Belowground DNA-based techniques: untangling the network of plant root interactions

Contains fulltext : 91591.pdf (publisher's version ) (Closed access)7 p

Simulating biosignatures from pre-oxygen photosynthesising life on TRAPPIST-1e

This is the author accepted manuscript. The final version is available on open access from Oxford University Press via the DOI in this recordData availability: The model output used for this study will be made available following this work’s acceptance for publicationIn order to assess observational evidence for potential atmospheric biosignatures on exoplanets, it will be essential to test whether spectral fingerprints from multiple gases can be explained by abiotic or biotic-only processes. Here, we develop and apply a coupled 1D atmosphere-ocean-ecosystem model to understand how primitive biospheres, which exploit abiotic sources of H2 , CO and O2 , could influence the atmospheric composition of rocky terrestrial exoplanets. We apply this to the Earth at 3.8 Ga and to TRAPPIST-1e. We focus on metabolisms that evolved before the evolution of oxygenic photosynthesis, which consume H2 and CO and produce potentially detectable levels of CH4 . O2 -consuming metabolisms are also considered for TRAPPIST-1e, as abiotic O2 production is predicted on M-dwarf orbiting planets. We show that these biospheres can lead to high levels of surface O2 (approximately 1–5 %) as a result of CO consumption, which could allow high O2 scenarios, by removing the main loss mechanisms of atomic oxygen. Increasing stratospheric temperatures, which increases atmospheric OH can reduce the likelihood of such a state forming. O2 -consuming metabolisms could also lower O2 levels to around 10 ppm and support a productive biosphere at low reductant inputs. Using predicted transmission spectral features from CH4 , CO, O2 /O3 and CO2 across the hypothesis space for tectonic reductant input, we show that biotically-produced CH4 may only be detectable at high reductant inputs. CO is also likely to be a dominant feature in transmission spectra for planets orbiting M-dwarfs, which could reduce the confidence in any potential biosignature observations linked to these biospheres.Science and Technology Facilities Council (STFC)UK Research and InnovationJohn Templeton FoundationLeverhulme TrustHill Family ScholarshipInstitute of Physic

γ-Secretase inhibitor enhances antitumour effect of radiation in Notch-expressing lung cancer

BACKGROUND: Notch receptor has an important role in both development and cancer. We previously reported that inhibition of the Notch3 by γ-secretase inhibitor (GSI) induces apoptosis and suppresses tumour proliferation in non-small-cell lung cancer. Although radiation is reported to induce Notch activation, little is known about the relationship between radiation and Notch pathway. METHODS: We examined the effect of combining GSI and radiation at different dosing in three Notch expressing lung cancer cell lines. The cytotoxic effect of GSI and radiation was evaluated using MTT assay and clonogenic assay in vitro and xenograft models. Expressions of Notch pathway, mitogen-activated protein kinase (MAPK) pathway and Bcl-2 family proteins were investigated using western blot analysis. RESULTS: We discovered that the antitumour effect of combining GSI and radiation was dependent on treatment schedule. γ-Secretase inhibitor administration after radiation had the greatest growth inhibition of lung cancer in vitro and in vivo. We showed that the combination induced apoptosis of lung cancer cell lines through the regulation of MAPK and Bcl-2 family proteins. Furthermore, activation of Notch after radiation was ameliorated by GSI administration, suggesting that treatment with GSI prevents Notch-induced radiation resistance. CONCLUSION: Notch has an important role in lung cancer. Treatment with GSI after radiation can significantly enhance radiation-mediated tumour cytotoxicity

Effects of Macromolecular Crowding on Protein Conformational Changes

Many protein functions can be directly linked to conformational changes. Inside cells, the equilibria and transition rates between different conformations may be affected by macromolecular crowding. We have recently developed a new approach for modeling crowding effects, which enables an atomistic representation of “test” proteins. Here this approach is applied to study how crowding affects the equilibria and transition rates between open and closed conformations of seven proteins: yeast protein disulfide isomerase (yPDI), adenylate kinase (AdK), orotidine phosphate decarboxylase (ODCase), Trp repressor (TrpR), hemoglobin, DNA β-glucosyltransferase, and Ap4A hydrolase. For each protein, molecular dynamics simulations of the open and closed states are separately run. Representative open and closed conformations are then used to calculate the crowding-induced changes in chemical potential for the two states. The difference in chemical-potential change between the two states finally predicts the effects of crowding on the population ratio of the two states. Crowding is found to reduce the open population to various extents. In the presence of crowders with a 15 Å radius and occupying 35% of volume, the open-to-closed population ratios of yPDI, AdK, ODCase and TrpR are reduced by 79%, 78%, 62% and 55%, respectively. The reductions for the remaining three proteins are 20–44%. As expected, the four proteins experiencing the stronger crowding effects are those with larger conformational changes between open and closed states (e.g., as measured by the change in radius of gyration). Larger proteins also tend to experience stronger crowding effects than smaller ones [e.g., comparing yPDI (480 residues) and TrpR (98 residues)]. The potentials of mean force along the open-closed reaction coordinate of apo and ligand-bound ODCase are altered by crowding, suggesting that transition rates are also affected. These quantitative results and qualitative trends will serve as valuable guides for expected crowding effects on protein conformation changes inside cells

