Swift thermal reaction norm evolution in a key coccolithopore species: Reaction norm assay data from an experiment in Kiel from December 2012 to June 2015

Temperature has a profound effect on the species composition and physiology of marine phytoplankton, a polyphyletic group of microbes responsible for half of global primary production. Here, we ask whether and how thermal reaction norms in a key calcifying species, the coccolithophore Emiliania huxleyi, change as a result of 2.5 years of experimental evolution to a temperature about 2��C below its upper thermal limit. Replicate experimental populations derived from a single genotype isolated from Norwegian coastal waters were grown at two temperatures for 2.5 years before assessing thermal responses at 6 temperatures ranging from 15 to 26��C, with pCO2 (400/1100/2200 ?atm) as a fully factorial additional factor. The two selection temperatures (15��/26.3��C) led to a marked divergence of thermal reaction norms. Optimal growth temperatures were 0.7��C higher in experimental populations selected at 26.3��C than those selected at 15.0��C. An additional negative effect of high pCO2 on maximal growth rate (8% decrease relative to lowest level) was observed. Finally, the maximum persistence temperature (Tmax) differed by 1-3��C between experimental treatments, as a result of an interaction between pCO2 and the temperature selection. Taken together, we demonstrate that several attributes of thermal reaction norms in phytoplankton may change faster than the predicted progression of ocean warming.

Application of Ultraintense Lasers to validate materials for laser fusion: production of ions and other relevant species

Justification of the need and demand of experimental facilities to test and validate materials for first wall in laser fusion reactors - Characteristics of the laser fusion products - Current ?possible? facilities for tests Ultraintense Lasers as ?complete? solution facility - Generation of ion pulses - Generation of X-ray pulses - Generation of other relevant particles (electrons, neutrons..)

Plasma���wall interaction in laser inertial fusion reactors: novel proposals for radiation tests of first wall materials

Dry-wall laser inertial fusion (LIF) chambers will have to withstand strong bursts of fast charged particles which will deposit tens of kJ m���2 and implant more than 1018 particles m���2 in a few microseconds at a repetition rate of some Hz. Large chamber dimensions and resistant plasma-facing materials must be combined to guarantee the chamber performance as long as possible under the expected threats: heating, fatigue, cracking, formation of defects, retention of light species, swelling and erosion. Current and novel radiation resistant materials for the first wall need to be validated under realistic conditions. However, at present there is a lack of facilities which can reproduce such ion environments. This contribution proposes the use of ultra-intense lasers and high-intense pulsed ion beams (HIPIB) to recreate the plasma conditions in LIF reactors. By target normal sheath acceleration, ultra-intense lasers can generate very short and energetic ion pulses with a spectral distribution similar to that of the inertial fusion ion bursts, suitable to validate fusion materials and to investigate the barely known propagation of those bursts through background plasmas/gases present in the reactor chamber. HIPIB technologies, initially developed for inertial fusion driver systems, provide huge intensity pulses which meet the irradiation conditions expected in the first wall of LIF chambers and thus can be used for the validation of materials too.

Plasma���wall interaction in laser inertial fusion reactors: novel proposals for radiation tests of first wall materials

Dry-wall laser inertial fusion (LIF) chambers will have to withstand strong bursts of fast charged particles which will deposit tens of kJ m���2 and implant more than 1018 particles m���2 in a few microseconds at a repetition rate of some Hz. Large chamber dimensions and resistant plasma-facing materials must be combined to guarantee the chamber performance as long as possible under the expected threats: heating, fatigue, cracking, formation of defects, retention of light species, swelling and erosion. Current and novel radiation resistant materials for the first wall need to be validated under realistic conditions. However, at present there is a lack of facilities which can reproduce such ion environments. This contribution proposes the use of ultra-intense lasers and high-intense pulsed ion beams (HIPIB) to recreate the plasma conditions in LIF reactors. By target normal sheath acceleration, ultra-intense lasers can generate very short and energetic ion pulses with a spectral distribution similar to that of the inertial fusion ion bursts, suitable to validate fusion materials and to investigate the barely known propagation of those bursts through background plasmas/gases present in the reactor chamber. HIPIB technologies, initially developed for inertial fusion driver systems, provide huge intensity pulses which meet the irradiation conditions expected in the first wall of LIF chambers and thus can be used for the validation of materials too.

