Beclin1-binding UVRAG targets the class C Vps complex to coordinate autophagosome maturation and endocytic trafficking

Autophagic and endocytic pathways are tightly regulated membrane rearrangement processes that are crucial for homeostasis, development and disease. Autophagic cargo is delivered from autophagosomes to lysosomes for degradation through a complex process that topologically resembles endosomal maturation. Here, we report that a Beclin1-binding autophagic tumour suppressor, UVRAG, interacts with the class C Vps complex, a key component of the endosomal fusion machinery. This interaction stimulates Rab7 GTPase activity and autophagosome fusion with late endosomes/lysosomes, thereby enhancing delivery and degradation of autophagic cargo. Furthermore, the UVRAG-class-C-Vps complex accelerates endosome-endosome fusion, resulting in rapid degradation of endocytic cargo. Remarkably, autophagosome/endosome maturation mediated by the UVRAG-class-C-Vps complex is genetically separable from UVRAG-Beclin1-mediated autophagosome formation. This result indicates that UVRAG functions as a multivalent trafficking effector that regulates not only two important steps of autophagy - autophagosome formation and maturation - but also endosomal fusion, which concomitantly promotes transport of autophagic and endocytic cargo to the degradative compartments.     

A high-throughput method for liquid chromatography–tandem mass spectrometry determination of plasma alkylresorcinols,biomarkers of whole grain wheat and rye intake

Plasma alkylresorcinols are increasingly analyzed in cohort studies to improve estimates of whole grain intake and their relationship with disease incidence. Current methods require large volumes of solvent (>10 ml/sample) and have relatively low daily sample throughput. We tested five different supported extraction methods for extracting alkylresorcinols from plasma and improved a normal-phase liquid chromatography coupled to a tandem mass spectrometer method to reduce sample analysis time. The method was validated and compared with gas chromatography–mass spectrometry analysis. Sample preparation with HybridSPE supported extraction was most effective for alkylresorcinol extraction, with recoveries of 77–82% from 100 μl of plasma. The use of 96-well plates allowed extraction of 160 samples per day. Using a 5-cm NH₂ column and heptane reduced run times to 3 min. The new method had a limit of detection and limit of quantification equivalent to 1.1–1.8 nmol/L and 3.5–6.1 nmol/L plasma, respectively, for the different alkylresorcinol homologues. Accuracy was 93–105%, and intra- and inter-batch precision values were 4–18% across different plasma concentrations. This method makes it possible to quantify plasma alkylresorcinols in 100 μl of plasma at a rate of at least 160 samples per day without the need for large volumes of organic solvents.     

Viral fusion peptides and identification of membrane-interacting segments

Viral envelope glycoproteins promote infection by mediating fusion between viral and cellular membranes. Fusion occurs after dramatic conformational changes within fusion proteins, leading to the exposure of a short stretch of mostly apolar residues, termed the fusion peptide, which is presumed to insert into the membrane and initiate the fusion process. The typical global composition of fusion peptides, rich in hydrophobic but also in small amino acids such as alanine and glycine, was used here as bait to detect other peptidic segments that can insert into membranes. We so evidenced a similar composition in several cytotoxic peptides, which promote pore formation such as peptides involved in amyloidoses and hydrophobic alpha-hairpins of pore-forming toxins. It is suggested that the structural plasticity observed for several membrane active peptides can be conferred by this particular global amino acid composition, which could be thus used to predict such functional behavior from genome data.     

How life‐history traits affect ecosystem properties: effects of dispersal in meta‐ecosystems

The concept of life‐history traits and the study of these traits are the hallmark of population biology. Acknowledging their variability and evolution has allowed us to understand how species adapt in response to their environment. The same traits are also involved in how species alter ecosystems and shape their dynamics and functioning. Some theories, such as the metabolic theory of ecology, ecological stoichiometry or pace‐of‐life theory, already recognize this junction, but only do so in an implicitly non‐spatial context. Meanwhile, for a decade now, it has been argued that ecosystem properties have to be understood at a larger scale using meta‐ecosystem theory because source–sink dynamics, community assembly and ecosystem stability are all modified by spatial structure. Here, we argue that some ecosystem properties can be linked to a single life‐history trait, dispersal, i.e. the tendency of organisms to live, compete and reproduce away from their birth place. By articulating recent theoretical and empirical studies linking ecosystem functioning and dynamics to species dispersal, we aim to highlight both the known connections between life‐history traits and ecosystem properties and the unknown areas, which deserve further empirical and theoretical developments.     

