The TORNADO1 and TORNADO2 genes function in several patterning processes during early leaf development in Arabidopsis thaliana

In multicellular organisms, patterning is a process that generates axes in the primary body plan, creates domains upon organ formation, and finally leads to differentiation into tissues and cell types. We identified the Arabidopsis thaliana TORNADO1 (TRN1) and TRN2 genes and their role in leaf patterning processes such as lamina venation, symmetry, and lateral growth. In trn mutants, the leaf venation network had a severely reduced complexity: incomplete loops, no tertiary or quaternary veins, and vascular islands. The leaf laminas were asymmetric and narrow because of a severely reduced cell number. We postulate that the imbalance between cell proliferation and cell differentiation and the altered auxin distribution in both trn mutants cause asymmetric leaf growth and aberrant venation patterning. TRN1 and TRN2 were epistatic to ASYMMETRIC LEAVES1 with respect to leaf asymmetry, consistent with their expression in the shoot apical meristem and leaf primordia. TRN1 codes for a large plant-specific protein with conserved domains also found in a variety of signaling proteins, whereas TRN2 encodes a transmembrane protein of the tetraspanin family whose phylogenetic tree is presented. Double mutant analysis showed that TRN1 and TRN2 act in the same pathway.     

Mapping information flow in sensorimotor networks

Biological organisms continuously select and sample information used by their neural structures for perception and action, and for creating coherent cognitive states guiding their autonomous behavior. Information processing, however, is not solely an internal function of the nervous system. Here we show, instead, how sensorimotor interaction and body morphology can induce statistical regularities and information structure in sensory inputs and within the neural control architecture, and how the flow of information between sensors, neural units, and effectors is actively shaped by the interaction with the environment. We analyze sensory and motor data collected from real and simulated robots and reveal the presence of information structure and directed information flow induced by dynamically coupled sensorimotor activity, including effects of motor outputs on sensory inputs. We find that information structure and information flow in sensorimotor networks (a) is spatially and temporally specific; (b) can be affected by learning, and (c) can be affected by changes in body morphology. Our results suggest a fundamental link between physical embeddedness and information, highlighting the effects of embodied interactions on internal (neural) information processing, and illuminating the role of various system components on the generation of behavior.     

Statistically integrated metabonomic-proteomic studies on a human prostate cancer xenograft model in mice

A novel statistically integrated proteometabonomic method has been developed and applied to a human tumor xenograft mouse model of prostate cancer. Parallel 2D-DIGE proteomic and 1H NMR metabolic profile data were collected on blood plasma from mice implanted with a prostate cancer (PC-3) xenograft and from matched control animals. To interpret the xenograft-induced differences in plasma profiles, multivariate statistical algorithms including orthogonal projection to latent structure (OPLS) were applied to generate models characterizing the disease profile. Two approaches to integrating metabonomic data matrices are presented based on OPLS algorithms to provide a framework for generating models relating to the specific and common sources of variation in the metabolite concentrations and protein abundances that can be directly related to the disease model. Multiple correlations between metabolites and proteins were found, including associations between serotransferrin precursor and both tyrosine and 3-D-hydroxybutyrate. Additionally, a correlation between decreased concentration of tyrosine and increased presence of gelsolin was also observed. This approach can provide enhanced recovery of combination candidate biomarkers across multi-omic platforms, thus, enhancing understanding of in vivo model systems studied by multiple omic technologies.     

Dynamic energy budget approaches for modelling organismal ageing

Ageing is a complex multifactorial process involving a progressive physiological decline that, ultimately, leads to the death of an organism. It involves multiple changes in many components that play fundamental roles under healthy and pathological conditions. Simultaneously, every organism undergoes accumulative 'wear and tear' during its lifespan, which confounds the effects of the ageing process. The scenario is complicated even further by the presence of both age-dependent and age-independent competing causes of death. Various manipulations have been shown to interfere with the ageing process. Calorie restriction, for example, has been reported to increase the lifespan of a wide range of organisms, which suggests a strong relation between energy metabolism and ageing. Such a link is also supported within the main theories for ageing: the free radical hypothesis, for instance, links oxidative damage production directly to energy metabolism. The Dynamic Energy Budgets (DEB) theory, which characterizes the uptake and use of energy by living organisms, therefore constitutes a useful tool for gaining insight into the ageing process. Here we compare the existing DEB-based modelling approaches and, then, discuss how new biological evidence could be incorporated within a DEB framework.     

