Tracing back to Sources of MAIC Using Farm records and Lab Techniques

For the implementation of tracing back systems (particular agents, unwanted observation and other), detection techniques should be available. For MAIC, a step-by-step approach was established, combining visual observation, lab based PCR-identification, tracing back to the farm of origin and finally the search for potential ports of entry

Comprehensive transcriptome analysis of the highly complex Pisum sativum genome using next generation sequencing

<p>Abstract</p> <p>Background</p> <p>The garden pea, <it>Pisum sativum</it>, is among the best-investigated legume plants and of significant agro-commercial relevance. <it>Pisum sativum </it>has a large and complex genome and accordingly few comprehensive genomic resources exist.</p> <p>Results</p> <p>We analyzed the pea transcriptome at the highest possible amount of accuracy by current technology. We used next generation sequencing with the Roche/454 platform and evaluated and compared a variety of approaches, including diverse tissue libraries, normalization, alternative sequencing technologies, saturation estimation and diverse assembly strategies. We generated libraries from flowers, leaves, cotyledons, epi- and hypocotyl, and etiolated and light treated etiolated seedlings, comprising a total of 450 megabases. Libraries were assembled into 324,428 unigenes in a first pass assembly.</p> <p>A second pass assembly reduced the amount to 81,449 unigenes but caused a significant number of chimeras. Analyses of the assemblies identified the assembly step as a major possibility for improvement. By recording frequencies of Arabidopsis orthologs hit by randomly drawn reads and fitting parameters of the saturation curve we concluded that sequencing was exhaustive. For leaf libraries we found normalization allows partial recovery of expression strength aside the desired effect of increased coverage. Based on theoretical and biological considerations we concluded that the sequence reads in the database tagged the vast majority of transcripts in the aerial tissues. A pathway representation analysis showed the merits of sampling multiple aerial tissues to increase the number of tagged genes. All results have been made available as a fully annotated database in fasta format.</p> <p>Conclusions</p> <p>We conclude that the approach taken resulted in a high quality - dataset which serves well as a first comprehensive reference set for the model legume pea. We suggest future deep sequencing transcriptome projects of species lacking a genomics backbone will need to concentrate mainly on resolving the issues of redundancy and paralogy during transcriptome assembly.</p

PLGG1, a plastidic glycolate glycerate transporter, is required for photorespiration and defines a unique class of metabolite transporters

Pick TR, Bräutigam A, Schulz MA, Obata T, Fernie AR, Weber APM. PLGG1, a plastidic glycolate glycerate transporter, is required for photorespiration and defines a unique class of metabolite transporters. Proceedings of the National Academy of Sciences of the United States of America. 2013;110(8):3185-3190.Photorespiratory carbon flux reaches up to a third of photosynthetic flux, thus contributes massively to the global carbon cycle. The pathway recycles glycolate-2-phosphate, the most abundant byproduct of RubisCO reactions. This oxygenation reaction of RubisCO and subsequent photorespiration significantly limit the biomass gains of many crop plants. Although photorespiration is a compartmentalized process with enzymatic reactions in the chloroplast, the peroxisomes, the mitochondria, and the cytosol, no transporter required for the core photorespiratory cycle has been identified at the molecular level to date. Using transcript coexpression analyses, we identified Plastidal glycolate glycerate translocator 1 (PLGG1) as a candidate core photorespiratory transporter. Related genes are encoded in the genomes of archaea, bacteria, fungi, and all Archaeplastida and have previously been associated with a function in programmed cell-death. A mutant deficient in PLGG1 shows WT-like growth only in an elevated carbon dioxide atmosphere. The mutant accumulates glycolate and glycerate, leading to the hypothesis that PLGG1 is a glycolate/glycerate transporter. This hypothesis was tested and supported by in vivo and in vitro transport assays and O-18(2)-metabolic flux profiling. Our results indicate that PLGG1 is the chloroplastidic glycolate/glycerate transporter, which is required for the function of the photorespiratory cycle. Identification of the PLGG1 transport function will facilitate unraveling the role of similar proteins in bacteria, archaea, and fungi in the future

