KCa2.3 channels, and we investigated the mechanisms responsible in rat mesenteric arteries. KCa3.1 channel input to EDHF hyperpolarization was enhanced by reducing external [Ca2+]o but blocked either with forskolin to activate protein kinase A or by limiting smooth muscle [Ca2+]i increases stimulated by phenylephrine depolarization. Imaging [Ca2+]i within the endothelial cell projections forming myoendothelial gap junctions revealed increases in cytoplasmic [Ca2+]i during endothelial stimulation with ACh that were unaffected by simultaneous increases in muscle [Ca2+]i evoked by phenylephrine. If gap junctions were uncoupled, KCa3.1 channels became the predominant input to EDHF hyperpolarization, and relaxation was inhibited with ouabain, implicating a crucial link through Na+/K+-ATPase. There was no evidence for an equivalent link through KCa2.3 channels nor between these channels and the putative EDHF pathway involving natriuretic peptide receptor-C. Reconstruction of confocal z-stack images from pressurized arteries revealed KCa2.3 immunostain at endothelial cell borders, including endothelial cell projections, whereas KCa3.1 channels and Na+/K+-ATPase {alpha}2/{alpha}3 subunits were highly concentrated in endothelial cell projections and adjacent to myoendothelial gap junctions. Thus, extracellular [Ca2+]o appears to modify KCa3.1 channel activity through a protein kinase A-dependent mechanism independent of changes in endothelial [Ca2+]i. The resulting hyperpolarization links to arterial relaxation largely through Na+/K+-ATPase, possibly reflecting K+ acting as an EDHF. In contrast, KCa2.3 hyperpolarization appears mainly to affect relaxation through myoendothelial gap junctions. Overall, these data suggest that K+ and myoendothelial coupling evoke EDHF-mediated relaxation through distinct, definable pathways.

Flatpack revolution

The project consists of a trilogy of films and a live performance. The Future trilogy takes IKEA riot of 2005 as the starting point for a speculative history of a fictional future, culminating in a choreographed re-enactment of the original event. Shot on 16mm and 8mm film, the series explores the possibility of collective action emerging from the capitalist relations inherent in the consumer riot. The live performance No Haus Like Bau, staged at the HAU 1 theatre in Berlin for the 5th Berlin Biennale, continues this research into re-enactment and post-1989 politics by dramatizing the rise and fall of the soviet union as a neo-Constructivist mime using a stage set made of flatpack furniture. Using the aesthetics of Modernism and the avant garde, from Constructivist and Futurist constumes to biomechanics and Bauhaus theatre theory, the project transposes early twentieth century utopian ideology to a present day setting where mass uprisings are motivated by cheap commodities. These explorations of consumerism and revolution have been widely exhibited internationally and supported by Film London, Arts Council England, Collective Gallery and the Berlin Biennale. The Future Trilogy formed the basis of a solo exhibition at the Te Tuhi Art Centre in Auckland, New Zealand and was screened as part of the Signal and Noise media art festival in Vancouver, as well as other exhibitions and screenings including “Roll it to Me” at Collective Gallery, Edinburgh, and Apocatopia, Castlefield Gallery, Manchester.

A novel, non-canonical mechanism of regulation of MST3 (mammalian Sterile20-related kinase 3)

