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Publikationen - Molekulare Signalverarbeitung

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Preprints

Niemeyer, M.; Moreno Castillo, E.; Ihling, C. H.; Iacobucci, C.; Wilde, V.; Hellmuth, A.; Hoehenwarter, W.; Samodelov, S. L.; Zurbriggen, M. D.; Kastritis, P. L.; Sinz, A.; Calderón Villalobos, L. I. A.; Flexibility of intrinsically disordered degrons in AUX/IAA proteins reinforces auxin receptor assemblies bioRxiv (2019) DOI: 10.1101/787770

Cullin RING-type E3 ubiquitin ligases SCFTIR1/AFB1-5 and their ubiquitylation targets, AUX/IAAs, sense auxin concentrations in the nucleus. TIR1 binds a surface-exposed degron in AUX/IAAs promoting their ubiquitylation and rapid auxin-regulated proteasomal degradation. Here, we resolved TIR1·auxin·IAA7 and TIR1·auxin·IAA12 complex topology, and show that flexible intrinsically disordered regions (IDRs) in the degron′s vicinity, cooperatively position AUX/IAAs on TIR1. The AUX/IAA PB1 interaction domain also assists in non-native contacts, affecting AUX/IAA dynamic interaction states. Our results establish a role for IDRs in modulating auxin receptor assemblies. By securing AUX/IAAs on two opposite surfaces of TIR1, IDR diversity supports locally tailored positioning for targeted ubiquitylation and might provide conformational flexibility for adopting a multiplicity of functional states. We postulate IDRs in distinct members of the AUX/IAA family to be an adaptive signature for protein interaction and initiation region for proteasome recruitment.
Publikation

Chutia, R.; Abel, S.; Ziegler, J.; Iron and Phosphate Deficiency Regulators Concertedly Control Coumarin Profiles in Arabidopsis thaliana Roots During Iron, Phosphate, and Combined Deficiencies Front. Plant Sci. 10, 113, (2019) DOI: 10.3389/fpls.2019.00113

Plants face varying nutrient conditions, to which they have to adapt to. Adaptive responses are nutrient-specific and strategies to ensure supply and homeostasis for one nutrient might be opposite to another one, as shown for phosphate (Pi) and iron (Fe) deficiency responses, where many genes are regulated in an opposing manner. This was also observed on the metabolite levels. Whereas root and exudate levels of catechol-type coumarins, phenylpropanoid-derived 2-benzopyranones, which facilitate Fe acquisition, are elevated after Fe deficiency, they are decreased after Pi deficiency. Exposing plants to combined Pi and Fe deficiency showed that the generation of coumarin profiles in Arabidopsis thaliana roots by Pi deficiency considerably depends on the availability of Fe. Similarly, the effect of Fe deficiency on coumarin profiles is different at low compared to high Pi availability. These findings suggest a fine-tuning of coumarin profiles, which depends on Fe and Pi availability. T-DNA insertion lines exhibiting aberrant expression of genes involved in the regulation of Pi starvation responses (PHO1, PHR1, bHLH32, PHL1, SPX1) and Fe starvation responses (BRUTUS, PYE, bHLH104, FIT) were used to analyze the regulation of the generation of coumarin profiles in Arabidopsis thaliana roots by Pi, Fe, and combined Pi and Fe deficiency. The analysis revealed a role of several Fe-deficiency response regulators in the regulation of Fe and of Pi deficiency-induced coumarin profiles as well as for Pi deficiency response regulators in the regulation of Pi and of Fe deficiency-induced coumarin profiles. Additionally, the regulation of Fe deficiency-induced coumarin profiles by Fe deficiency response regulators is influenced by Pi availability. Conversely, regulation of Pi deficiency-induced coumarin profiles by Pi deficiency response regulators is modified by Fe availability.
Publikation

Naumann, C.; Müller, J.; Sakhonwasee, S.; Wieghaus, A.; Hause, G.; Heisters, M.; Bürstenbinder, K.; Abel, S.; The Local Phosphate Deficiency Response Activates Endoplasmic Reticulum Stress-Dependent Autophagy Plant Physiol. 179, 460-476, (2019) DOI: 10.1104/pp.18.01379