TIM-3: a novel regulatory molecule of alloimmune activation

T cell Ig domain and mucin domain (TIM)-3 has previously been established as a central regulator of Th1 responses and immune tolerance. In this study, we examined its functions in allograft rejection in a murine model of vascularized cardiac transplantation. TIM-3 was constitutively expressed on dendritic cells and natural regulatory T cells (Tregs) but only detected on CD4(+)FoxP3(-) and CD8(+) T cells in acutely rejecting graft recipients. A blocking anti-TIM-3 mAb accelerated allograft rejection only in the presence of host CD4(+) T cells. Accelerated rejection was accompanied by increased frequencies of alloreactive IFN-γ-, IL-6-, and IL-17-producing splenocytes, enhanced CD8(+) cytotoxicity against alloantigen, increased alloantibody production, and a decline in peripheral and intragraft Treg/effector T cell ratio. Enhanced IL-6 production by CD4(+) T cells after TIM-3 blockade plays a central role in acceleration of rejection. Using an established alloreactivity TCR transgenic model, blockade of TIM-3 increased allospecific effector T cells, enhanced Th1 and Th17 polarization, and resulted in a decreased frequency of overall number of allospecific Tregs. The latter is due to inhibition in induction of adaptive Tregs rather than prevention of expansion of allospecific natural Tregs. In vitro, targeting TIM-3 did not inhibit nTreg-mediated suppression of Th1 alloreactive cells but increased IL-17 production by effector T cells. In summary, TIM-3 is a key regulatory molecule of alloimmunity through its ability to broadly modulate CD4(+) T cell differentiation, thus recalibrating the effector and regulatory arms of the alloimmune response.     