The canonical pathway of regulation of the germinal centre kinase (GCK) III subgroup member, mammalian Sterile20-related kinase 3 (MST3), involves a caspase-mediated cleavage between N-terminal catalytic and C-terminal regulatory domains with possible concurrent autophosphorylation of the activation loop MST3(Thr178-), induction of Ser-/Thr-protein kinase activity and nuclear localisation. We identified an alternative ‘non-canonical’ pathway of MST3 activation (regulated primarily through dephosphorylation) which may also be applicable to other GCKIII (and GCKVI) subgroup members. In the basal state, inactive MST3 co-immunoprecipitated with the Golgi protein, GOLGA2/gm130. Activation of MST3 by calyculin A (a protein Ser-/Thr- phosphatase 1/2A inhibitor) stimulated (auto)phosphorylation of MST3(Thr178-) in the catalytic domain with essentially simultaneous cis-autophosphorylation of MST3(Thr328-) in the regulatory domain, an event also requiring the MST3(341-376) sequence which acts as a putative docking domain. MST3(Thr178-) phosphorylation increased MST3 kinase activity but this activity was independent of MST3(Thr328-) phosphorylation. Interestingly, MST3(Thr328-) lies immediately C-terminal to a STRAD pseudokinase-like site recently identified as being involved in binding of GCKIII/GCKVI members to MO25 scaffolding proteins. MST3(Thr178- /Thr328-) phosphorylation was concurrent with dissociation of MST3 from GOLGA2/gm130 and association of MST3 with MO25, and MST3(Thr328-) phosphorylation was necessary for formation of the activated MST3-MO25 holocomplex.

Cryopreservation of winter-dormant apple buds: 1 - variation in recovery with cultivar and winter conditions

The widely-adopted protocol for the cryopreservation of winter buds of fruit trees, such as Malus and Pyrus, was developed in a region with a continental climate, that provides relatively hard winters with a consequent effect on adaptive plant hardiness. In this study the protocol was evaluated in a typical maritime climate (eastern Denmark) where milder winters can be expected. The survival over two winters was evaluated, looking at variation between seasons and cultivars together with the progressive reduction in survival due to individual steps in the protocol. The study confirms that under such conditions significant variation in survival can be expected and that an extended period of imposed dehydration at -4oC is critical for bud survival. The occurrence of freezing events during this treatment suggests that cryodehydration may be involved, as well as evaporative water loss. To optimize the protocol for maritime environments, further investigation into the water status of the explants during cryopreservation is proposed. Keywords: Malus x domestica, cryopreservation, dormant bud, survival, grafting

Regulation of platelet biology by platelet endothelial cell adhesion molecule-1.

Platelet endothelial cell adhesion molecule-1 (PECAM-1), an immunoreceptor tyrosine-based inhibitory motif containing receptor, plays diverse and apparently contradictory roles in regulating the response of platelets to stimuli; inhibiting platelet response to immunoreceptor tyrosine-based activation motif and G protein-coupled receptor signalling following stimulation with collagen, adenosine diphosphate, and thrombin, as well as enhancing integrin outside-in signalling. These dual, and opposing, roles suggest an important and complex role for PECAM-1 in orchestrating platelet response to vascular damage. Indeed, during thrombus formation, the influence of PECAM-1 on the multiple signalling pathways combines leading to a relatively large inhibitory effect on thrombus formation.

DC-SIGN increases the affinity of HIV-1 envelope glycoprotein interaction with CD4

Mannose-binding C-type lectin receptors, expressed on Langerhans cells and subepithelial dendritic cells (DCs) of cervico-vaginal tissues, play an important role in HIV-1 capture and subsequent dissemination to lymph nodes. DC-SIGN has been implicated in both productive infection of DCs and the DC-mediated trans infection of CD4(+) T cells that occurs in the absence of replication. However, the molecular events that underlie this efficient transmission have not been fully defined. In this study, we have examined the effect of the extracellular domains of DC-SIGN and Langerin on the stability of the interaction of the HIV-1 envelope glycoprotein with CD4 and also on replication in permissive cells. Surface plasmon resonance analysis showed that DC-SIGN increases the binding affinity of trimeric gp140 envelope glycoproteins to CD4. In contrast, Langerin had no effect on the stability of the gp140:CD4 complex. In vitro infection experiments to compare DC-SIGN enhancement of CD4-dependent and CD4-independent strains demonstrated significantly lower enhancement of the CD4-independent strain. In addition DC-SIGN increased the relative rate of infection of the CD4-dependent strain but had no effect on the CD4-independent strain. DC-SIGN binding to the HIV envelope protein effectively increases exposure of the CD4 binding site, which in turn contributes to enhancement of infection.