Transcriptional changes and oxidative stress associated with the synergistic interaction between potato virus X and potato virus Y and their relationship with symptom expression.

Many virus diseases of economic importance to agriculture result from mixtures of different pathogens invading the host at a given time. This contrasts with the relatively scarce studies available on the molecular events associated with virus---host interactions in mixed infections. Compared with single infections, co-infection of Nicotiana benthamiana with Potato virus X (PVX) and Potato virus Y (PVY) resulted in increased systemic symptoms (synergism) that led to necrosis of the newly emerging leaves and death of the plant. A comparative transcriptional analysis was undertaken to identify quantitative and qualitative differences in gene expression during this synergistic infection and correlate these changes with the severe symptoms it caused. Global transcription profiles of doubly infected leaves were compared with those from singly infected leaves using gene ontology enrichment analysis and metabolic pathway annotator software. Functional gene categories altered by the double infection comprise suites of genes regulated coordinately, which are associated with chloroplast functions (downregulated), protein synthesis and degradation (upregulated), carbohydrate metabolism (upregulated), and response to biotic stimulus and stress (upregulated). The expressions of reactive oxygen species?generating enzymes as well as several mitogen-activated protein kinases were also significantly induced. Accordingly, synergistic infection induced a severe oxidative stress in N. benthamiana leaves, as judged by increases in lipid peroxidation and by the generation of superoxide radicals in chloroplasts, which correlated with the misregulation of antioxidative genes in microarray data. Interestingly, expression of genes encoding oxylipin biosynthesis was uniquely upregulated by the synergistic infection. Virus-induced gene silencing of ?-dioxygenase1 delayed cell death during PVX?PVY infection.

New insights into the regulation of NADPH oxidase dependent reactive oxygen species signaling during the plant immune response

