The Plastid Outer Envelope – A Highly Dynamic Interface between Plastid and Cytoplasm

Plastids are the defining organelles of all photosynthetic eukaryotes. They are the site of photosynthesis and of a large number of other essential metabolic pathways, such as fatty acid and amino acid biosyntheses, sulfur and nitrogen assimilation, and aromatic and terpenoid compound production, to mention only a few examples. The metabolism of plastids is heavily intertwined and connected with that of the surrounding cytosol, thus causing massive traffic of metabolic precursors, intermediates, and products. Two layers of biological membranes that are called the inner (IE) and the outer (OE) plastid envelope membranes bound the plastids of Archaeplastida. While the IE is generally accepted as the osmo-regulatory barrier between cytosol and stroma, the OE was considered to represent an unspecific molecular sieve, permeable for molecules of up to 10 kDa. However, after the discovery of small substrate specific pores in the OE, this view has come under scrutiny. In addition to controlling metabolic fluxes between plastid and cytosol, the OE is also crucial for protein import into the chloroplast. It contains the receptors and translocation channel of the TOC complex that is required for the canonical post-translational import of nuclear-encoded, plastid-targeted proteins. Further, the OE is a metabolically active compartment of the chloroplast, being involved in, e.g., fatty acid metabolism and membrane lipid production. Also, recent findings hint on the OE as a defense platform against several biotic and abiotic stress conditions, such as cold acclimation, freezing tolerance, and phosphate deprivation. Moreover, dynamic non-covalent interactions between the OE and the endomembrane system are thought to play important roles in lipid and non-canonical protein trafficking between plastid and endoplasmic reticulum. While proteomics and bioinformatics has provided us with comprehensive but still incomplete information on proteins localized in the plastid IE, the stroma, and the thylakoids, our knowledge of the protein composition of the plastid OE is far from complete. In this article, we report on the recent progress in discovering novel OE proteins to draw a conclusive picture of the OE. A “parts list” of the plastid OE will be presented, using data generated by proteomics of plastids isolated from various plant sources

Understanding metabolite transport and metabolism in C-4 plants through RNA-seq

Schlueter U, Denton AK, Bräutigam A. Understanding metabolite transport and metabolism in C-4 plants through RNA-seq. Current Opinion in Plant Biology. 2016;31:83-90.RNA-seq, the measurement of steady-state RNA levels by next generation sequencing, has enabled quantitative transcriptome analyses of complex traits in many species without requiring the parallel sequencing of their genomes. The complex trait of C-4 photosynthesis, which increases photosynthetic efficiency via a biochemical pump that concentrates CO2 around RubisCO, has evolved convergently multiple times. Due to these interesting properties, C-4 photosynthesis has been analyzed in a series of comparative RNA-seq projects. These projects compared both species with and without the C-4 trait and different tissues or organs within a C-4 plant. The RNA-seq studies were evaluated by comparing to earlier single gene studies. The studies confirmed the marked changes expected for C-4 signature genes, but also revealed numerous new players in C-4 metabolism showing that the C-4 cycle is more complex than previously thought, and suggesting modes of integration into the underlying C-3 metabolism

Plastid Signals and the Bundle Sheath: Mesophyll Development in Reticulate Mutants

Lundquist PK, Rosar C, Bräutigam A, Weber APM. Plastid Signals and the Bundle Sheath: Mesophyll Development in Reticulate Mutants. Molecular Plant. 2014;7(1):14-29.The development of a plant leaf is a meticulously orchestrated sequence of events producing a complex organ comprising diverse cell types. The reticulate class of leaf variegation mutants displays contrasting pigmentation between veins and interveinal regions due to specific aberrations in the development of mesophyll cells. Thus, the reticulate mutants offer a potent tool to investigate cell-type-specific developmental processes. The discovery that most mutants are affected in plastid-localized, metabolic pathways that are strongly expressed in vasculature-associated tissues implicates a crucial role for the bundle sheath and their chloroplasts in proper development of the mesophyll cells. Here, we review the reticulate mutants and their phenotypic characteristics, with a focus on those in Arabidopsis thaliana. Two alternative models have been put forward to explain the relationship between plastid metabolism and mesophyll cell development, which we call here the supply and the signaling hypotheses. We critically assess these proposed models and discuss their implications for leaf development and bundle sheath function in C3 species. The characterization of the reticulate mutants supports the significance of plastid retrograde signaling in cell development and highlights the significance of the bundle sheath in C3 photosynthesis

