136 research outputs found

    Whole-Genome Analysis of the SHORT-ROOT Developmental Pathway in Arabidopsis

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    Stem cell function during organogenesis is a key issue in developmental biology. The transcription factor SHORT-ROOT (SHR) is a critical component in a developmental pathway regulating both the specification of the root stem cell niche and the differentiation potential of a subset of stem cells in the Arabidopsis root. To obtain a comprehensive view of the SHR pathway, we used a statistical method called meta-analysis to combine the results of several microarray experiments measuring the changes in global expression profiles after modulating SHR activity. Meta-analysis was first used to identify the direct targets of SHR by combining results from an inducible form of SHR driven by its endogenous promoter, ectopic expression, followed by cell sorting and comparisons of mutant to wild-type roots. Eight putative direct targets of SHR were identified, all with expression patterns encompassing subsets of the native SHR expression domain. Further evidence for direct regulation by SHR came from binding of SHR in vivo to the promoter regions of four of the eight putative targets. A new role for SHR in the vascular cylinder was predicted from the expression pattern of several direct targets and confirmed with independent markers. The meta-analysis approach was then used to perform a global survey of the SHR indirect targets. Our analysis suggests that the SHR pathway regulates root development not only through a large transcription regulatory network but also through hormonal pathways and signaling pathways using receptor-like kinases. Taken together, our results not only identify the first nodes in the SHR pathway and a new function for SHR in the development of the vascular tissue but also reveal the global architecture of this developmental pathway

    A Quantitative and Dynamic Model for Plant Stem Cell Regulation

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    Plants maintain pools of totipotent stem cells throughout their entire life. These stem cells are embedded within specialized tissues called meristems, which form the growing points of the organism. The shoot apical meristem of the reference plant Arabidopsis thaliana is subdivided into several distinct domains, which execute diverse biological functions, such as tissue organization, cell-proliferation and differentiation. The number of cells required for growth and organ formation changes over the course of a plants life, while the structure of the meristem remains remarkably constant. Thus, regulatory systems must be in place, which allow for an adaptation of cell proliferation within the shoot apical meristem, while maintaining the organization at the tissue level. To advance our understanding of this dynamic tissue behavior, we measured domain sizes as well as cell division rates of the shoot apical meristem under various environmental conditions, which cause adaptations in meristem size. Based on our results we developed a mathematical model to explain the observed changes by a cell pool size dependent regulation of cell proliferation and differentiation, which is able to correctly predict CLV3 and WUS over-expression phenotypes. While the model shows stem cell homeostasis under constant growth conditions, it predicts a variation in stem cell number under changing conditions. Consistent with our experimental data this behavior is correlated with variations in cell proliferation. Therefore, we investigate different signaling mechanisms, which could stabilize stem cell number despite variations in cell proliferation. Our results shed light onto the dynamic constraints of stem cell pool maintenance in the shoot apical meristem of Arabidopsis in different environmental conditions and developmental states

    Accurate and versatile 3D segmentation of plant tissues at cellular resolution

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    Quantitative analysis of plant and animal morphogenesis requires accurate segmentation of individual cells in volumetric images of growing organs. In the last years, deep learning has provided robust automated algorithms that approach human performance, with applications to bio-image analysis now starting to emerge. Here, we present PlantSeg, a pipeline for volumetric segmentation of plant tissues into cells. PlantSeg employs a convolutional neural network to predict cell boundaries and graph partitioning to segment cells based on the neural network predictions. PlantSeg was trained on fixed and live plant organs imaged with confocal and light sheet microscopes. PlantSeg delivers accurate results and generalizes well across different tissues, scales, acquisition settings even on non plant samples. We present results of PlantSeg applications in diverse developmental contexts. PlantSeg is free and open-source, with both a command line and a user-friendly graphical interface

    Agri-Environmental Policy Measures in Israel: The Potential of Using Market-Oriented Instruments

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    This paper examines the possibilities of developing agri-environmental policy measures in Israel, focusing on market-oriented instruments. A conceptual framework for developing agri-environmental policy measures is presented, first in very broad lines (mandatory regulations, economic instruments and advisory measures) and subsequently focusing on economic instruments, and specifically, on market-oriented ones. Two criteria of choice between the measures are suggested: their contribution to improving the effectiveness of the policy; and the feasibility of their implementation. This is the framework used for analyzing agri-environmental measures in Israel. Israel currently implements a mix of mandatory regulations, economic instruments and advisory measures to promote the agri-environment. The use of additional economic instruments may improve the effectiveness of the policy. When comparing the effectiveness of various economic measures, we found that the feasibility of implementation of market-oriented instruments is greater, due to the Israeli public’s preference for strengthening market orientation in the agricultural sector. Four market-oriented instruments were practiced in a pilot project conducted in an Israeli rural area. We found that in this case study, the institutional feasibility and acceptance by stakeholders were the major parameters influencing the implementation of the market-oriented instruments, whereas the instruments’ contribution to enhancing the ecological or economic effectiveness were hardly considered by the stakeholders as arguments in favor of their use