Las NADPH oxidasas de plantas, denominadas ���respiratory burst oxidase homologues��� (RBOHs), producen especies reactivas del ox��geno (ROS) que median un amplio rango de funciones. En la c��lula vegetal, el ajuste preciso de la producci��n de ROS aporta la especificidad de se��al para generar una respuesta apropiada ante las amenazas ambientales. RbohD y RbohF, dos de los diez genes Rboh de Arabidopsis, son pleiotr��picos y median diversos procesos fisiol��gicos en respuesta a pat��genos. El control espacio-temporal de la expresi��n de los genes RbohD y RbohF podr��a ser un aspecto cr��tico para determinar la multiplicidad de funciones de estas oxidasas. Por ello, generamos l��neas transg��nicas de Arabidopsis con fusiones de los promoters de RbohD y RbohF a los genes delatores de la B-glucuronidasa y la luciferasa. Estas l��neas fueron empleadas para revelar el patr��n de expresi��n diferencial de RbohD y RbohF durante la respuesta inmune de Arabidopsis a la bacteria pat��gena Pseudomonas syringae pv. tomato DC3000, el hongo necr��trofo Plectosphaerella cucumerina y en respuesta a se��ales relacionadas con la respuesta inmune. Nuestros experimentos revelan un patr��n de expresi��n diferencial de los promotores de RbohD y RbohF durante el desarrollo de la planta y en la respuesta inmune de Arabidopsis. Adem��s hemos puesto de manifiesto que existe una correlaci��n entre el nivel de actividad de los promotores de RbohD y RbohF con la acumulaci��n de ROS y el nivel de muerte celular en respuesta a pat��genos. La expression de RbohD y RbohF tambi��n es modulada de manera diferencial en respuesta a patrones moleculares asociados a pat��genos (PAMPs) y por ��cido absc��sico (ABA). Cabe destacar que, mediante una estrategia de intercambio de promotores, hemos revelado que la regi��n promotora de RbohD, es necesaria para dirigir la producci��n de ROS en respuesta a P. cucumerina. Adicionalmente, la activaci��n del promotor de RbohD en respuesta al aislado de P. cucumerina no adaptado a Arabidopsis 2127, nos llev�� a realizar ensayos de susceptibilidad con el doble mutante rbohD rbohF que han revelado un papel desconocido de estas oxidasas en resistencia no-huesped. La interacci��n entre la se��alizaci��n dependiente de las RBOHs y otros componentes de la respuesta inmune de plantas podr��a explicar tambi��n las distintas funciones que median estas oxidasas en relaci��n con la respuesta inmune. Entre la gran cantidad de se��ales coordinadas con la actividad de las RBOHs, existen evidencias gen��ticas y farmacol��gicas que indican que las prote��nas G heterotrim��ricas est��n implicadas en algunas de las rutas de se��alizaci��n mediadas por ROS derivadas de los RBOHs en respuesta a se��ales ambientales. Por ello hemos estudiado la relaci��n entre estas RBOH-NADPH oxidasas y AGB1, la subunidad �� de las prote��nas G heterotrim��ricas en la respuesta inmune de Arabidopsis. An��lisis de epistasis indican que las prote��nas G heterotrim��ricas est��n implicadas en distintas rutas de se��alizaci��n en defensa mediadas por las RBOHs. Nuestros resultados ilustran la relaci��n compleja entre la se��alizaci��n mediada por las RBOHs y las prote��nas G heterotrim��ricas, que var��a en funci��n de la interacci��n planta-pat��geno analizada. Adem��s, hemos explorado la posible asociaci��n entre AGB1 con RBOHD y RBOHF en eventos tempranos de la respuesta immune. Cabe se��alar que experimentos de co��mmunoprecipitaci��n apuntan a una posible asociaci��n entre AGB1 y la kinasa citoplasm��tica reguladora de RBOHD, BIK1. Esto indica un posible mecanismo de control de la funci��n de esta NADPH oxidase por AGB1. En conjunto, estos datos aportan nuevas perspectivas sobre c��mo, a trav��s del control transcripcional o mediante la interacci��n con las prote��nas G heterotrim��ricas, las NADPH oxidases de plantas median la producci��n de ROS y la se��alizaci��n por ROS en la respuesta inmune. Nuestro trabajo ejemplifica c��mo la regulaci��n diferencial de dos miembros de una familia multig��nica, les permite realizar distintas funciones fisiol��gicas especializadas usando un mismo mecanismo enzim��tico. ABSTRACT The plant NADPH oxidases, termed respiratory burst oxidase homologues (RBOHs), produce reactive oxygen species (ROS) which mediate a wide range of functions. Fine tuning this ROS production provides the signaling specificity to the plant cell to produce the appropriate response to environmental threats. RbohD and RbohF, two of the ten Rboh genes present in Arabidopsis, are pleiotropic and mediate diverse physiological processes in response to pathogens. One aspect that may prove critical to determine the multiplicity of functions of RbohD and RbohF is the spatio-temporal control of their gene expression. Thus, we generated Arabidopsis transgenic lines with RbohD- and RbohF-promoter fusions to the ��-glucuronidase and the luciferase reporter genes. These transgenics were employed to reveal RbohD and RbohF promoter activity during Arabidopsis immune response to the pathogenic bacterium Pseudomonas syringae pv tomato DC3000, the necrotrophic fungus Plectosphaerella cucumerina and in response to immunity-related cues. Our experiments revealed a differential expression pattern of RbohD and RbohF throughout plant development and during Arabidopsis immune response. Moreover, we observed a correlation between the level of RbohD and RbohF promoter activity, the accumulation of ROS and the amount of cell death in response to pathogens. RbohD and RbohF gene expression was also differentially modulated by pathogen associated molecular patterns and abscisic acid. Interestingly, a promoter-swap strategy revealed the requirement for the promoter region of RbohD to drive the production of ROS in response to P. cucumerina. Additionally, since the RbohD promoter was activated during Arabidopsis interaction with a non-adapted P. cucumerina isolate 2127, we performed susceptibility tests to this fungal isolate that uncovered a new role of these oxidases on non-host resistance. The interplay between RBOH-dependent signaling with other components of the plant immune response might also explain the different immunity-related functions mediated by these oxidases. Among the plethora of signals coordinated with RBOH activity, pharmacological and genetic evidence indicates that heterotrimeric G proteins are involved in some of the signaling pathways mediated by RBOH���derived ROS in response to environmental cues. Therefore, we analysed the interplay between these RBOH-NADPH oxidases and AGB1, the Arabidopsis ��-subunit of heterotrimeric G proteins during Arabidopsis immune response. We carried out epistasis studies that allowed us to test the implication of AGB1 in different RBOH-mediated defense signaling pathways. Our results illustrate the complex relationship between RBOH and heterotrimeric G proteins signaling, that varies depending on the type of plant-pathogen interaction. Furthermore, we tested the potential association between AGB1 with RBOHD and RBOHF during early immunity. Interestingly, our co-immunoprecipitation experiments point towards an association of AGB1 and the RBOHD regulatory kinase BIK1, thus providing a putative mechanism in the control of the NADPH oxidase function by AGB1. Taken all together, these studies provide further insights into the role that transcriptional control or the interaction with heterotrimeric G-proteins have on RBOH-NADPH oxidase-dependent ROS production and signaling in immunity. Our work exemplifies how, through a differential regulation, two members of a multigenic family achieve specialized physiological functions using a common enzymatic mechanism.