Expression-system-dependent modulation of HIV-1 envelope glycoprotein antigenicity and immunogenicity.

Recombinant expression systems differ in the type of glycosylation they impart on expressed antigens such as the human immunodeficiency virus type 1 (HIV-1) envelope glycoproteins, potentially affecting their biological properties. We performed head-to-head antigenic, immunogenic and molecular profiling of two distantly related Env surface (gp120) antigens produced in different systems: (a) mammalian (293 FreeStyle cells; 293F) cells in the presence of kifunensine, which impart only high-mannose glycans; (b) insect cells (Spodoptera frugiperda, Sf9), which confer mainly paucimannosidic glycans; (c) Sf9 cells recombinant for mammalian glycosylation enzymes (Sf9 Mimic), which impart high-mannose, hybrid and complex glycans without sialic acid; and (d) 293F cells, which impart high-mannose, hybrid and complex glycans with sialic acid. Molecular models revealed a significant difference in gp120 glycan coverage between the Sf9-derived and wild-type mammalian-cell-derived material that is predicted to affect ligand binding sites proximal to glycans. Modeling of solvent-exposed surface electrostatic potentials showed that sialic acid imparts a significant negative surface charge that may influence gp120 antigenicity and immunogenicity. Gp120 expressed in systems that do not incorporate sialic acid displayed increased ligand binding to the CD4 binding and CD4-induced sites compared to those expressed in the system that do, and imparted other more subtle differences in antigenicity in a gp120 subtype-specific manner. Non-sialic-acid-containing gp120 was significantly more immunogenic than the sialylated version when administered in two different adjuvants, and induced higher titers of antibodies competing for CD4 binding site ligand-gp120 interaction. These findings suggest that non-sialic-acid-imparting systems yield gp120 immunogens with modified antigenic and immunogenic properties, considerations that should be considered when selecting expression systems for glycosylated antigens to be used for structure-function studies and for vaccine use.

Three dimensional picture of the local structure of 1,4-Polybutadiene from a complete atomistic model and neutron scattering data

An efficient method of combining neutron diffraction data over an extended Q range with detailed atomistic models is presented. A quantitative and qualitative mapping of the organization of the chain conformation in both glass and liquid phase has been performed. The proposed structural refinement method is based on the exploitation of the intrachain features of the diffraction pattern by the use of internal coordinates for bond lengths, valence angles and torsion rotations. Models are built stochastically by assignment of these internal coordinates from probability distributions with limited variable parameters. Variation of these parameters is used in the construction of models that minimize the differences between the observed and calculated structure factors. A series of neutron scattering data of 1,4-polybutadiene at the region 20320 K is presented. Analysis of the experimental data yield bond lengths for C-C and C=C of 1.54 and 1.35 Å respectively. Valence angles of the backbone were found to be at 112 and 122.8 for the CCC and CC=C respectively. Three torsion angles corresponding to the double bond and the adjacent R and β bonds were found to occupy cis and trans, s(, trans and g( and trans states, respectively. We compare our results with theoretical predictions, computer simulations, RIS models, and previously reported experimental results.

Barefoot in the Head

A co-edited book (Article Press, UCE, 2011), and book chapter written in response to the event: 'Barefoot in the Head', co-curated with Alun Rowlands and Mark Beasley at Bruce High Quality Foundation University, 225 West Broadway, as part of Performa 09 New York, 12 November 2009 Also including authored chapter: Russell, J. ‘A largely intolerable combination of two mainly unconnected texts: 1. Description of the Barefoot in the Head event, at BHQU, NY, 12/11/09; 2. Fictioning and the End.’ pp 68-85.