    KIRMES: kernel-based identification of regulatory modules in euchromatic sequences

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    Motivation: Understanding transcriptional regulation is one of the main challenges in computational biology. An important problem is the identification of transcription factor (TF) binding sites in promoter regions of potential TF target genes. It is typically approached by position weight matrix-based motif identification algorithms using Gibbs sampling, or heuristics to extend seed oligos. Such algorithms succeed in identifying single, relatively well-conserved binding sites, but tend to fail when it comes to the identification of combinations of several degenerate binding sites, as those often found in cis-regulatory modules

    TOR acts as a metabolic gatekeeper for auxin-dependent lateral root initiation in Arabidopsis thaliana

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    Plant organogenesis requires matching the available metabolic resources to developmental programs. In Arabidopsis, the root system is determined by primary root-derived lateral roots (LRs), and adventitious roots (ARs) formed from non-root organs. Lateral root formation entails the auxin-dependent activation of transcription factors ARF7, ARF19, and LBD16. Adventitious root formation relies on LBD16 activation by auxin and WOX11. The allocation of shoot-derived sugar to the roots influences branching, but how its availability is sensed for LRs formation remains unknown. We combine metabolic profiling with cell-specific interference to show that LRs switch to glycolysis and consume carbohydrates. The target-of-rapamycin (TOR) kinase is activated in the lateral root domain. Interfering with TOR kinase blocks LR initiation while promoting AR formation. The target-of-rapamycin inhibition marginally affects the auxin-induced transcriptional response of the pericycle but attenuates the translation of ARF19, ARF7, and LBD16. TOR inhibition induces WOX11 transcription in these cells, yet no root branching occurs as TOR controls LBD16 translation. TOR is a central gatekeeper for root branching that integrates local auxin-dependent pathways with systemic metabolic signals, modulating the translation of auxin-induced genes.The authors gratefully acknowledge [...] the Cluster of Excellence Cellular Networks of the University of Heidelberg (CellNetworks) through grant EcTOP6 “Metabolism and Development” and the German Research Foundation (DFG) through grants INST 35/1314-1 FUGG, INST 35/1503-1 FUGG and FOR2581. Open Access funding enabled and organized by ProjektDEAL. Open Access funding enabled and organized by Projekt DEAL.Peer reviewe

    TOR acts as a metabolic gatekeeper for auxin-dependent lateral root initiation in Arabidopsis thaliana [Dataset]

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    Appendix Figure S1. Lateral root deficiency leads to starch hyperaccumulation in leaves. -- Appendix Figure S2. Comparable starch content in foliage at the end of the light period in plants producing LR or not. -- Appendix Figure S3. Glucose and Sucrose levels in shoots of IAA-treated Col-0 and slr seedlings. -- Appendix Figure S4. Auxin/slr-dependent signaling reconfigures the carbon metabolism-related transcriptome during LR formation is influenced. -- Appendix Figure S5. TOR over-activation leads to longer primary roots, whereas impairment of the TOR machinery results in reduced primary root length. -- Appendix Figure S6. Silencing efficiency in UB10pro>>amiR-TOR line. -- Appendix Figure S7. IAA or external carbohydrate sources in TOR-deficient seedlings can not rescue lateral root formation. -- Appendix Figure S8. Foliar accumulation of starch upon TOR silencing. -- Appendix Figure S9. Transcriptome analysis upon auxin-induced induction of lateral root formation in UB10pro>>amiR-TOR. -- Appendix Figure S10. Expression of TOR, GATA23, and ARF7 transcripts upon inhibition of TOR via AZD8055. -- Appendix Figure S11. IAA-responsive genes detected during ribosome profiling and TOR inhibition IAA induced in the RNA-seq experiment under TOR deficiency vastly overlap. -- Appendix Figure S12. WOX11 expression upon TOR-knockdown or TOR inhibition. -- Expanded view figures EV1-EV3. -- Dataset EV1. -- Dataset EV2.embj2022111273-sup-0001-appendix.pdfembj2022111273-sup-0002-evfigs.pdfembj2022111273-sup-0003-datasetev1.xlsxembj2022111273-sup-0004-datasetev2.xlsxembj2022111273-sup-0006-sdataev.zipPeer reviewe

    When the Choice Is Ours: Context and Agency Modulate the Neural Bases of Decision-Making

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    The option to choose between several courses of action is often associated with the feeling of being in control. Yet, in certain situations, one may prefer to decline such agency and instead leave the choice to others. In the present functional magnetic resonance imaging (fMRI) study, we provide evidence that the neural processes involved in decision-making are modulated not only by who controls our choice options (agency), but also by whether we have a say in who is in control (context). The fMRI results are noteworthy in that they reveal specific contributions of the anterior frontomedian cortex (viz. BA 10) and the rostral cingulate zone (RCZ) in decision-making processes. The RCZ is engaged when conditions clearly present us with the most choice options. BA 10 is engaged in particular when the choice is completely ours, as well as when it is completely up to others to choose for us which in turn gives rise to an attribution of control to oneself or someone else, respectively. After all, it does not only matter whether we have any options to choose from, but also who decides on that