Conversion of cucumber linoleate 13-lipoxygenase to a 9-lipoxygenating species by site-directed mutagenesis

Multiple lipoxygenase sequence alignments and structural modeling of the enzyme/substrate interaction of the cucumber lipid body lipoxygenase suggested histidine 608 as the primary determinant of positional specificity. Replacement of this amino acid by a less-space-filling valine altered the positional specificity of this linoleate 13-lipoxygenase in favor of 9-lipoxygenation. These alterations may be explained by the fact that H608V mutation may demask the positively charged guanidino group of R758, which, in turn, may force an inverse head-to-tail orientation of the fatty acid substrate. The R758L+H608V double mutant exhibited a strongly reduced reaction rate and a random positional specificity. Trilinolein, which lacks free carboxylic groups, was oxygenated to the corresponding (13S)-hydro(pero)xy derivatives by both the wild-type enzyme and the linoleate 9-lipoxygenating H608V mutant. These data indicate the complete conversion of a linoleate 13-lipoxygenase to a 9-lipoxygenating species by a single point mutation. It is hypothesized that H608V exchange may alter the orientation of the substrate at the active site and/or its steric configuration in such a way that a stereospecific dioxygen insertion at C-9 may exclusively take place.

Aggregate community properties and the strength of species��� interactions

Although humanity depends on the continued, aggregate functioning of natural ecosystems, few studies have explored the impact of community structure on the stability of aggregate community properties. Here we derive the stability of the aggregate property of community biomass as a function of species��� competition coefficients for a two-species model. The model predicts that the stability of community biomass is relatively independent of the magnitude of the interaction strengths. Instead, the degree of asymmetry of the interactions appears to be key to community stability.

