Evolution of alternative biosynthetic pathways for vitamin C following plastid acquisition in photosynthetic eukaryotes.

Ascorbic acid (vitamin C) is an enzyme co-factor in eukaryotes that also plays a critical role in protecting photosynthetic eukaryotes against damaging reactive oxygen species derived from the chloroplast. Many animal lineages, including primates, have become ascorbate auxotrophs due to the loss of the terminal enzyme in their biosynthetic pathway, L-gulonolactone oxidase (GULO). The alternative pathways found in land plants and Euglena use a different terminal enzyme, L-galactonolactone dehydrogenase (GLDH). The evolutionary processes leading to these differing pathways and their contribution to the cellular roles of ascorbate remain unclear. Here we present molecular and biochemical evidence demonstrating that GULO was functionally replaced with GLDH in photosynthetic eukaryote lineages following plastid acquisition. GULO has therefore been lost repeatedly throughout eukaryote evolution. The formation of the alternative biosynthetic pathways in photosynthetic eukaryotes uncoupled ascorbate synthesis from hydrogen peroxide production and likely contributed to the rise of ascorbate as a major photoprotective antioxidant.

Critical Role for STAT4 Activation by Type 1 Interferons in the Interferon-Y Response to Viral Infection

Interferons (IFNs) are essential for host defense. Although the antiviral effects of the type 1 IFNs IFN- and IFN- (IFN-/) have been established, their immunoregulatory functions, especially their ability to regulate IFN- production, are poorly understood. Here we show that IFN-/ activate STAT4 directly (STAT, signal transducers and activators of transcription) and that this is required for IFN- production during viral infections of mice, in concert with T cell receptor-derived signals. In contrast, STAT1 appears to negatively regulate IFN-/ induction of IFN-. Thus, type 1 IFNs, in addition to interleukin-12, provide pathways for innate regulation of adaptive immunity, and their immunoregulatory functions are controlled by modulating the activity of individual STATs.

Muscle-specific variations in use-dependent crossed-facilitation of corticospinal pathways mediated by transcranial direct current (DC) stimulation

The tendency for contractions of muscles in the upper limb to give rise to increases in the excitability of corticospinal projections to the homologous muscles of the opposite limb is well known. Although the suppression of this tendency is integral to tasks of daily living, its exploitation may prove to be critical in the rehabilitation of acquired hemiplegias. Transcranial direct current (DC) stimulation induces changes in cortical excitability that outlast the period of application. We present evidence that changes in the reactivity of the corticospinal pathway induced by DC stimulation of the motor cortex interact systematically with those brought about by contraction of the muscles of the ipsilateral limb. During the application of flexion torques (up to 50% of maximum) applied at the left wrist, motor evoked potentials (MEPs) were evoked in the quiescent muscles of the right arm by magnetic stimulation of the left motor cortex (M1). The MEPs were obtained prior to and following 10 min of anodal, cathodal or sham DC stimulation of left M1. Cathodal stimulation counteracted increases in the crossed-facilitation of projections to the (right) wrist flexors that otherwise occurred as a result of repeated flexion contractions at the left wrist. In addition, cathodal stimulation markedly decreased the excitability of corticospinal projections to the wrist extensors of the right limb. Thus changes in corticospinal excitability induced by DC stimulation can be shaped (i.e. differentiated by muscle group) by focal contractions of muscles in the limb ipsilateral to the site of stimulation. (C) 2008 Elsevier Ireland Ltd. All rights reserved.

