24 results on '"Lucyshyn D"'
Search Results
2. Determination of masked mycotoxins using HPLC–tandem mass spectrometry
- Author
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Berthiller, F., primary, Schuhmacher, R., additional, Poppenberger, B., additional, Lucyshyn, D., additional, Lemmens, M., additional, Adam, G., additional, and Krska, R., additional
- Published
- 2006
- Full Text
- View/download PDF
3. First results of GEN-AU: Cloning of Deoxynivalenol- and Zearalenone-inactivating UDP-glucosyltransferase genes from Arabidopsis thaliana and expression in yeast for production of mycotoxin-glucosides
- Author
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Poppenberger, B., Berthiller, F., Lucyshyn, D., Schuhmacher, R., Krska, R., and Adam, G.
- Published
- 2005
- Full Text
- View/download PDF
4. Root Hair Imaging Using Confocal Microscopy.
- Author
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Mutanwad KV, Debreczeny M, and Lucyshyn D
- Subjects
- Plants, Genetically Modified genetics, Meristem growth & development, Meristem genetics, Microscopy, Confocal methods, Plant Roots growth & development, Plant Roots genetics, Plant Roots cytology, Arabidopsis genetics
- Abstract
Plant genetics plays a key role in determining root hair initiation and development. A complex network of genetic interactions therefore closely monitors and influences root hair phenotype and morphology. The significance of these genes can be studied by employing, for instance, loss-of-function mutants, overexpression plant lines, and fluorescently labeled constructs. Confocal laser scanning microscopy is a great tool to visually observe and document these morphological features. This chapter elaborates the techniques involved in handling of microscopic setup to acquire images displaying root hair distribution along the fully elongated zone of Arabidopsis thaliana roots. Additionally, we illustrate an approach to visualize early fate determination of epidermal cells in the root apical meristem, by describing a method for imaging YFP tagged transgenic plant lines., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)
- Published
- 2024
- Full Text
- View/download PDF
5. Balancing O-GlcNAc and O-fucose in plants.
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Mutanwad KV and Lucyshyn D
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- Acetylglucosamine metabolism, Animals, Cell Nucleus metabolism, Glycosylation, Protein Processing, Post-Translational, Signal Transduction, Fucose metabolism, N-Acetylglucosaminyltransferases genetics, N-Acetylglucosaminyltransferases metabolism
- Abstract
O-linked modification of nuclear and cytosolic proteins with monosaccharides is essential in all eukaryotes. While many aspects of this post-translational modification are highly conserved, there are striking differences between plants and the animal kingdom. In animals, dynamic cycling of O-GlcNAc is established by two essential single copy enzymes, the O-GlcNAc transferase OGT and O-GlcNAc hydrolase OGA. In contrast, plants balance O-GlcNAc with O-fucose modifications, catalyzed by the OGT SECRET AGENT (SEC) and the protein O-fucosyltransferase (POFUT) SPINDLY (SPY). However, specific glycoside hydrolases for either of the two modifications have not yet been identified. Nucleocytoplasmic O-glycosylation is still not very well understood in plants, even though a high number of proteins were found to be affected. One important open question is how specificity is established in a system where only two enzymes modify hundreds of proteins. Here, we discuss the possibility that O-GlcNAc- and O-fucose-binding proteins could introduce an additional flexible layer of regulation in O-glycosylation-mediated signaling pathways, with the potential of integrating internal or external signals., (© 2021 The Authors. The FEBS Journal published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)
- Published
- 2022
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6. Author Correction: Light triggers PILS-dependent reduction in nuclear auxin signalling for growth transition.
- Author
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Béziat C, Barbez E, Feraru MI, Lucyshyn D, and Kleine-Vehn J
- Abstract
A Correction to this paper has been published: https://doi.org/10.1038/s41477-021-00924-y.
- Published
- 2021
- Full Text
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7. Retraction Note: PPP1, a plant-specific regulator of transcription controls Arabidopsis development and PIN expression.
- Author
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Benjamins R, Barbez E, Ortbauer M, Terpstra I, Lucyshyn D, Moulinier-Anzola J, Khan MA, Leitner J, Malenica N, Butt H, Korbei B, Scheres B, Kleine-Vehn J, and Luschnig C
- Published
- 2021
- Full Text
- View/download PDF
8. Targeting alternative splicing by RNAi: from the differential impact on splice variants to triggering artificial pre-mRNA splicing.