Linkage and Association Mapping for Two Major Traits Used in the Maritime Pine Breeding Program: Height Growth and Stem Straightness

BackgroundIncreasing our understanding of the genetic architecture of complex traits, through analyses of genotype-phenotype associations and of the genes/polymorphisms accounting for trait variation, is crucial, to improve the integration of molecular markers into forest tree breeding. In this study, two full-sib families and one breeding population of maritime pine were used to identify quantitative trait loci (QTLs) for height growth and stem straightness, through linkage analysis (LA) and linkage disequilibrium (LD) mapping approaches.ResultsThe populations used for LA consisted of two unrelated three-generation full-sib families (n = 197 and n = 477). These populations were assessed for height growth or stem straightness and genotyped for 248 and 217 markers, respectively. The population used for LD mapping consisted of 661 founders of the first and second generations of the breeding program. This population was phenotyped for the same traits and genotyped for 2,498 single-nucleotide polymorphism (SNP) markers corresponding to 1,652 gene loci. The gene-based reference genetic map of maritime pine was used to localize and compare the QTLs detected by the two approaches, for both traits. LA identified three QTLs for stem straightness and two QTLs for height growth. The LD study yielded seven significant associations (P ≤ 0.001): four for stem straightness and three for height growth. No colocalisation was found between QTLs identified by LA and SNPs detected by LD mapping for the same trait.ConclusionsThis study provides the first comparison of LA and LD mapping approaches in maritime pine, highlighting the complementary nature of these two approaches for deciphering the genetic architecture of two mandatory traits of the breeding program.     

Mitochondrial F1F0-ATP synthase and organellar internal architecture

The mitochondrial F₁F₀-ATP synthase adopts supramolecular structures. The interaction domains between monomers involve components belonging to the F₀ domains. In Saccharomyces cerevisiae, alteration of these components destabilizes the oligomeric structures, leading concomitantly to the appearance of monomeric species of ATP synthase and anomalous mitochondrial morphologies in the form of onion-like structures. The mitochondrial ultrastructure at the cristae level is thus modified. Electron microscopy on cross-sections of wild type mitochondria display many short cristae with narrowed intra-cristae space, whereas yeast mutants defected in supramolecular ATP synthases assembly present a low number of large lamellar cristae of constant thickness and traversing the whole organelle. The growth of these internal structures leads finally to mitochondria with sphere-like structures with a mean diameter of 1 μm that are easily identified by epifluorescence microscopy. As a result, ATP synthase is an actor of the mitochondrial ultrastructure in yeast. This paper reviews the ATP synthase components whose modifications lead to anomalous mitochondrial morphology and also provides a schema showing the formation of the so-called onion-like structures.     

Different expression levels of the TAP peptide transporter lead to recognition of different antigenic peptides by tumor-specific CTL

Decreased antigenicity of cancer cells is a major problem in tumor immunology. This is often acquired by an expression defect in the TAP. However, it has been reported that certain murine Ags appear on the target cell surface upon impairment of TAP expression. In this study, we identified a human CTL epitope belonging to this Ag category. This epitope is derived from preprocalcitonin (ppCT) signal peptide and is generated within the endoplasmic reticulum by signal peptidase and signal peptide peptidase. Lung cancer cells bearing this antigenic peptide displayed low levels of TAP, but restoration of their expression by IFN-γ treatment or TAP1 and TAP2 gene transfer abrogated ppCT Ag presentation. In contrast, TAP upregulation in the same tumor cells increased their recognition by proteasome/TAP-dependent peptide-specific CTLs. Thus, to our knowledge, ppCT(16-25) is the first human tumor epitope whose surface expression requires loss or downregulation of TAP. Lung tumors frequently display low levels of TAP molecules and might thus be ignored by the immune system. Our results suggest that emerging signal peptidase-generated peptides represent alternative T cell targets, which permit CTLs to destroy TAP-impaired tumors and thus overcome tumor escape from CD8(+) T cell immunity.     

