Auxin fluxes through plasmodesmata modify root-tip auxin distribution

© 2020. Published by The Company of Biologists Ltd. Auxin is a key signal regulating plant growth and development. It is well established that auxin dynamics depend on the spatial distribution of efflux and influx carriers on the cell membranes. In this study, we employ a systems approach to characterise an alternative symplastic pathway for auxin mobilisation via plasmodesmata, which function as intercellular pores linking the cytoplasm of adjacent cells. To investigate the role of plasmodesmata in auxin patterning, we developed a multicellular model of the Arabidopsis root tip. We tested the model predictions using the DII-VENUS auxin response reporter, comparing the predicted and observed DII-VENUS distributions using genetic and chemical perturbations designed to affect both carrier-mediated and plasmodesmatal auxin fluxes. The model revealed that carrier-mediated transport alone cannot explain the experimentally determined auxin distribution in the root tip. In contrast, a composite model that incorporates both carrier-mediated and plasmodesmatal auxin fluxes re-capitulates the root-tip auxin distribution. We found that auxin fluxes through plasmodesmata enable auxin reflux and increase total root-tip auxin. We conclude that auxin fluxes through plasmodesmata modify the auxin distribution created by efflux and influx carriers

Volumetric Segmentation of Cell Cycle Markers in Confocal Images Using Machine Learning and Deep Learning

© Copyright © 2020 Khan, Voß, Pound and French. Understanding plant growth processes is important for many aspects of biology and food security. Automating the observations of plant development—a process referred to as plant phenotyping—is increasingly important in the plant sciences, and is often a bottleneck. Automated tools are required to analyze the data in microscopy images depicting plant growth, either locating or counting regions of cellular features in images. In this paper, we present to the plant community an introduction to and exploration of two machine learning approaches to address the problem of marker localization in confocal microscopy. First, a comparative study is conducted on the classification accuracy of common conventional machine learning algorithms, as a means to highlight challenges with these methods. Second, a 3D (volumetric) deep learning approach is developed and presented, including consideration of appropriate loss functions and training data. A qualitative and quantitative analysis of all the results produced is performed. Evaluation of all approaches is performed on an unseen time-series sequence comprising several individual 3D volumes, capturing plant growth. The comparative analysis shows that the deep learning approach produces more accurate and robust results than traditional machine learning. To accompany the paper, we are releasing the 4D point annotation tool used to generate the annotations, in the form of a plugin for the popular ImageJ (FIJI) software. Network models and example datasets will also be available online

AtMYB93 is a novel negative regulator of lateral root development in Arabidopsis

Plant root system plasticity is critical for survival in changing environmental conditions. One important aspect of root architecture is lateral root development, a complex process regulated by hormone, environmental and protein signalling pathways. Here we show, using molecular genetic approaches, that the MYB transcription factor AtMYB93 is a novel negative regulator of lateral root development in Arabidopsis. We identify AtMYB93 as an interaction partner of the lateral-root-promoting ARABIDILLO proteins. Atmyb93 mutants have faster lateral root developmental progression and enhanced lateral root densities, while AtMYB93-overexpressing lines display the opposite phenotype. AtMYB93 is expressed strongly, specifically and transiently in the endodermal cells overlying early lateral root primordia and is additionally induced by auxin in the basal meristem of the primary root. Furthermore, Atmyb93 mutant lateral root development is insensitive to auxin, indicating that AtMYB93 is required for normal auxin responses during lateral root development. We propose that AtMYB93 is part of a novel auxin-induced negative feedback loop stimulated in a select few endodermal cells early during lateral root development, ensuring that lateral roots only develop when absolutely required. Putative AtMYB93 homologues are detected throughout flowering plants and represent promising targets for manipulating root systems in diverse crop species

Dioxygenase-encoding AtDAO1 gene controls IAA oxidation and homeostasis in Arabidopsis

Auxin represents a key signal in plants, regulating almost every aspect of their growth and development. Major breakthroughs have been made dissecting the molecular basis of auxin transport, perception, and response. In contrast, how plants control the metabolism and homeostasis of the major form of auxin in plants, indole-3-acetic acid (IAA), remains unclear. In this paper, we initially describe the function of the Arabidopsis thaliana gene DIOXYGENASE FOR AUXIN OXIDATION 1 (AtDAO1). Transcriptional and translational reporter lines revealed that AtDAO1 encodes a highly root expressed, cytoplasmically localized IAA oxidase. Stable isotope-labeled IAA feeding studies of loss and gain of function AtDAO1 lines showed that this oxidase represents the major regulator of auxin degradation to 2-oxoindole 3-acetic acid (oxIAA) in Arabidopsis. Surprisingly, AtDAO1 loss and gain of function lines exhibited relatively subtle auxin-related phenotypes, such as altered root hair length. Metabolite profiling of mutant lines revealed that disrupting AtDAO1 regulation resulted in major changes in steady-state levels of oxIAA and IAA conjugates but not IAA. Hence, IAA conjugation and catabolism seem to regulate auxin levels in Arabidopsis in a highly redundant manner. We observed that transcripts of AtDOA1 IAA oxidase and GH3 IAA-conjugating enzymes are auxin-inducible, providing a molecular basis for their observed functional redundancy. We conclude that the AtDAO1 gene plays a key role regulating auxin homeostasis in Arabidopsis, acting in concert with GH3 genes, to maintain auxin concentration at optimal levels for plant growth and development