Pathways to state-selective control of vibrational wavepackets in the deuterium molecular ion

The capability of intense ultrashort laser pulses to initiate, control and image vibrational wavepacket dynamics in the deuterium molecular ion has been simulated with a view to inform and direct future femtosecond pump-control-probe experiments. The intense-field coherent control of the vibrational superposition has been studied as a function of pulse intensity and delay time, to provide an indication of key constraints for experimental studies. For selected cases of the control mechanism, probing of the subsequent vibrational wavepacket dynamics has been simulated via the photodissociation (PD) channel. Such PD probing is shown to elucidate the modified wavepacket dynamics where the position of the quantum revival is sensitive to the control process. Through Fourier transform analysis the PD yield is also shown to provide a characterisation of the vibrational distribution. It has been shown that a simple 'critical R cut-off' approximation can be used to reproduce the effect of a probe pulse interaction, providing a convenient and efficient alternative to intensive computer simulations of the PD mechanism in the deuterium molecular ion.

Pathways for delivery of surface water nutrients to receptors

Diffuse contaminants can make their way into rivers via a number of different pathways, including overland flow, interflow, and shallow and deep groundwater. Identification of the key pathway(s) delivering contaminants to a receptor is important for implementing effective water management strategies. The ‘Pathways Project’, funded by the Irish Environmental Protection Agency, is developing a catchment management tool that will enable practitioners to identify the critical source areas for diffuse contaminants, and the key pathways of interest in assessing contaminant problems on a catchment and sub-catchment scale.
One of the aims of the project is to quantify the flow and contaminant loadings being delivered to the stream via each of the main pathways. Chemical separation of stream event hydrographs is being used to supplement more traditional physical hydrograph separation methods. Distinct, stable chemical signatures are derived for each of the pathway end members, and the proportion of flow from each during a rainfall event can be determined using a simple mass balance approach.
Event sampling was carried out in a test catchment underlain by poorly permeable soils and bedrock, which is predominantly used for grazing with a number of one-off rural residential houses. Results show that artificial field drainage, which includes subterranean land drains and collector drains around the perimeters of the 1 to 10 ha fields, plays an important role in the delivery of flow and nutrients to the streams in these types of hydrogeological settings.
Nitrate infiltrates with recharge and is delivered to the stream primarily via the artificial drains and the shallow groundwater pathway. Longitudinal stream profiles show that the nitrate load input is relatively uniform over the 8 km length of the stream at high flows, suggesting widespread diffuse contaminant input. In contrast, phosphorus is adsorbed in the clay-rich soil and is transported mainly via the overland flow pathway and the artificial drains. Longitudinal stream profiles for phosphorus suggest a pattern of more discrete points of phosphorus inputs, which may be related to point sources of contamination.
These techniques have application elsewhere within a toolkit of methods for determining the key pathways delivering contaminants to surface water receptors.

Developing an integrated hydrograph separation and lumped modelling approach to quantifying hydrological pathways in Irish river catchments

An appreciation of the quantity of streamflow derived from the main hydrological pathways involved in transporting diffuse contaminants is critical when addressing a wide range of water resource management issues. In order to assess hydrological pathway contributions to streams, it is necessary to provide feasible upper and lower bounds for flows in each pathway. An important first step in this process is to provide reliable estimates of the slower responding groundwater pathways and subsequently the quicker overland and interflow pathways. This paper investigates the effectiveness of a multi-faceted approach applying different hydrograph separation techniques, supplemented by lumped hydrological modelling, for calculating the Baseflow Index (BFI), for the development of an integrated approach to hydrograph separation. A semi-distributed, lumped and deterministic rainfall runoff model known as NAM has been applied to ten catchments (ranging from 5 to 699 km2). While this modelling approach is useful as a validation method, NAM itself is also an important tool for investigation. These separation techniques provide a large variation in BFI, a difference of 0.741 predicted for BFI in a catchment with the less reliable fixed and sliding interval methods and local minima turning point methods included. This variation is reduced to 0.167 with these methods omitted. The Boughton and Eckhardt algorithms, while quite subjective in their use, provide quick and easily implemented approaches for obtaining physically realistic hydrograph separations. It is observed that while the different separation techniques give varying BFI values for each of the catchments, a recharge coefficient approach developed in Ireland, when applied in conjunction with the Master recession Curve Tabulation method, predict estimates in agreement with those obtained using the NAM model, and these estimates are also consistent with the study catchments’ geology. These two separation methods, in conjunction with the NAM model, were selected to form an integrated approach to assessing BFI in catchments.