Peptides of type II collagen can induce the cleavage of type II collagen and aggrecan in articular cartilage.

The objective of this study was to determine whether a fragment(s) of type II collagen can induce cartilage degradation. Fragments generated by cyanogen bromide (CB) cleavage of purified bovine type II collagen were separated by HPLC. These fragments together with selected overlapping synthetic peptides were first analysed for their capacity to induce cleavage of type II collagen by collagenases in chondrocyte and explant cultures of healthy adult bovine articular cartilage. Collagen cleavage was measured by immunoassay and degradation of proteoglycan (mainly aggrecan) was determined by analysis of cleavage products of core protein by Western blotting. Gene expression of matrix metalloproteinases MMP-13 and MMP-1 was measured using Real-time PCR. Induction of denaturation of type II collagen in situ in cartilage matrix with exposure of the CB domain was identified with a polyclonal and monoclonal antibodies that only react with this domain in denatured but not native type II collagen. As well as the mixture of CB fragments and peptide CB12, a single synthetic peptide CB12-II (residues 195-218), but not synthetic peptide CB12-IV (residues 231-254), potently and consistently induced in explant cultures at 10 microM and 25 microM, in a time, cell and dose dependent manner, collagenase-induced cleavage of type II collagen accompanied by upregulation of MMP-13 expression but not MMP-1. In isolated chondrocyte cultures CB12-II induced very limited upregulation of MMP-13 as well as MMP-1 expression. Although this was accompanied by concomitant induction of cleavage of type II collagen by collagenases, this was not associated by aggrecan cleavage. Peptide CB12-IV, which had no effect on collagen cleavage, clearly induced aggrecanase specific cleavage of the core protein of this proteoglycan. Thus these events involving matrix molecule cleavage can importantly occur independently of each other, contrary to popular belief. Denaturation of type II collagen with exposure of the CB12-II domain was also shown to be much increased in osteoarthritic human cartilage compared to non-arthritic cartilage. These observations reveal that peptides of type II collagen, to which there is increased exposure in osteoarthritic cartilage, can when present in sufficient concentration induce cleavage of type II collagen (CB12-II) and aggrecan (CB12-IV) accompanied by increased expression of collagenases. Such increased concentrations of denatured collagen are present in adult and osteoarthritic cartilages and the exposure of chondrocytes to the sequences they encode, either in soluble or more likely insoluble form, may therefore play a role in the excessive resorption of matrix molecules that is seen in arthritis and development.     

Trade‐offs between land and water requirements for large‐scale bioenergy production

Bioenergy is expected to play an important role in the future energy mix as it can substitute fossil fuels and contribute to climate change mitigation. However, large‐scale bioenergy cultivation may put substantial pressure on land and water resources. While irrigated bioenergy production can reduce the pressure on land due to higher yields, associated irrigation water requirements may lead to degradation of freshwater ecosystems and to conflicts with other potential users. In this article, we investigate the trade‐offs between land and water requirements of large‐scale bioenergy production. To this end, we adopt an exogenous demand trajectory for bioenergy from dedicated energy crops, targeted at limiting greenhouse gas emissions in the energy sector to 1100 Gt carbon dioxide equivalent until 2095. We then use the spatially explicit global land‐ and water‐use allocation model MAgPIE to project the implications of this bioenergy target for global land and water resources. We find that producing 300 EJ yr^?1 of bioenergy in 2095 from dedicated bioenergy crops is likely to double agricultural water withdrawals if no explicit water protection policies are implemented. Since current human water withdrawals are dominated by agriculture and already lead to ecosystem degradation and biodiversity loss, such a doubling will pose a severe threat to freshwater ecosystems. If irrigated bioenergy production is prohibited to prevent negative impacts of bioenergy cultivation on water resources, bioenergy land requirements for meeting a 300 EJ yr^?1 bioenergy target increase substantially (+ 41%) – mainly at the expense of pasture areas and tropical forests. Thus, avoiding negative environmental impacts of large‐scale bioenergy production will require policies that balance associated water and land requirements.