Inorganic phosphate (Pi) is often a limiting plant nutrient. In members of the Brassicaceae family, such as Arabidopsis (Arabidopsis thaliana), Pi deprivation reshapes root system architecture to favor topsoil foraging. It does so by inhibiting primary root extension and stimulating lateral root formation. Root growth inhibition from phosphate (Pi) deficiency is triggered by iron-stimulated, apoplastic reactive oxygen species generation and cell wall modifications, which impair cell-to-cell communication and meristem maintenance. These processes require LOW PHOSPHATE RESPONSE1 (LPR1), a cell wall-targeted ferroxidase, and PHOSPHATE DEFICIENCY RESPONSE2 (PDR2), the single endoplasmic reticulum (ER)-resident P5-type ATPase (AtP5A), which is thought to control LPR1 secretion or activity. Autophagy is a conserved process involving the vacuolar degradation of cellular components. While the function of autophagy is well established under nutrient starvation (C, N, or S), it remains to be explored under Pi deprivation. Because AtP5A/PDR2 likely functions in the ER stress response, we analyzed the effect of Pi limitation on autophagy. Our comparative study of mutants defective in the local Pi deficiency response, ER stress response, and autophagy demonstrated that ER stress-dependent autophagy is rapidly activated as part of the developmental root response to Pi limitation and requires the genetic PDR2-LPR1 module. We conclude that Pi-dependent activation of autophagy in the root apex is a consequence of local Pi sensing and the associated ER stress response, rather than a means for systemic recycling of the macronutrient.
Publikation

Mitra, D.; Klemm, S.; Kumari, P.; Quegwer, J.; Möller, B.; Poeschl, Y.; Pflug, P.; Stamm, G.; Abel, S.; Bürstenbinder, K.; Microtubule-associated protein IQ67 DOMAIN5 regulates morphogenesis of leaf pavement cells in Arabidopsis thaliana J. Exp. Bot. 70, 529-543, (2019) DOI: 10.1093/jxb/ery395

Plant microtubules form a highly dynamic intracellular network with important roles for regulating cell division, cell proliferation and cell morphology. Its organization and dynamics are coordinated by various microtubule-associated proteins (MAPs) that integrate environmental and developmental stimuli to fine-tune and adjust cytoskeletal arrays. IQ67 DOMAIN (IQD) proteins recently emerged as a class of plant-specific MAPs with largely unknown functions. Here, using a reverse genetics approach, we characterize Arabidopsis IQD5 in terms of its expression domains, subcellular localization and biological functions. We show that IQD5 is expressed mostly in vegetative tissues, where it localizes to cortical microtubule arrays. Our phenotypic analysis of iqd5 loss-of-function lines reveals functions of IQD5 in pavement cell (PC) shape morphogenesis. Histochemical analysis of cell wall composition further suggests reduced rates of cellulose deposition in anticlinal cell walls, which correlate with reduced anisotropic expansion. Lastly, we demonstrate IQD5-dependent recruitment of calmodulin calcium sensors to cortical microtubule arrays and provide first evidence for important roles of calcium in regulation of PC morphogenesis. Our work thus identifies IQD5 as a novel player in PC shape regulation, and, for the first time, links calcium signaling to developmental processes that regulate anisotropic growth in PCs.
Bücher und Buchkapitel

Ziegler, J.; Hussain, H.; Neubert, R. H. H.; Abel, S.; Sensitive and Selective Amino Acid Profiling of Minute Tissue Amounts by HPLC/Electrospray Negative Tandem Mass Spectrometry Using 9-Fluorenylmethoxycarbonyl (Fmoc-Cl) Derivatization (Alterman, M. A., ed.). Methods Mol. Biol. 2030, 365-379, (2019) ISBN: 978-1-4939-9639-1 DOI: 10.1007/978-1-4939-9639-1_27

A method for selective and sensitive quantification of amino acids is described. The combination of established derivatization procedures of secondary and primary amino groups with 9-fluorenylmethoxycarbonyl chloride (Fmoc-Cl) and subsequent detection of derivatized amino acids by LC-ESI-MS/MS using multiple reaction monitoring provides high selectivity. The attachment of an apolar moiety enables purification of derivatized amino acids from matrix by a single solid-phase extraction step, which increases sensitivity by reduced ion suppression during LC-ESI-MS/MS detection. Additionally, chromatography of all amino acids can be performed on reversed-phase HPLC columns using eluents without additives, which are known to cause significant decreases in signal to noise ratios. The method has been routinely applied for amino acid profiling of low amounts of liquids and tissues of various origins with a sample throughput of about 50–100 samples a day. In addition to a detailed description of the method, some representative examples are presented.
Bücher und Buchkapitel

Möller, B.; Bürstenbinder, K.; Semi-Automatic Cell Segmentation from Noisy Image Data for Quantification of Microtubule Organization on Single Cell Level 199-203, (2019) ISBN: 978-1-5386-3641-1 DOI: 10.1109/ISBI.2019.8759145