Critical role of water in the direct oxidation of CO and hydrocarbons in diesel exhaust after treatment catalysis

CO and C3H6 oxidation have been carried out in the absence and presence of water over a Pd/Al2O3catalyst. It is clear that water promotes CO and, as a consequence, C3H6oxidation takes place at muchlower temperatures compared with the dry feed. The significant increase in the catalyst’s activity withrespect to CO oxidation is not simply associated with changes in surface concentration as a result ofcompetitive adsorption effects. Utilising18O2as the reactant allows the pathways whereby the oxidationdue to gaseous dioxygen and where the water activates the CO and C3H6to be distinguished. In thepresence of water, the predominant pathway is via water activation with C16O2and C16O18O being themajor species formed and oxidation with dioxygen plays a secondary role. The importance of wateractivation is further supported by the significant decrease in its effect when using D2O versus H2O.

Disruption of SF3B1 results in deregulated expression and splicing of key genes and pathways in myelodysplastic syndrome hematopoietic stem and progenitor cells

The splicing factor SF3B1 is the most commonly mutated gene in the myelodysplastic syndrome (MDS), particularly in patients with refractory anemia with ring sideroblasts (RARS). We investigated the functional effects of SF3B1 disruption in myeloid cell lines: SF3B1 knockdown resulted in growth inhibition, cell cycle arrest and impaired erythroid differentiation and deregulation of many genes and pathways, including cell cycle regulation and RNA processing. MDS is a disorder of the hematopoietic stem cell and we thus studied the transcriptome of CD34 + cells from MDS patients with SF3B1 mutations using RNA sequencing. Genes significantly differentially expressed at the transcript andor exon level in SF3B1 mutant compared with wild-type cases include genes that are involved in MDS pathogenesis (ASXL1 and CBL), iron homeostasis and mitochondrial metabolism (ALAS2, ABCB7 and SLC25A37) and RNA splicingprocessing (PRPF8 and HNRNPD). Many genes regulated by a DNA damage-induced BRCA1-BCLAF1-SF3B1 protein complex showed differential expressionsplicing in SF3B1 mutant cases. This is the first study to determine the target genes of SF3B1 mutation in MDS CD34 + cells. Our data indicate that SF3B1 has a critical role in MDS by affecting the expression and splicing of genes involved in specific cellular processespathways, many of which are relevant to the known RARS pathophysiology, suggesting a causal link.

Divergent androgen regulation of unfolded protein response pathways drives prostate cancer

The unfolded protein response (UPR) is a homeostatic mechanism to maintain endoplasmic reticulum (ER) function. The UPR is activated by various physiological conditions as well as in disease states, such as cancer. As androgens regulate secretion and development of the normal prostate and drive prostate cancer (PCa) growth, they may affect UPR pathways. Here, we show that the canonical UPR pathways are directly and divergently regulated by androgens in PCa cells, through the androgen receptor (AR), which is critical for PCa survival. AR bound to gene regulatory sites and activated the IRE1α branch, but simultaneously inhibited PERK signaling. Inhibition of the IRE1α arm profoundly reduced PCa cell growth in vitro as well as tumor formation in preclinical models of PCa in vivo. Consistently, AR and UPR gene expression were correlated in human PCa, and spliced XBP-1 expression was significantly upregulated in cancer compared with normal prostate. These data establish a genetic switch orchestrated by AR that divergently regulates the UPR pathways and suggest that targeting IRE1α signaling may have therapeutic utility in PCa.