Tipping the Balance: Robustness of Tip Cell Selection, Migration and Fusion in Angiogenesis

Vascular abnormalities contribute to many diseases such as cancer and diabetic retinopathy. In angiogenesis new blood vessels, headed by a migrating tip cell, sprout from pre-existing vessels in response to signals, e.g., vascular endothelial growth factor (VEGF). Tip cells meet and fuse (anastomosis) to form blood-flow supporting loops. Tip cell selection is achieved by Dll4-Notch mediated lateral inhibition resulting, under normal conditions, in an interleaved arrangement of tip and non-migrating stalk cells. Previously, we showed that the increased VEGF levels found in many diseases can cause the delayed negative feedback of lateral inhibition to produce abnormal oscillations of tip/stalk cell fates. Here we describe the development and implementation of a novel physics-based hierarchical agent model, tightly coupled to in vivo data, to explore the system dynamics as perpetual lateral inhibition combines with tip cell migration and fusion. We explore the tipping point between normal and abnormal sprouting as VEGF increases. A novel filopodia-adhesion driven migration mechanism is presented and validated against in vivo data. Due to the unique feature of ongoing lateral inhibition, ‘stabilised’ tip/stalk cell patterns show sensitivity to the formation of new cell-cell junctions during fusion: we predict cell fates can reverse. The fusing tip cells become inhibited and neighbouring stalk cells flip fate, recursively providing new tip cells. Junction size emerges as a key factor in establishing a stable tip/stalk pattern. Cell-cell junctions elongate as tip cells migrate, which is shown to provide positive feedback to lateral inhibition, causing it to be more susceptible to pathological oscillations. Importantly, down-regulation of the migratory pathway alone is shown to be sufficient to rescue the sprouting system from oscillation and restore stability. Thus we suggest the use of migration inhibitors as therapeutic agents for vascular normalisation in cancer

Targeting Toll-like receptor 7/8 enhances uptake of apoptotic leukemic cells by monocyte-derived dendritic cells but interferes with subsequent cytokine-induced maturation

Therapeutic vaccination with dendritic cells (DC) is an emerging investigational therapy for eradication of minimal residual disease in acute myeloid leukemia. Various strategies are being explored in manufacturing DC vaccines ex vivo, e.g., monocyte-derived DC (MoDC) loaded with leukemia-associated antigens (LAA). However, the optimal source of LAA and the choice of DC-activating stimuli are still not well defined. Here, loading with leukemic cell preparations (harboring both unknown and known LAA) was explored in combination with a DC maturation-inducing cytokine cocktail (CC; IL-1β, IL-6, TNF-α, and PGE2) and Toll-like receptor ligands (TLR-L) to optimize uptake. Since heat shock induced apoptotic blasts were more efficiently taken up than lysates, we focused on uptake of apoptotic leukemic cells. Uptake of apoptotic blast was further enhanced by the TLR7/8-L R848 (20–30%); in contrast, CC-induced maturation inhibited uptake. CC, and to a lesser extent R848, enhanced the ability of MoDC to migrate and stimulate T cells. Furthermore, class II-associated invariant chain peptide expression was down-modulated after R848- or CC-induced maturation, indicating enhanced processing and presentation of antigenic peptides. To improve both uptake and maturation, leukemic cells and MoDC were co-incubated with R848 for 24 h followed by addition of CC. However, this approach interfered with CC-mediated MoDC maturation as indicated by diminished migratory and T cell stimulatory capacity, and the absence of IL-12 production. Taken together, our data demonstrate that even though R848 improved uptake of apoptotic leukemic cells, the sequential use of R848 and CC is counter-indicated due to its adverse effects on MoDC maturation

Pain Management in Patients with Cancer: Focus on Opioid Analgesics

Cancer pain is generally treated with pharmacological measures, relying on using opioids alone or in combination with adjuvant analgesics. Weak opioids are used for mild-to-moderate pain as monotherapy or in a combination with nonopioids. For patients with moderate-to-severe pain, strong opioids are recommended as initial therapy rather than beginning treatment with weak opioids. Adjunctive therapy plays an important role in the treatment of cancer pain not fully responsive to opioids administered alone (ie, neuropathic, bone, and visceral colicky pain). Supportive drugs should be used wisely to prevent and treat opioids’ adverse effects. Understanding the pharmacokinetics, pharmacodynamics, interactions, and cautions with commonly used opioids can help determine appropriate opioid selection for individual cancer patients