SufU Is an Essential Iron-Sulfur Cluster Scaffold Protein in Bacillus subtilis

Bacteria use three distinct systems for iron-sulfur (Fe/S) cluster biogenesis: the ISC, SUF, and NIF machineries. The ISC and SUF systems are widely distributed, and many bacteria possess both of them. In Escherichia coli, ISC is the major and constitutive system, whereas SUF is induced under iron starvation and/or oxidative stress. Genomic analysis of the Fe/S cluster biosynthesis genes in Bacillus subtilis suggests that this bacterium''s genome encodes only a SUF system consisting of a sufCDSUB gene cluster and a distant sufA gene. Mutant analysis of the putative Fe/S scaffold genes sufU and sufA revealed that sufU is essential for growth under minimal standard conditions, but not sufA. The drastic growth retardation of a conditional mutant depleted of SufU was coupled with a severe reduction of aconitase and succinate dehydrogenase activities in total-cell lysates, suggesting a crucial function of SufU in Fe/S protein biogenesis. Recombinant SufU was devoid of Fe/S clusters after aerobic purification. Upon in vitro reconstitution, SufU bound an Fe/S cluster with up to ∼1.5 Fe and S per monomer. The assembled Fe/S cluster could be transferred from SufU to the apo form of isopropylmalate isomerase Leu1, rapidly forming catalytically active [4Fe-4S]-containing holo-enzyme. In contrast to native SufU, its D43A variant carried a Fe/S cluster after aerobic purification, indicating that the cluster is stabilized by this mutation. Further, we show that apo-SufU is an activator of the cysteine desulfurase SufS by enhancing its activity about 40-fold in vitro. SufS-dependent formation of holo-SufU suggests that SufU functions as an Fe/S cluster scaffold protein tightly cooperating with the SufS cysteine desulfurase.Iron-sulfur (Fe/S) clusters are one of the most ubiquitous and versatile cofactors employed by nature for catalyzing a variety of redox reactions or for serving as redox sensors in a broad range of regulatory processes (10). Iron and sulfide are toxic for the cells in concentrations needed for spontaneous chemical Fe/S protein maturation. Hence, cells have developed complex biosynthesis machineries which are essential in vivo to assemble Fe/S proteins. Three phylogenetically distinct biosynthesis systems have been found in bacteria: ISC (iron-sulfur cluster), SUF (sulfur mobilization), and NIF (nitrogen fixation) (9, 11, 24). The ISC machinery is the most widely distributed bacterial Fe/S cluster biogenesis system and is also present in eukaryotes (31). In Escherichia coli a second system for Fe/S cluster assembly, SUF, is induced under conditions of iron limitation and/or oxidative stress, thus replacing the housekeeping ISC system for assembly of Fe/S proteins. In contrast, SUF was found as the exclusive Fe/S biogenesis system in mycobacteria (22) and Enterococcus faecalis (39) and hence may also serve as a constitutive system. Furthermore, SUF is present in plastids of green plants, resembling the situation found in their cyanobacterial ancestors (25, 53). The NIF system is responsible for the dedicated maturation of the complex Fe/S protein nitrogenase involved in nitrogen fixation, e.g., in Azotobacter vinelandii. Some NIF genes are associated with anaerobic or microaerobic growth in Helicobacter pylori and Entamoeba histolytica (24).Common principles for Fe/S protein assembly in each system have been defined (30). The de novo assembly of an Fe/S cluster occurs on scaffold proteins which transiently bind the Fe/S cluster before transfer to target apoproteins. Cysteine desulfurases such as IscS and SufS serve as sulfur donors, which acquire sulfur from free l-cysteine by pyridoxal-5′-phosphate-dependent desulfuration. The sulfur is transiently bound in the form of a persulfide to an active-site cysteine of the desulfurase and is subsequently transferred to the scaffold protein. Several SUF systems contain SufE, which specifically forms a complex with the cysteine desulfurase SufS (36, 42). SufE enhances SufS activity significantly and assists the sulfur transfer to scaffold proteins. In this case the persulfide is transiently bound to SufE and not to the desulfurase. Recent studies show that the E. coli cysteine desulfurase CsdA is able to complement the SUF system and interacts with SufE if SufS is inactivated (50). A general iron donor involved in Fe/S cluster assembly is not known yet; however, frataxin homologs in prokaryotes and eukaryotes are postulated to deliver iron to the scaffold protein IscU in the ISC system (5, 9, 31, 34).Several components have been suggested to act as scaffold proteins. U-type scaffold proteins such as bacterial NifU, IscU, and eukaryotic Isu1 preferentially bind [2Fe-2S] clusters. However, the assembly of [4Fe-4S] clusters was described to proceed by reductive coupling of two [2Fe-2S] clusters that bind successively to an IscU dimer (1). A-type scaffolds like bacterial SufA or IscA can bind [2Fe-2S] clusters in their monomeric state and were found to be involved in the maturation of [4Fe-4S] proteins such as aconitase (17, 32, 47). Overproduced IscA was also shown to bind mononuclear iron which could be used for Fe/S cluster assembly on IscU in vitro.The ISC system characteristically contains the molecular chaperone pair HscA and HscB that are involved in Fe/S cluster transfer from the IscU scaffold protein to the target proteins. The Hsp70-type HscA specifically binds to a highly conserved LPPVK motif located near the third strictly conserved cluster-binding cysteine in IscU. Specific IscU-HscA complex formation was found to be necessary and sufficient to stimulate the ATPase activity of HscA (12, 21, 48). The SUF system, in contrast, does not contain HscA and HscB chaperones in the suf gene cluster. Instead it comprises the SufBCD proteins. The SufC protein has intrinsic ATPase activity and forms a complex with SufB, a putative scaffold protein, and SufD (29). The precise molecular function of the ATP-hydrolyzing SufBCD complex is not yet clear.The best-characterized SUF system from E. coli contains the gene cluster sufABCDSE (5). However, many bacteria, in particular members of the phylum Firmicutes, contain a different suf gene cluster encoding sufCDSUB, which has been studied so far only by bioinformatic approaches (39, 46). While SufS and SufBCD in the two gene clusters appear to be similar proteins, SufE is lacking in most Firmicutes and also in the mycobacterial and Thermotoga maritima SUF machineries (22, 24). The additionally present SufU shares similarities with IscU of the ISC system (39). The protein contains all three conserved cysteine residues involved in Fe/S cluster association and yet characteristically lacks the LPPVK motif, consistent with the absence of HscA and HscB proteins in sufCDSUB species.In this study, we made use of the Gram-positive bacterium Bacillus subtilis to initiate functional analysis of the sufCDSUB genes in Fe/S cluster biosynthesis by genetic and biochemical approaches. In particular, since no functional information is available for SufU, we tested its putative role as an Fe/S scaffold protein. SufU was found to be crucial for cell viability and for Fe/S-dependent enzyme activities in crude cell lysates. In vitro cluster reconstitution with recombinant SufU indicated that SufU binds a labile Fe/S cluster which can be transferred to apo-Leu1 in a fast and efficient way, fully activating its catalytic function as a [4Fe-4S] cluster-dependent isopropylmalate isomerase. The B. subtilis SufU was found to activate the desulfurase activity of purified B. subtilis SufS. Our results suggest that SufS and the SufU scaffold protein closely act together in the Fe/S cluster biogenesis in B. subtilis.     