Growth factor activation of ErbB2/ErbB3 signaling pathways regulate the activity of Estrogen Receptors (ER)

La signalisation par l’estrogène a longtemps été considérée comme jouant un rôle critique dans le développement et la progression des cancers hormono-dépendants tel que le cancer du sein. Deux tiers des cancers du sein expriment le récepteur des estrogènes (ER) qui constitue un élément indiscutable dans cette pathologie. L’acquisition d’une résistance endocrinienne est cependant un obstacle majeur au traitement de cette forme de cancer. L’émergence de cancers hormono-indépendants peut est produite par l’activation de ER en absence d’estrogène, l’hypersensibilité du récepteur aux faibles concentrations plasmique d’estrogène ainsi que l’activation de ER par des modulateurs sélectifs. L’activité du ER est fortement influencée par l’environnement cellulaire tel que l’activation de voie de signalisation des facteurs de croissances, la disponibilité de protéines co-régulatrices et des séquences promotrices ciblées. Présentement, les études ont principalement considérées le rôle de ERα, cependant avec la découverte de ERβ, notre compréhension de la diversité des mécanismes potentiels impliquant des réponses ER-dépendantes s’est améliorée. L’activation des voies des kinases par les facteurs de croissance entraîne le développement d’un phénotype tumoral résistant aux traitements actuels. Nos connaissances des voies impliquées dans l’activation de ER sont restreintes. ERα est considéré comme le sous-type dominant et corrèle avec la plupart des facteurs de pronostic dans le cancer du sein. Le rôle de ERβ reste imprécis. Les résultats présentés dans cette thèse ont pour objectif de mieux comprendre l’implication de ERβ dans la prolifération cellulaire par l’étude du comportement de ERβ et ERα suite à l’activation des voies de signalisation par les facteurs de croissance. Nous démontrons que l’activation des récepteurs de surfaces de la famille ErbB, spécifiquement ErbB2/ErbB3, inhibe l’activité transcriptionnelle de ERβ, malgré la présence du coactivateur CBP, tout en activant ERα. De plus, l’inhibition de ERβ est attribuée à un résidu sérine (Ser-255) situé dans la région charnière, absente dans ERα. Des études supplémentaires de ErbB2/ErbB3 ont révélé qu’ils activent la voie PI3K/Akt ciblant à son tour la Ser-255. En effet, cette phosphorylation de ERβ par PI3K/Akt induit une augmentation de l’ubiquitination du récepteur qui promeut sa dégradation par le système ubiquitine-protéasome. Cette dégradation est spécifique pour ERβ. De façon intéressante, la dégradation par le protéasome requiert la présence du coactivateur CBP normalement requis pour l’activité transcriptionnelle des récepteurs nucléaires. Malgré le fait que l’activation de la voie PI3K/Akt corrèle avec une diminution de l’expression des gènes sous le contrôle de ERβ, on observe une augmentation de la prolifération des cellules cancéreuses. L’inhibition de la dégradation de ERβ réduit cette prolifération excessive causée par le traitement avec Hrgβ1, un ligand de ErbB3. Un nombre croissant d’évidences indique que les voies de signalisations des facteurs de croissance peuvent sélectivement réguler l’activité transcriptionnelle de sous-types de ER. De plus, le ratio ERα/ERβ dans les cancers du sein devient un outil de diagnostique populaire afin de déterminer la sévérité d’une tumeur. En conclusion, la caractérisation moléculaire du couplage entre la signalisation des facteurs de croissance et la fonction des ERs permettra le développement de nouveaux traitements afin de limiter l’apparition de cellules tumorales résistantes aux thérapies endocriniennes actuelles.