Total colonic aganglionosis : multicentre study of surgical treatment and patient-reported outcomes up to adulthood

Background: Surgery for total colonic aganglionosis (TCA) is designed to preserve continence and achieve satisfactory quality of life. This study evaluated a comprehensive group of clinical and social outcomes. Methods: An international multicentre study from eight Nordic hospitals involving examination of case records and a patient-reported questionnaire survey of all patients born with TCA between 1987 and 2006 was undertaken. Results: Of a total of 116 patients, five (4 center dot 3 per cent) had died and 102 were traced. Over a median follow-up of 12 (range 0 center dot 3-33) years, bowel continuity was established in 75 (73 center dot 5 per cent) at a median age of 11 (0 center dot 5-156) months. Mucosectomy with a short muscular cuff and straight ileoanal anastomosis (SIAA) (29 patients) or with aJpouch (JIAA) (26) were the most common reconstructions (55 of 72, 76 per cent). Major early postoperative complications requiring surgical intervention were observed in four (6 per cent) of the 72 patients. In 57 children aged over 4 years, long-term functional bowel symptoms after reconstruction included difficulties in holding back defaecation in 22 (39 per cent), more than one faecal accident per week in nine (16 per cent), increased frequency of defaecation in 51 (89 per cent), and social restrictions due to bowel symptoms in 35 (61 per cent). Enterocolitis occurred in 35 (47 per cent) of 72 patients. Supplementary enteral and/or parenteral nutrition was required by 51 (55 per cent) of 93 patients at any time during follow-up. Of 56 responders aged 2-20 years, true low BMI for age was found in 20 (36 per cent) and 13 (23 per cent) were short for age. Conclusion: Reconstruction for TCA was associated with persistent bowel symptoms, and enterocolitis remained common. Multidisciplinary follow-up, including continuity of care in adulthood, might improve care standards in patients with TCA.Peer reviewe

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

Modified bibenzimidazole ligands as spectator ligands in photoactive molecular functional Ru-polypyridine units? Implications from spectroscopy

The photophysical properties of Ruthenium-bipyridine complexes bearing a bibenzimidazole ligand were investigated. The nitrogens on the bibenzimidazole-ligand were protected, by adding either a phenylene group or a 1,2-ethandiyl group, to remove the photophysical dependence of the complex on the protonation state of the bibenzimidazole ligand. This protection results in the bibenzimidazole ligand contributing to the MLCT transition, which is experimentally evidenced by (resonance) Raman scattering in concert with DFT calculations for a detailed mode assignment in the (resonance) Raman spectra

Glioblastoma and glioblastoma stem cells are dependent on functional MTH1

Glioblastoma multiforme (GBM) is an aggressive form of brain cancer with poor prognosis. Cancer cells are characterized by a specific redox environment that adjusts metabolism to its specific needs and allows the tumor to grow and metastasize. As a consequence, cancer cells and especially GBM cells suffer from elevated oxidative pressure which requires antioxidant-defense and other sanitation enzymes to be upregulated. MTH1, which degrades oxidized nucleotides, is one of these defense enzymes and represents a promising cancer target. We found MTH1 expression levels elevated and correlated with GBM aggressiveness and discovered that siRNA knock-down or inhibition of MTH1 with small molecules efficiently reduced viability of patient-derived GBM cultures. The effect of MTH1 loss on GBM viability was likely mediated through incorporation of oxidized nucleotides and subsequent DNA damage. We revealed that MTH1 inhibition targets GBM independent of aggressiveness as well as potently kills putative GBM stem cells in vitro. We used an orthotopic zebrafish model to confirm our results in vivo and light-sheet microscopy to follow the effect of MTH1 inhibition in GBM in real time. In conclusion, MTH1 represents a promising target for GBM therapy and MTH1 inhibitors may also be effective in patients that suffer from recurring disease