Changing climate in Hungary and trends in the annual number of heat stress days

Global climate change can have serious direct effects on animal health and production through heat stress. In Hungary, the number of heat stress days per year (YNHD), i.e., days when the temperature humidity index (THI) is above a specific comfort threshold, has increased in recent years based on observed meteorological data. Between 1973 and 2008, the countrywide average increase in YNHD was 4.1% per year. Climate scenarios based on regional climate models (RCM) were used to predict possible changes in YNHD for the near future (2021–2050) relative to the reference period (1961–1990). This comparison shows that, in Hungary, the 30-year mean of YNHD is expected to increase by between 1 and 27 days, depending on the RCM used. Half of the scenarios investigated in this study predicted that, in large parts of Hungary, YNHD will increase by at least 1 week. However, the increase observed in the past, and that predicted for the near future, is spatially heterogeneous, and areas that currently have large cattle populations are expected to be affected more severely than other regions.     

The Interplay of Proton,Electron, and Metabolite Supply for Photosynthetic H2 Production in Chlamydomonas reinhardtii

To obtain a detailed picture of sulfur deprivation-induced H₂ production in microalgae, metabolome analyses were performed during key time points of the anaerobic H₂ production process of Chlamydomonas reinhardtii. Analyses were performed using gas chromatography coupled to mass spectrometry (GC/MS), two-dimensional gas chromatography combined with time-of-flight mass spectrometry (GCxGC-TOFMS), lipid and starch analysis, and enzymatic determination of fermentative products. The studies were designed to provide a detailed metabolite profile of the solar Bio-H₂ production process. This work reports on the differential analysis of metabolic profiles of the high H₂-producing strain Stm6Glc4 and the wild-type cc406 (WT) before and during the H₂ production phase. Using GCxGC-TOFMS analysis the number of detected peaks increased from 128 peaks, previously detected by GC/MS techniques, to ∼1168. More detailed analysis of the anaerobic H₂ production phase revealed remarkable differences between wild-type and mutant cells in a number of metabolic pathways. Under these physiological conditions the WT produced up to 2.6 times more fatty acids, 2.2 times more neutral lipids, and up to 4 times more fermentation products compared with Stm6Glc4. Based on these results, specific metabolic pathways involving the synthesis of fatty acids, neutral lipids, and fermentation products during anaerobiosis in C. reinhardtii have been identified as potential targets for metabolic engineering to further enhance substrate supply for the hydrogenase(s) in the chloroplast.     

Measuring Preferences on Environmental Damages in LCIA. Part 1: Cognitive Limits in Panel Surveys (9 pp)

-

Part 1: Cognitive Limits in Panel Surveys · Part 2: The Question Format in Panel Surveys This series of two papers discusses the elicitation of weights for damage categories in LCIA with the aid of panel surveys. The papers focus especially on methodological aspects in panel surveys. Part 1 discusses potential cognitive limits of the panel members to understand the reference that is weighted. Part 2 focuses on the influence of the question format and compares results from two different weighting tasks: discrete choice (between alternatives) and score allocation.

Goal, Scope and Background

The weighting of environmental impacts and damages on the safeguard subjects Human Health, Ecosystems, and Resources is a significant step of full aggregated LCIA. Panel surveys have become a common approach in LCIA research to investigate the preferences of stakeholders on environmental impacts and damages. Despite the numerous studies, the knowledge on how to elicit reliable weights is still poor and inconsistent. We present a questionnaire study with 58 environmental science students to investigate so-called framing effects in panel surveys.

Main Features

The study investigates the significance of different framings, which were provided by three references. In addition, the significance of quantitative information provided in the questionnaire is tested. The references are (a1) safeguard subjects without specified additional information, (a2) damages in Europe as they are perceived by the panelist, and (a3) quantified scenarios derived from Eco-indicator99. All participants ranked and rated the importance of the safeguard subjects three times, once within each reference system. According to a test-of-scope study, quantitative information given to the panelist was varied. One level (b1) included data from the Ecoindicator99 methodology, whereas the other group (b2) received data with significantly higher Human Health damages and lower Ecosystem damages, ceteris paribus. This design allows testing the influence of quantitative data on the rating.

Results

The weighting of the safeguard subjects (a1) reveals that Human Health is considered a slightly more important safeguard subject than Ecosystems. However, both are judged to be significantly more important than Resources. This picture changes for the references (a2) and (a3) where damages were weighted. For both references, the respondents rated damages to Ecosystems as most important followed by Resources and Human Health, showing by far the lowest weights. Therefore, the framing of the reference that was weighted played a significant role. The ratings of the subgroups (b1) and (b2) did not differ with respect to the importance of damages, though substantially different quantitative information was given.

Conclusion and Outlook

The participants of the study were obviously insensitive with respect to quantitative information provided. This raises three questions, which are discussed. What is the mental model upon which respondents base their beliefs and values? Can we expect that 'more sophisticated' subjects would respond differently? Which prerequisites should an empirical weighting procedure fulfill in order to incorporate numerical data? We propose different approaches for future procedures in order to accurately analyze these questions.