- Author
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Fuchs A, Riegler S, Ayatollahi Z, Cavallari N, Giono LE, Nimeth BA, Mutanwad KV, Schweighofer A, Lucyshyn D, Barta A, Petrillo E, and Kalyna M
- Subjects
- Arabidopsis Proteins biosynthesis, Exons, Genes, Plant, HeLa Cells, Humans, MicroRNAs genetics, Plants, Genetically Modified, Protein Isoforms biosynthesis, Protoplasts metabolism, RNA Precursors genetics, RNA Processing, Post-Transcriptional, RNA, Plant genetics, Serine-Arginine Splicing Factors biosynthesis, Serine-Arginine Splicing Factors genetics, Transcription, Genetic, Transfection, Alternative Splicing genetics, Arabidopsis genetics, Arabidopsis Proteins genetics, Gene Expression Regulation, Plant genetics, Gene Knockdown Techniques, Nonsense Mediated mRNA Decay, Protein Isoforms genetics, RNA Interference, RNA Precursors metabolism, RNA, Plant metabolism
- Abstract
Alternative splicing generates multiple transcript and protein isoforms from a single gene and controls transcript intracellular localization and stability by coupling to mRNA export and nonsense-mediated mRNA decay (NMD). RNA interference (RNAi) is a potent mechanism to modulate gene expression. However, its interactions with alternative splicing are poorly understood. We used artificial microRNAs (amiRNAs, also termed shRNAmiR) to knockdown all splice variants of selected target genes in Arabidopsis thaliana. We found that splice variants, which vary by their protein-coding capacity, subcellular localization and sensitivity to NMD, are affected differentially by an amiRNA, although all of them contain the target site. Particular transcript isoforms escape amiRNA-mediated degradation due to their nuclear localization. The nuclear and NMD-sensitive isoforms mask RNAi action in alternatively spliced genes. Interestingly, Arabidopsis SPL genes, which undergo alternative splicing and are targets of miR156, are regulated in the same manner. Moreover, similar results were obtained in mammalian cells using siRNAs, indicating cross-kingdom conservation of these interactions among RNAi and splicing isoforms. Furthermore, we report that amiRNA can trigger artificial alternative splicing, thus expanding the RNAi functional repertoire. Our findings unveil novel interactions between different post-transcriptional processes in defining transcript fates and regulating gene expression., (© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.)
- Published
- 2021
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9. The Arabidopsis O-fucosyltransferase SPINDLY regulates root hair patterning independently of gibberellin signaling.
- Author
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Mutanwad KV, Zangl I, and Lucyshyn D
- Subjects
- Arabidopsis genetics, Arabidopsis Proteins genetics, DNA-Binding Proteins genetics, DNA-Binding Proteins metabolism, Gene Expression Regulation, Plant, Glycosylation, Homeodomain Proteins genetics, Homeodomain Proteins metabolism, Plant Roots genetics, Repressor Proteins genetics, Signal Transduction, Arabidopsis metabolism, Arabidopsis Proteins metabolism, Gibberellins metabolism, Plant Roots metabolism, Repressor Proteins metabolism
- Abstract
Root hairs are able to sense soil composition and play an important role in water and nutrient uptake. In Arabidopsis thaliana , root hairs are distributed in the epidermis in a specific pattern, regularly alternating with non-root hair cells in continuous cell files. This patterning is regulated by internal factors such as a number of hormones, as well as by external factors like nutrient availability. Thus, root hair patterning is an excellent model for studying the plasticity of cell fate determination in response to environmental changes. Here, we report that loss-of-function mutants for the Protein O-fucosyltransferase SPINDLY (SPY) show defects in root hair patterning. Using transcriptional reporters, we show that patterning in spy-22 is affected upstream of GLABRA2 (GL2) and WEREWOLF (WER). O-fucosylation of nuclear and cytosolic proteins is an important post-translational modification that is still not very well understood. So far, SPY is best characterized for its role in gibberellin signaling via fucosylation of the growth-repressing DELLA protein REPRESSOR OF ga1-3 (RGA). Our data suggest that the epidermal patterning defects in spy-22 are independent of RGA and gibberellin signaling., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2020. Published by The Company of Biologists Ltd.)
- Published
- 2020
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10. Direct Control of SPEECHLESS by PIF4 in the High-Temperature Response of Stomatal Development.