Systems approaches reveal that ABCB and PIN proteins mediate co-dependent auxin efflux

Members of the B family of membrane-bound ATP-binding cassette (ABC) transporters represent key components of the auxin-efflux machinery in plants. Over the last two decades experimental studies have shown that modifying ABCB expression affects auxin distribution and plant phenotypes. However, precisely how ABCB proteins transport auxin in conjunction with the more widely studied family of PIN-formed (PIN) auxin efflux transporters is unclear, and studies using heterologous systems have produced conflicting results.Here, we integrate ABCB localization data into a multicellular model of auxin transport in the Arabidopsis thaliana root tip to predict how ABCB-mediated auxin transport impacts organ-scale auxin distribution. We use our model to test five potential ABCB–PIN regulatory interactions, simulating the auxin dynamics for each interaction and quantitatively comparing the predictions with experimental images of the DII-VENUS auxin reporter in wild type and abcb single and double loss-of-function mutants. Only specific ABCB–PIN regulatory interactions result in predictions that recreate the experimentally observed DII-VENUS distributions and long-distance auxin transport. Our results suggest that ABCBs enable auxin efflux independently of PINs; however, PIN-mediated auxin efflux is predominantly through a co-dependent efflux where co-localised with ABCBs

Einführung des cloudbasierten Bibliothekssystems Alma in Berlin – ein Erfahrungsbericht

Die enge Zusammenarbeit der vier Berliner Universitätsbibliotheken [Freie Universität Berlin (FU), Humboldt-Universität zu Berlin (HU), Technische Universität Berlin (TU), Universität der Künste Berlin (UdK)] reicht weit zurück. Bereits vor der Jahrtausendwende haben die Berliner Universitätsbibliotheken (UBs) gemeinsam das Bibliothekssystem Aleph 500 ausgewählt und implementiert, danach folgten weitere Systeme – das Linking System SFX, die Digitale Bibliothek MetaLib und das Bibliotheksportal Primo. Es war daher folgerichtig und selbstverständlich, dass auch die Auswahl und Implementierung eines neuen Bibliothekssystems in enger Abstimmung und Zusammenarbeit erfolgte. Die Erfahrungen bei Vertragsverhandlungen und Implementierung von Alma sind Gegenstand des folgenden Berichtes.As regards implementing new library technology, the Berlin University libraries have been working closely together for more than 20 years. It was the case for the implementation of the legacy system Aleph 500, the linking system SFX, the digital library MetaLib and the library portal Primo, and therefore it was a matter of course to continue the close cooperation during the implementation of the new cloud-based library system, too. The experience gained during the contract negotiations and the implementation project, and lessons learned are the focus of this report.Peer Reviewe

Dual expression and anatomy lines allow simultaneous visualization of gene expression and anatomy

Studying the developmental genetics of plant organs, requires following gene expression in specific tissues. To facilitate this, we have developed the Dual Expression Anatomy Lines (DEAL), which incorporate a red plasma membrane marker alongside a fluorescent reporter for a gene of interest in the same vector. Here, we adapted the GreenGate cloning vectors to create two destination vectors showing strong marking of cell membranes in either the whole root or specifically in the lateral roots. This system can also be used in both embryos and whole seedlings. As proof of concept, we follow both gene expression and anatomy in Arabidopsis (Arabidopsis thaliana) during lateral root organogenesis for a period of over 24h,. and cCoupled with the development of a flow cell and perfusion system, we follow changes in activity of the DII auxin sensor following application of auxin

Rye Bread Defects: Analysis of Composition and Further Influence Factors as Determinants of Dry-Baking

For decades, the evaluation of rye milling products have been aimed at detecting raw material defects that are linked to excessive enzyme activity. Those defects were indirectly characterized by the rheological methods of the dough or the final products. However, such methods do not sufficiently reflect the baking properties of all rye flours present on the market. A further problem is the continuing climate change, which affects compound composition in rye. So far, these bread defects can only be corrected by process engineering (e.g., extended dough resting). Therefore, it is necessary to characterize the main determinants of the quality defects prior to the baking process in order to predict baking quality and not waste raw material, energy, and time. In this study, it was found that the water accessibility of starch for gelatinization and its partial inhibition by certain components play a major role in baking quality. Specifically, high amounts of insoluble nonstarch-polysaccharides (NSPSs) and a hindered denaturation of proteins seem to be an indication and reason for poor baking quality. However, traditional quantitative analysis of the ingredients and properties of the rye milling products (e.g., falling number, protein content, amylographic data) does not allow any reliable conclusions about rye flour suitability for use as bread rye. It can be concluded that more complex compositional aspects (e.g., complexation of compounds) need to be characterized for future quality control of rye.TU Berlin, Open-Access-Mittel – 202