Variation in per capita interaction strength: Thresholds due to nonlinear dynamics and nonequilibrium conditions

I measured the strength of interaction between a marine herbivore and its growing resource over a realistic range of absolute and relative abundances. The herbivores (hermit crabs: Pagurus spp.) have slow and/or weak functional and numerical responses to epiphytic diatoms (Isthmia nervosa), which show logistic growth in the absence of consumers. By isolating this interaction in containers in the field, I mimicked many of the physical and biological variables characteristic of the intertidal while controlling the densities of focal species. The per capita effects of consumers on the population dynamics of their resource (i.e., interaction strength) were defined by using the relationship between hermit crab density and proportional change in the resource. When this relationship is fit by a Weibull function, a single parameter distinguishes constant interaction strength from one that varies as a function of density. Constant interaction strength causes the proportion of diatoms to fall linearly or proportionally as hermit crab density increases whereas per capita effects that increase with density cause an accelerating decline. Although many mathematical models of species interactions assume linear dynamics and invariant parameters, at least near equilibrium, the per capita effects of hermit crabs on diatoms varied substantially, apparently crossing a threshold from weak to strong when consumption exceeded resource production. This threshold separates a domain of coexistence from one of local extinction of the resource. Such thresholds may help explain trophic cascades, resource compensation, and context-dependent interaction strengths, while indicating a way to predict trophic effects, despite nonlinearities, as a function of vital rates.

Primate species richness is determined by plant productivity: Implications for���conservation

The explanation of patterns in species richness ranks among the most important tasks of ecology. Current theories emphasize the interaction between historical and geographical factors affecting the size of the regional species pool and of locally acting processes such as competitive exclusion, disturbance, productivity, and seasonality. Local species richness, or alpha diversity, of plants and primary consumers has been claimed to peak in habitats of low and intermediate productivity, which, if true, has major implications for conservation. Here, by contrast, we show that local richness of Neotropical primates (platyrrhines) is influenced by both historical biogeography and productivity but not by tree species richness or seasonality. This pattern indicates that habitats with the highest plant productivity are also the richest for many important primary consumers. We show further that fragmentation of Amazonian rain forests in the Pleistocene, if it occurred, appears to have had a negligible influence on primate alpha species richness.

Molecular Interaction Map of the Mammalian Cell Cycle Control and DNA Repair Systems

Eventually to understand the integrated function of the cell cycle regulatory network, we must organize the known interactions in the form of a diagram, map, and/or database. A diagram convention was designed capable of unambiguous representation of networks containing multiprotein complexes, protein modifications, and enzymes that are substrates of other enzymes. To facilitate linkage to a database, each molecular species is symbolically represented only once in each diagram. Molecular species can be located on the map by means of indexed grid coordinates. Each interaction is referenced to an annotation list where pertinent information and references can be found. Parts of the network are grouped into functional subsystems. The map shows how multiprotein complexes could assemble and function at gene promoter sites and at sites of DNA damage. It also portrays the richness of connections between the p53-Mdm2 subsystem and other parts of the network.

Direct interaction of flagellin termini essential for polymorphic ability of flagellar���filament

We report the structures of flagellar filaments reconstituted from various flagellins with small terminal truncations. Flagellins from Salmonella typhimurium strains SJW1103 (wild type), SJW1660, and SJW1655 were used, which form a left-handed supercoil, the L- and R-type straight forms, respectively. Structure analyses were done by electron cryomicroscopy and helical image reconstruction with a help of x-ray fiber diffraction for determining precise helical symmetries. Truncation of either terminal region, irrespective of the original flagellin species, results in a straight filament having a helical symmetry distinct either from the L- or R-type. This filament structure is named Lt-type. Although the local subunit packing is similar in all three types, a close comparison shows that the Lt-type packing is almost identical to the R-type but distinct from the L-type, which demonstrates the strong two-state preference of the subunit interactions. The structure clearly suggests that both termini are located in the inner tube of the concentric double-tubular structure of the filament core, and their proper interaction is responsible for the correct folding of fairly large terminal regions that form the inner tube. The double tubular structure appears to be essential for the polymorphic ability of flagellar filaments, which is required for the swimming���tumbling of bacterial taxis.