Barrier-to-Autointegration Factor Proteome Reveals Chromatin-Regulatory Partners

Nuclear lamin filaments and associated proteins form a nucleoskeletal (“lamina”) network required for transcription, replication, chromatin organization and epigenetic regulation in metazoans. Lamina defects cause human disease (“laminopathies”) and are linked to aging. Barrier-to-autointegration factor (BAF) is a mobile and essential component of the nuclear lamina that binds directly to histones, lamins and LEM-domain proteins, including the inner nuclear membrane protein emerin, and has roles in chromatin structure, mitosis and gene regulation. To understand BAF's mechanisms of action, BAF associated proteins were affinity-purified from HeLa cell nuclear lysates using BAF-conjugated beads, and identified by tandem mass spectrometry or independently identified and quantified using the iTRAQ method. We recovered A- and B-type lamins and core histones, all known to bind BAF directly, plus four human transcription factors (Requiem, NonO, p15, LEDGF), disease-linked proteins (e.g., Huntingtin, Treacle) and several proteins and enzymes that regulate chromatin. Association with endogenous BAF was independently validated by co-immunoprecipitation from HeLa cells for seven candidates including Requiem, poly(ADP-ribose) polymerase 1 (PARP1), retinoblastoma binding protein 4 (RBBP4), damage-specific DNA binding protein 1 (DDB1) and DDB2. Interestingly, endogenous BAF and emerin each associated with DDB2 and CUL4A in a UV- and time-dependent manner, suggesting BAF and emerin have dynamic roles in genome integrity and might help couple DNA damage responses to the nuclear lamina network. We conclude this proteome is a rich source of candidate partners for BAF and potentially also A- and B-type lamins, which may reveal how chromatin regulation and genome integrity are linked to nuclear structure

A Complete Pathway Model for Lipid A Biosynthesis in Escherichia coli.

Lipid A is a highly conserved component of lipopolysaccharide (LPS), itself a major component of the outer membrane of Gram-negative bacteria. Lipid A is essential to cells and elicits a strong immune response from humans and other animals. We developed a quantitative model of the nine enzyme-catalyzed steps of Escherichia coli lipid A biosynthesis, drawing parameters from the experimental literature. This model accounts for biosynthesis regulation, which occurs through regulated degradation of the LpxC and WaaA (also called KdtA) enzymes. The LpxC degradation signal appears to arise from the lipid A disaccharide concentration, which we deduced from prior results, model results, and new LpxK overexpression results. The model agrees reasonably well with many experimental findings, including the lipid A production rate, the behaviors of mutants with defective LpxA enzymes, correlations between LpxC half-lives and cell generation times, and the effects of LpxK overexpression on LpxC concentrations. Its predictions also differ from some experimental results, which suggest modifications to the current understanding of the lipid A pathway, such as the possibility that LpxD can replace LpxA and that there may be metabolic channeling between LpxH and LpxB. The model shows that WaaA regulation may serve to regulate the lipid A production rate when the 3-deoxy-D-manno-oct-2-ulosonic acid (KDO) concentration is low and/or to control the number of KDO residues that get attached to lipid A. Computation of flux control coefficients showed that LpxC is the rate-limiting enzyme if pathway regulation is ignored, but that LpxK is the rate-limiting enzyme if pathway regulation is present, as it is in real cells. Control also shifts to other enzymes if the pathway substrate concentrations are not in excess. Based on these results, we suggest that LpxK may be a much better drug target than LpxC, which has been pursued most often

"Nested" cryptic diversity in a widespread marine ecosystem engineer: a challenge for detecting biological invasions

<p>Abstract</p> <p>Background</p> <p>Ecosystem engineers facilitate habitat formation and enhance biodiversity, but when they become invasive, they present a critical threat to native communities because they can drastically alter the receiving habitat. Management of such species thus needs to be a priority, but the poorly resolved taxonomy of many ecosystem engineers represents a major obstacle to correctly identifying them as being either native or introduced. We address this dilemma by studying the sea squirt <it>Pyura stolonifera</it>, an important ecosystem engineer that dominates coastal communities particularly in the southern hemisphere. Using DNA sequence data from four independently evolving loci, we aimed to determine levels of cryptic diversity, the invasive or native status of each regional population, and the most appropriate sampling design for identifying the geographic ranges of each evolutionary unit.</p> <p>Results</p> <p>Extensive sampling in Africa, Australasia and South America revealed the existence of "nested" levels of cryptic diversity, in which at least five distinct species can be further subdivided into smaller-scale genetic lineages. The ranges of several evolutionary units are limited by well-documented biogeographic disjunctions. Evidence for both cryptic native diversity and the existence of invasive populations allows us to considerably refine our view of the native versus introduced status of the evolutionary units within <it>Pyura stolonifera </it>in the different coastal communities they dominate.</p> <p>Conclusions</p> <p>This study illustrates the degree of taxonomic complexity that can exist within widespread species for which there is little taxonomic expertise, and it highlights the challenges involved in distinguishing between indigenous and introduced populations. The fact that multiple genetic lineages can be native to a single geographic region indicates that it is imperative to obtain samples from as many different habitat types and biotic zones as possible when attempting to identify the source region of a putative invader. "Nested" cryptic diversity, and the difficulties in correctly identifying invasive species that arise from it, represent a major challenge for managing biodiversity.</p