Early growth response protein 1 (Egr-1) expression by Insulin-like growth factor 1 (IGF-1) involves MAPKs and PKB pathways

Un remodelage vasculaire anormal est à la base de la pathogenèse des maladies cardio-vasculaires (MCV) telles que l’athérosclérose et l’hypertension. Des dysfonctionnements au niveau de la migration, l’hypertrophie et la prolifération des cellules musculaires lisses vasculaires (CMLV) sont des évènements cellulaires qui jouent un rôle primordial dans le remodelage vasculaire. L’insulin-like growth factor 1 (IGF-1), puissant facteur mitogène, contribue au développement des MCV, notamment via l’activation des protéines MAPK et PI3-K/PKB, composantes clés impliquées dans les voies de croissance cellulaire. Ces molécules sont également impliquées dans la modulation de l’expression de nombreux facteurs de transcription, incluant le facteur Egr-1. Egr-1 est régulé à la hausse dans différents types de maladies vasculaires impliquant les voies de signalisation de croissance et de stress oxydant qui par ailleurs peuvent être déclenchées par l’IGF-1. Cependant, la question d’une possible modulation de l’expression d’Egr-1 dans les CMLV demeure inabordée; plus spécifiquement, la caractérisation de la voie de signalisation reliant l’action d’IGF-1 à l’expression d’Egr-1 reste à établir. Dans cette optique, l’objectif de cette étude a été d’examiner l’implication de MAPK, PKB et des dérivés réactifs de l’oxygène (DRO) dans l’expression d’Egr-1 induite par l’IGF-1 dans les CMLV. L’IGF-1 a induit une augmentation marquée du niveau protéique de l’Egr-1 en fonction du temps et de la concentration utilisés. Cette augmentation a été inhibée en fonction des doses d’agents pharmacologiques qui ciblent les voies de signalisation de MAPK, PKB et DRO. De plus, l’expression du facteur de transcription, Egr-1, en réponse de l’IGF-1, a été atténuée suite à un blocage pharmacologique des processus cellulaires responsables de la synthèse d’ARN et de synthèse protéique. Pour conclure, on a démontré que l’IGF-1 stimule l’expression d’Egr-1 via les voies de signalisation, impliquant ERK1/2/JNK, PI3K/PKB. On a également proposé que les DRO jouent un rôle important dans ce processus. Dans l’ensemble, nous avons suggéré un nouveau mécanisme par lequel l’IGF-1 promeut la prolifération et l’hypertrophie cellulaire, processus à la base des anomalies vasculaires.

Analysis of water-level response to rainfall and implications for recharge pathways in the Chalk aquifer, SE England

Water table response to rainfall was investigated at six sites in the Upper, Middle and Lower Chalk of southern England. Daily time series of rainfall and borehole water level were cross-corretated to investigate seasonal variations in groundwater-level response times, based on periods of 3-month duration. The time tags (in days) yielding significant correlations were compared with the average unsaturated zone thickness during each 3-month period. In general, for cases when the unsaturated zone was greater than 18 m thick, the time tag for a significant water-level response increased rapidly once the depth to the water table exceeded a critical value, which varied from site to site. For shallower water tables, a linear relationship between the depth to the water table and the water-level response time was evident. The observed variations in response time can only be partially accounted for using a diffusive model for propagation through the unsaturated matrix, suggesting that some fissure flow was occurring. The majority of rapid responses were observed during the winter/spring recharge period, when the unsaturated zone is thinnest and the unsaturated zone moisture content is highest, and were more likely to occur when the rainfall intensity exceeded 5 mm/day. At some sites, a very rapid response within 24 h of rainfall was observed in addition to the longer term responses even when the unsaturated zone was up to 64 m thick. This response was generally associated with the autumn period. The results of the cross-correlation analysis provide statistical support for the presence of fissure flow and for the contribution of multiple pathways through the unsaturated zone to groundwater recharge. (c) 2006 Elsevier B.V. All rights reserved.