    Regulation of plant stem cell quiescence by a brassinosteroid signaling module

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    Referred to by: Josep Vilarrasa-Blasi, Mary-Paz González-García, David Frigola, Norma Fàbregas-Vallvé, Konstantinos G. Alexiou, Nuria López-Bigas, Susana Rivas, Alain Jauneau, Jan U. Lohmann, Philip N. Benfey, Marta Ibañes, Ana I. Caño-Delgado Regulation of Plant Stem Cell Quiescence by a Brassinosteroid Signaling Module Developmental Cell, Volume 33, Issue 2, 20 April 2015, Pages 238.The quiescent center (QC) maintains the activity of the surrounding stem cells within the root stem cell niche, yet specific molecular players sustaining the low rate of QC cell division remain poorly understood. Here, we identified a R2R3-MYB transcription factor, BRAVO (BRASSINOSTEROIDS AT VASCULAR AND ORGANIZING CENTER), acting as a cell-specific repressor of QC divisions in the primary root of Arabidopsis. Ectopic BRAVO expression restricts overall root growth and ceases root regeneration upon damage of the stem cells, demonstrating the role of BRAVO in counteracting Brassinosteroid (BR)-mediated cell division in the QC cells. Interestingly, BR-regulated transcription factor BES1 (BRI1-EMS SUPRESSOR 1) directly represses and physically interacts with BRAVO in vivo, creating a switch that modulates QC divisions at the root stem cell niche. Together, our results define a mechanism for BR-mediated regulation of stem cell quiescence in plants.J.V.-B. and N.F.-V. are funded by FI PhD fellowship from the Generalitat de Catalunya (GC) in the A.I.C.-D. laboratory. J.V.-B. received a short-term fellowship (BE1-00924) in the Lohmann (J.U.L.) laboratory supported by the SFB873 of the DFG. Research by D.F and M.I. is funded by FIS2012-37655-C02-02 by the Spanish Ministry de Economy and Competitiveness and 2009SGR14 from GC, and D.F. has a PhD fellowship (FPU-AP2009-3736). S.R. is funded by the Laboratoire d’Excellence (LABEX) TULIP (ANR-10-LABX-41). M.-P.G.-G. received a “Juan de la Cierva” postdoctoral contract from the Spanish Ministry of Science in the Ana Caño (A.I.C.-D.) laboratory, and an HFSP short-term fellowship in the Benfey (P.N.B.) laboratory. P.N.B. is funded by NSF Arabidopsis 2010 grant. Work in the Ana Caño (A.I.C.-D.) laboratory is funded by a BIO2010/007 grant from the Spanish Ministry of Innovation and Science and a Marie-Curie Initial Training Network “BRAVISSIMO” (grant no. PITN-GA-2008-215118).Peer reviewe

    DEB025 (Alisporivir) Inhibits Hepatitis C Virus Replication by Preventing a Cyclophilin A Induced Cis-Trans Isomerisation in Domain II of NS5A

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    DEB025/Debio 025 (Alisporivir) is a cyclophilin (Cyp)-binding molecule with potent anti-hepatitis C virus (HCV) activity both in vitro and in vivo. It is currently being evaluated in phase II clinical trials. DEB025 binds to CypA, a peptidyl-prolyl cis-trans isomerase which is a crucial cofactor for HCV replication. Here we report that it was very difficult to select resistant replicons (genotype 1b) to DEB025, requiring an average of 20 weeks (four independent experiments), compared to the typically <2 weeks with protease or polymerase inhibitors. This indicates a high genetic barrier to resistance for DEB025. Mutation D320E in NS5A was the only mutation consistently selected in the replicon genome. This mutation alone conferred a low-level (3.9-fold) resistance. Replacing the NS5A gene (but not the NS5B gene) from the wild type (WT) genome with the corresponding sequence from the DEB025res replicon resulted in transfer of resistance. Cross-resistance with cyclosporine A (CsA) was observed, whereas NS3 protease and NS5B polymerase inhibitors retained WT-activity against DEB025res replicons. Unlike WT, DEB025res replicon replicated efficiently in CypA knock down cells. However, DEB025 disrupted the interaction between CypA and NS5A regardless of whether the NS5A protein was derived from WT or DEB025res replicon. NMR titration experiments with peptides derived from the WT or the DEB025res domain II of NS5A corroborated this observation in a quantitative manner. Interestingly, comparative NMR studies on two 20-mer NS5A peptides that contain D320 or E320 revealed a shift in population between the major and minor conformers. These data suggest that D320E conferred low-level resistance to DEB025 probably by reducing the need for CypA-dependent isomerisation of NS5A. Prolonged DEB025 treatment and multiple genotypic changes may be necessary to generate significant resistance to DEB025, underlying the high barrier to resistance
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