Processing of regular and irregular past tense morphology in highly proficient second language learners of English: a self-paced reading study

Dual-system models suggest that English past tense morphology involves two processing routes: rule application for regular verbs and memory retrieval for irregular verbs (Pinker, 1999). In second language (L2) processing research, Ullman (2001a) suggested that both verb types are retrieved from memory, but more recently Clahsen and Felser (2006) and Ullman (2004) argued that past tense rule application can be automatised with experience by L2 learners. To address this controversy, we tested highly proficient Greek-English learners with naturalistic or classroom L2 exposure compared to native English speakers in a self-paced reading task involving past tense forms embedded in plausible sentences. Our results suggest that, irrespective to the type of exposure, proficient L2 learners of extended L2 exposure apply rule-based processing.

Phosphorus flux from wetland ditch sediments

The accumulation of phosphorus (P) in the bottom sediment of field drainage ditches poses a threat to the ecology both of the ditch water and downstream water courses. We investigated the amounts, forms and internal loading of sediment-bound P along two drainage ditches that regulate water levels in a basin fen (~ 200 ha) supporting a mixture of restored wetland and drained agricultural fields. Water levels in the Lady's Drove Rhyne are currently managed to enhance the biodiversity of the wetland (Catcott Lows Reserve — an area formerly cultivated for arable crop production); whereas, the East Ditch is managed to drain adjoining land that remains under arable and livestock production. Laboratory-based chemical fractionation schemes were used to characterise the forms and potential mobility of the sediment-bound P, whilst pore-water equilibrators were employed in situ to evaluate the diffusive flux of P through the sediment–water column, and to characterise the corresponding redox conditions. Along both ditches, sediment pore-water profiles indicated conditions ranging from weakly to very reducing conditions with increasing depth, and net fluxes of P from the sediment to overlying water. P flux values ranged from 0.33 to 1.30 mg m− 2 day− 1. Both the degree of P saturation (DPS) of the sediment and NaOH extractable (Fe/Al-bound) P correlated significantly (P < 0.05) with P flux. Both in the wetland and agricultural ditches, by far the highest values for P flux were recorded at sites closest to points of drainage water entry from the corresponding, adjoining land. Although the P flux data were obtained from only a single sampling event, this study highlights the contribution of historical as well as ongoing agricultural land use on the sustained elevated P status of ditch sediments in lowland catchments.

Stress field control of eruption dynamics at a rift volcano: Nyamuragira, D.R.Congo

Using the record of 30 flank eruptions over the last 110 years at Nyamuragira, we have tested the relationship between the eruption dynamics and the local stress field. There are two groups of eruptions based on their duration (< 80days >) that are also clustered in space and time. We find that the eruptions fed by dykes parallel to the East African Rift Valley have longer durations (and larger volumes) than those eruptions fed by dykes with other orientations. This is compatible with a model for compressible magma transported through an elastic-walled dyke in a differential stress field from an over-pressured reservoir (Woods et al., 2006). The observed pattern of eruptive fissures is consistent with a local stress field modified by a northwest-trending, right lateral slip fault that is part of the northern transfer zone of the Kivu Basin rift segment. We have also re-tested with new data the stochastic eruption models for Nyamuragira of Burt et al. (1994). The time-predictable, pressure-threshold model remains the best fit and is consistent with the typically observed declining rate of sulphur dioxide emission during the first few days of eruption with lava emission from a depressurising, closed, crustal reservoir. The 2.4-fold increase in long-term eruption rate that occurred after 1977 is confirmed in the new analysis. Since that change, the record has been dominated by short-duration eruptions fed by dykes perpendicular to the Rift. We suggest that the intrusion of a major dyke during the 1977 volcano-tectonic event at neighbouring Nyiragongo volcano inhibited subsequent dyke formation on the southern flanks of Nyamuragira and this may also have resulted in more dykes reaching the surface elsewhere. Thus that sudden change in output was a result of a changed stress field that forced more of the deep magma supply to the surface. Another volcano-tectonic event in 2002 may also have changed the magma output rate at Nyamuragira.

A carbamazepine-indomethacin (1:1) cocrystal produced by milling

An X-ray amorphous mixture of carbamazepine and indomethacin transforms upon annealing to produce a novel 1:1 cocrystal, whose structure has been determined from laboratory powder X-ray diffraction (PXRD) data.