Glycogen synthase kinase 3 (GSK3) in the heart: a point of integration in hypertrophic signalling and a therapeutic target? A critical analysis

Glycogen synthase kinase 3 (GSK3, of which there are two isoforms, GSK3alpha and GSK3beta) was originally characterized in the context of regulation of glycogen metabolism, though it is now known to regulate many other cellular processes. Phosphorylation of GSK3alpha(Ser21) and GSK3beta(Ser9) inhibits their activity. In the heart, emphasis has been placed particularly on GSK3beta, rather than GSK3alpha. Importantly, catalytically-active GSK3 generally restrains gene expression and, in the heart, catalytically-active GSK3 has been implicated in anti-hypertrophic signalling. Inhibition of GSK3 results in changes in the activities of transcription and translation factors in the heart and promotes hypertrophic responses, and it is generally assumed that signal transduction from hypertrophic stimuli to GSK3 passes primarily through protein kinase B/Akt (PKB/Akt). However, recent data suggest that the situation is far more complex. We review evidence pertaining to the role of GSK3 in the myocardium and discuss effects of genetic manipulation of GSK3 activity in vivo. We also discuss the signalling pathways potentially regulating GSK3 activity and propose that, depending on the stimulus, phosphorylation of GSK3 is independent of PKB/Akt. Potential GSK3 substrates studied in relation to myocardial hypertrophy include nuclear factors of activated T cells, beta-catenin, GATA4, myocardin, CREB, and eukaryotic initiation factor 2Bvarepsilon. These and other transcription factor substrates putatively important in the heart are considered. We discuss whether cardiac pathologies could be treated by therapeutic intervention at the GSK3 level but conclude that any intervention would be premature without greater understanding of the precise role of GSK3 in cardiac processes.

Oxidised low-density lipoproteins induce rapid platelet activation and shape change through tyrosine kinase and Rho kinase-signaling pathways

Oxidized low-density lipoproteins (oxLDL) generated in the hyperlipidemic state may contribute to unregulated platelet activation during thrombosis. Although the ability of oxLDL to activate platelets is established, the underlying signaling mechanisms remain obscure. Weshow that oxLDL stimulate platelet activation through phosphorylation of the regulatory light chains of the contractile protein myosin IIa (MLC). oxLDL, but not native LDL, induced shape change, spreading, and phosphorylation of MLC (serine 19) through a pathway that was ablated under conditions that blocked CD36 ligation or inhibited Src kinases, suggesting a tyrosine kinase–dependent mechanism. Consistent with this, oxLDL induced tyrosine phosphorylation of a number of proteins including Syk and phospholipase C g2. Inhibition of Syk, Ca21 mobilization, and MLC kinase (MLCK) only partially inhibited MLC phosphorylation, suggesting the presence of a second pathway. oxLDL activated RhoA and RhoA kinase (ROCK) to induce inhibitory phosphorylation of MLC phosphatase (MLCP). Moreover, inhibition of Src kinases prevented the activation of RhoA and ROCK, indicating that oxLDL regulates contractile signaling through a tyrosine kinase–dependent pathway that induces MLC phosphorylation through the dual activation of MLCK and inhibition of MLCP. These data reveal new signaling events downstream of CD36 that are critical in promoting platelet aggregation by oxLDL.

Full effects of land use change in the representative concentration pathways

Future land use change (LUC) is an important component of the IPCC representative concentration pathways (RCPs), but in these scenarios' radiative forcing targets the climate impact of LUC only includes greenhouse gases. However, climate effects due to physical changes of the land surface can be as large. Here we show the critical importance of including non-carbon impacts of LUC when considering the RCPs. Using an ensemble of climate model simulations with and without LUC, we show that the net climate effect is very different from the carbon-only effect. Despite opposite signs of LUC, all the RCPs assessed here have a small net warming from LUC because of varying biogeophysical effects, and in RCP4.5 the warming is outside of the expected variability. The afforestation in RCP4.5 decreases surface albedo, making the net global temperature anomaly over land around five times larger than RCPs 2.6 and 8.5, for around twice the amount of LUC. Consequent changes to circulation in RCP4.5 in turn reduce Arctic sea ice cover. The small net positive temperature effect from LUC could make RCP4.5's universal carbon tax, which incentivizes retaining and growing forest, counter productive with respect to climate. However, there are spatial differences in the balance of impacts, and potential climate gains would need to be assessed against other environmental aims.