Systematic Analysis of Stability Patterns in Plant Primary Metabolism

Metabolic networks are characterized by complex interactions and regulatory mechanisms between many individual components. These interactions determine whether a steady state is stable to perturbations. Structural kinetic modeling (SKM) is a framework to analyze the stability of metabolic steady states that allows the study of the system Jacobian without requiring detailed knowledge about individual rate equations. Stability criteria can be derived by generating a large number of structural kinetic models (SK-models) with randomly sampled parameter sets and evaluating the resulting Jacobian matrices. Until now, SKM experiments applied univariate tests to detect the network components with the largest influence on stability. In this work, we present an extended SKM approach relying on supervised machine learning to detect patterns of enzyme-metabolite interactions that act together in an orchestrated manner to ensure stability. We demonstrate its application on a detailed SK-model of the Calvin-Benson cycle and connected pathways. The identified stability patterns are highly complex reflecting that changes in dynamic properties depend on concerted interactions between several network components. In total, we find more patterns that reliably ensure stability than patterns ensuring instability. This shows that the design of this system is strongly targeted towards maintaining stability. We also investigate the effect of allosteric regulators revealing that the tendency to stability is significantly increased by including experimentally determined regulatory mechanisms that have not yet been integrated into existing kinetic models

MutT homologue 1 (MTH1) catalyzes the hydrolysis of mutagenic O6-methyl-dGTP.

Nucleotides in the free pool are more susceptible to nonenzymatic methylation than those protected in the DNA double helix. Methylated nucleotides like O6-methyl-dGTP can be mutagenic and toxic if incorporated into DNA. Removal of methylated nucleotides from the nucleotide pool may therefore be important to maintain genome integrity. We show that MutT homologue 1 (MTH1) efficiently catalyzes the hydrolysis of O6-methyl-dGTP with a catalytic efficiency similar to that for 8-oxo-dGTP. O6-methyl-dGTP activity is exclusive to MTH1 among human NUDIX proteins and conserved through evolution but not found in bacterial MutT. We present a high resolution crystal structure of human and zebrafish MTH1 in complex with O6-methyl-dGMP. By microinjecting fertilized zebrafish eggs with O6-methyl-dGTP and inhibiting MTH1 we demonstrate that survival is dependent on active MTH1 in vivo. O6-methyl-dG levels are higher in DNA extracted from zebrafish embryos microinjected with O6-methyl-dGTP and inhibition of O6-methylguanine-DNA methyl transferase (MGMT) increases the toxicity of O6-methyl-dGTP demonstrating that O6-methyl-dGTP is incorporated into DNA. MTH1 deficiency sensitizes human cells to the alkylating agent Temozolomide, a sensitization that is more pronounced upon MGMT inhibition. These results expand the cellular MTH1 function and suggests MTH1 also is important for removal of methylated nucleotides from the nucleotide pool

A remote secondary binding pocket promotes heteromultivalent targeting of DC-SIGN

Dendritic cells (DC) are antigen-presenting cells coordinating the interplay of the innate and the adaptive immune response. The endocytic C-type lectin receptors DC-SIGN and Langerin display expression profiles restricted to distinct DC subtypes and have emerged as prime targets for next-generation immunotherapies and anti-infectives. Using heteromultivalent liposomes copresenting mannosides bearing aromatic aglycones with natural glycan ligands, we serendipitously discovered striking cooperativity effects for DC-SIGN+ but not for Langerin+ cell lines. Mechanistic investigations combining NMR spectroscopy with molecular docking and molecular dynamics simulations led to the identification of a secondary binding pocket for the glycomimetics. This pocket, located remotely of DC-SIGN’s carbohydrate bindings site, can be leveraged by heteromultivalent avidity enhancement. We further present preliminary evidence that the aglycone allosterically activates glycan recognition and thereby contributes to DC-SIGN-specific cell targeting. Our findings have important implications for both translational and basic glycoscience, showcasing heteromultivalent targeting of DCs to improve specificity and supporting potential allosteric regulation of DC-SIGN and CLRs in general

Queering Development? The Unsettling Geographies of South–South Cooperation

This paper deploys queer theory as a way of approaching South–South Cooperation (SSC). It examines the ways in which Southern development partners are not simply up-ending the long-standing spatialities, imaginaries and identities (re)produced through the mainstream international development regime, but queering terminologies and definitions, while presenting themselves in fluid ways, enrolling different identities and attributes in different places and to different audiences. At the same time, a queer lens reveals the (re)inscription of gendered, sexualised and racialised identities and hierarchies through the relationships, intimacies and practices of SSC. The paper proposes that queer theory can offer productive insights into the complex and compelling phenomenon of SSC, and the transgressive challenges to the postcolonial hierarchies and binaries of “traditional” international development