The structure of the microtubule cytoskeleton provides valuable information related to morphogenesis of cells. The cytoskeleton organizes into diverse patterns that vary in cells of different types and tissues, but also within a single tissue. To assess differences in cytoskeleton organization methods are needed that quantify cytoskeleton patterns within a complete cell and which are suitable for large data sets. A major bottleneck in most approaches, however, is a lack of techniques for automatic extraction of cell contours. Here, we present a semi-automatic pipeline for cell segmentation and quantification of microtubule organization. Automatic methods are applied to extract major parts of the contours and a handy image editor is provided to manually add missing information efficiently. Experimental results prove that our approach yields high-quality contour data with minimal user intervention and serves a suitable basis for subsequent quantitative studies.
Bücher und Buchkapitel

Möller, B.; Zergiebel, L.; Bürstenbinder, K.; Quantitative and Comparative Analysis of Global Patterns of (Microtubule) Cytoskeleton Organization with CytoskeletonAnalyzer2D (Cvrčková, F. & Žárský, V., eds.). Methods Mol. Biol. 1992, 151-171, (2019) ISBN: 978-1-4939-9469-4 DOI: 10.1007/978-1-4939-9469-4_10

The microtubule cytoskeleton plays important roles in cell morphogenesis. To investigate the mechanisms of cytoskeletal organization, for example, during growth or development, in genetic studies, or in response to environmental stimuli, image analysis tools for quantitative assessment are needed. Here, we present a method for texture measure-based quantification and comparative analysis of global microtubule cytoskeleton patterns and subsequent visualization of output data. In contrast to other approaches that focus on the extraction of individual cytoskeletal fibers and analysis of their orientation relative to the growth axis, CytoskeletonAnalyzer2D quantifies cytoskeletal organization based on the analysis of local binary patterns. CytoskeletonAnalyzer2D thus is particularly well suited to study cytoskeletal organization in cells where individual fibers are difficult to extract or which lack a clearly defined growth axis, such as leaf epidermal pavement cells. The tool is available as ImageJ plugin and can be combined with publicly available software and tools, such as R and Cytoscape, to visualize similarity networks of cytoskeletal patterns.
Bücher und Buchkapitel

Möller, B.; Poeschl, Y.; Klemm, S.; Bürstenbinder, K.; Morphological Analysis of Leaf Epidermis Pavement Cells with PaCeQuant (Cvrčková, F. & Žárský, V., eds.). Methods Mol. Biol. 1992, 329-349, (2019) ISBN: 978-1-4939-9469-4 DOI: 10.1007/978-1-4939-9469-4_22

Morphological analysis of cell shapes requires segmentation of cell contours from input images and subsequent extraction of meaningful shape descriptors that provide the basis for qualitative and quantitative assessment of shape characteristics. Here, we describe the publicly available ImageJ plugin PaCeQuant and its associated R package PaCeQuantAna, which provides a pipeline for fully automatic segmentation, feature extraction, statistical analysis, and graphical visualization of cell shape properties. PaCeQuant is specifically well suited for analysis of jigsaw puzzle-like leaf epidermis pavement cells from 2D input images and supports the quantification of global, contour-based, skeleton-based, and pavement cell-specific shape descriptors.
Publikation

Wang, Q.; Grubb, C. D.; Abel, S.; Direct analysis of single leaf disks for chemopreventive glucosinolates Phytochem. Anal. 13, 152-157, (2002) DOI: 10.1002/pca.636

Natural isothiocyanates, produced during plant tissue damage from methionine‐derived glucosinolates, are potent inducers of mammalian phase 2 detoxification enzymes such as quinone reductase (QR). A greatly simplified bioassay for glucosinolates based on induction and colorimetric detection of QR activity in murine hepatoma cells is described. It is demonstrated that excised leaf disks of Arabidopsis thaliana (ecotype Columbia) can directly and reproducibly substitute for cell‐free leaf extracts as inducers of murine QR, which reduces sample preparation to a minimum and maximizes throughput. A comparison of 1 and 3 mm diameter leaf disks indicated that QR inducer potency was proportional to disk circumference (extent of tissue damage) rather than to area. When compared to the QR inducer potency of the corresponding amount of extract, 1 mm leaf disks were equally effective, whereas 3 mm disks were 70% as potent. The QR inducer potency of leaf disks correlated positively with the content of methionine‐derived glucosinolates, as shown by the analysis of wild‐type plants and mutant lines with lower or higher glucosinolate content. Thus, the microtitre plate‐based assay of single leaf disks provides a robust and inexpensive visual method for rapidly screening large numbers of plants in mapping populations or mutant collections and may be applicable to other glucosinolate‐producing species.
  • Die Leibniz-Gemeinschaft
  • Digitalisierung in der Landwirtschaft
  • Martin-Luther Universität Halle
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