Interaction between inducible nitric oxide synthase and cyclooxygenase-2 after cerebral ischemia

Focal cerebral ischemia is associated with expression of both inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), enzymes whose reaction products contribute to the evolution of ischemic brain injury. We tested the hypothesis that, after cerebral ischemia, nitric oxide (NO) produced by iNOS enhances COX-2 activity, thereby increasing the toxic potential of this enzyme. Cerebral ischemia was produced by middle cerebral artery occlusion in rats or mice. Twenty-four hours after ischemia in rats, iNOS-immunoreactive neutrophils were observed in close proximity (<20 ��m) to COX-2-positive cells at the periphery of the infarct. In the olfactory bulb, only COX-2 positive cells were observed. Cerebral ischemia increased the concentration of the COX-2 reaction product prostaglandin E2 (PGE2) in the ischemic area and in the ipsilateral olfactory bulb. The iNOS inhibitor aminoguanidine reduced PGE2 concentration in the infarct, where both iNOS and COX-2 were expressed, but not in the olfactory bulb, where only COX-2 was expressed. Postischemic PGE2 accumulation was reduced significantly in iNOS null mice compared with wild-type controls (C57BL/6 or SV129). The data provide evidence that NO produced by iNOS influences COX-2 activity after focal cerebral ischemia. Pro-inflammatory prostanoids and reactive oxygen species produced by COX-2 may be a previously unrecognized factor by which NO contributes to ischemic brain injury. The pathogenic effect of the interaction between NO, or a derived specie, and COX-2 is likely to play a role also in other brain diseases associated with inflammation.

Analysis of the Xenopus laevis CCAAT-enhancer binding protein �� gene promoter demonstrates species-specific differences in the mechanisms for both auto-activation and regulation by Sp1

Transcription factors belonging to the CCAAT-enhancer binding protein (C/EBP) family have been implicated in the regulation of gene expression during differentiation, development and disease. Autoregulation is relatively common in the modulation of C/EBP gene expression and the murine and human C/EBP�� genes have been shown to be auto-activated by different mechanisms. In the light of this finding, it is essential that autoregulation of C/EBP�� genes from a wider range of different species be investigated in order to gauge the degree of commonality, or otherwise, that may exist. We report here studies that investigate the regulation of the Xenopus laevis C/EBP�� gene (xC/EBP��). The ���1131/+41 promoter region was capable of directing high levels of expression in both the human hepatoma Hep3B and the Xenopus kidney epithelial A6 cell lines, and was auto-activated by expression vectors specifying for xC/EBP�� or xC/EBP��. Deletion analysis showed that the ���321/+41 sequence was sufficient for both the constitutive promoter activity and auto-activation and electrophoretic mobility shift assays identified the interaction of C/EBPs and Sp1 to this region. Although deletion of either the C/EBP or the Sp1 site drastically reduced the xC/EBP�� promoter activity, multimers of only the C/EBP site could confer autoregulation to a heterologous SV40 promoter. These results indicate that, in contrast to the human promoter and in common with the murine gene, the xC/EBP�� promoter was subject to direct autoregulation. In addition, we demonstrate a novel species-specific action of Sp1 in the regulation of C/EBP�� expression, with the factor able to repress the murine promoter but activate the Xenopus gene.

A Xenopus distal-less gene in transgenic mice: conserved regulation in distal limb epidermis and other sites of epithelial-mesenchymal interaction.

In this paper, we show the conserved regulation of the homeodomain gene Distal-less-3 (Dlx-3) by analyzing the expression of a promoter from the Xenopus ortholog, Xdll-2, in transgenic mice. A 470-bp frog regulatory sequence confers appropriate expression on a lacZ reporter gene in the ectodermal component of structures derived from epithelial-mesenchymal interactions. Remarkably, this includes structures absent in Xenopus, such as the hair follicle and mammary gland, suggesting that conserved regulatory elements can be used to control the formation of structures peculiar to individual species. In addition, expression of Dlx-3 in developing limbs is highest at the most distal portion. This pattern is duplicated by the Xenopus promoter, indicating that this DNA may include sequences responsive to conserved proximodistal patterning signals in the vertebrate limb.