- Author
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Lau OS, Song Z, Zhou Z, Davies KA, Chang J, Yang X, Wang S, Lucyshyn D, Tay IHZ, Wigge PA, and Bergmann DC
- Subjects
- Arabidopsis metabolism, Arabidopsis Proteins physiology, Basic Helix-Loop-Helix Transcription Factors genetics, Basic Helix-Loop-Helix Transcription Factors physiology, Cell Differentiation, Cell Lineage, Gene Expression Regulation, Plant genetics, Hot Temperature, Phytochrome metabolism, Plant Development, Plant Stomata physiology, Signal Transduction, Temperature, Transcription Factors metabolism, Arabidopsis Proteins metabolism, Basic Helix-Loop-Helix Transcription Factors metabolism, Plant Stomata metabolism
- Abstract
Environmental factors shape the phenotypes of multicellular organisms. The production of stomata-the epidermal pores required for gas exchange in plants-is highly plastic and provides a powerful platform to address environmental influence on cell differentiation [1-3]. Rising temperatures are already impacting plant growth, a trend expected to worsen in the near future [4]. High temperature inhibits stomatal production, but the underlying mechanism is not known [5]. Here, we show that elevated temperature suppresses the expression of SPEECHLESS (SPCH), the basic-helix-loop-helix (bHLH) transcription factor that serves as the master regulator of stomatal lineage initiation [6, 7]. Our genetic and expression analyses indicate that the suppression of SPCH and stomatal production is mediated by the bHLH transcription factor PHYTOCHROME-INTERACTING FACTOR 4 (PIF4), a core component of high-temperature signaling [8]. Importantly, we demonstrate that, upon exposure to high temperature, PIF4 accumulates in the stomatal precursors and binds to the promoter of SPCH. In addition, we find SPCH feeds back negatively to the PIF4 gene. We propose a model where warm-temperature-activated PIF4 binds and represses SPCH expression to restrict stomatal production at elevated temperatures. Our work identifies a molecular link connecting high-temperature signaling and stomatal development and reveals a direct mechanism by which production of a specific cell lineage can be controlled by a broadly expressed environmental signaling factor., (Copyright © 2018 Elsevier Ltd. All rights reserved.)
- Published
- 2018
- Full Text
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11. Sudden acquired retinal degeneration syndrome in western Canada: 93 cases.
- Author
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Leis ML, Lucyshyn D, Bauer BS, Grahn BH, and Sandmeyer LS
- Subjects
- Animals, Blindness epidemiology, Blindness veterinary, Canada epidemiology, Dog Diseases etiology, Dogs, Female, Male, Prevalence, Retinal Degeneration epidemiology, Syndrome, Dog Diseases epidemiology, Retinal Degeneration veterinary
- Abstract
This study reviewed clinical data from dogs diagnosed with sudden acquired retinal degeneration syndrome (SARDS) in western Canada. Medical records from the Western College of Veterinary Medicine from 2002 to 2016 showed that 93 cases of SARDS were diagnosed based on presentation for sudden blindness and a bilaterally extinguished electroretinogram. The most common pure breeds were the miniature schnauzer, dachshund, and pug. The mean age at diagnosis was 8.1 years and males and females were equally affected. Most of the dogs were presented with normal non-chromatic, but abnormal chromatic pupillary light reflexes. The incidence of retinal degeneration as detected via ophthalmoscopy increased over time after SARDS diagnosis. Polyuria, polydipsia, polyphagia, weight gain, elevated liver enzyme values, isosthenuria, and proteinuria were common clinical and laboratory findings. Chromatic pupillary light reflex testing may be more valuable than non-chromatic pupillary light testing in detecting pupil response abnormalities in dogs with SARDS, although electroretinography remains the definitive diagnostic test., Competing Interests: Use of this article is limited to a single copy for personal study. Anyone interested in obtaining reprints should contact the CVMA office (hbroughton@cvma-acmv.org) for additional copies or permission to use this material elsewhere.
- Published
- 2017
12. Light triggers PILS-dependent reduction in nuclear auxin signalling for growth transition.
- Author
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Béziat C, Barbez E, Feraru MI, Lucyshyn D, and Kleine-Vehn J
- Subjects
- Arabidopsis growth & development, Arabidopsis Proteins genetics, Arabidopsis Proteins metabolism, Cell Nucleus metabolism, Cell Nucleus radiation effects, Gene Expression Regulation, Plant, Phytochrome B metabolism, Arabidopsis metabolism, Arabidopsis radiation effects, Arabidopsis Proteins physiology, Indoleacetic Acids metabolism, Light, Plant Growth Regulators metabolism, Signal Transduction radiation effects
- Abstract
The phytohormone auxin induces or represses growth depending on its concentration and the underlying tissue type. However, it remains unknown how auxin signalling is modulated to allow tissues transiting between repression and promotion of growth. Here, we used apical hook development as a model for growth transitions in plants. A PIN-FORMED (PIN)-dependent intercellular auxin transport module defines an auxin maximum that is causal for growth repression during the formation of the apical hook. Our data illustrate that growth transition for apical hook opening is largely independent of this PIN module, but requires the PIN-LIKES (PILS) putative auxin carriers at the endoplasmic reticulum. PILS proteins reduce nuclear auxin signalling in the apical hook, leading to the de-repression of growth and the onset of hook opening. We also show that the phytochrome (phy) B-reliant light-signalling pathway directly regulates PILS gene activity, thereby enabling light perception to repress nuclear auxin signalling and to control growth. We propose a novel mechanism, in which PILS proteins allow external signals to alter tissue sensitivity to auxin, defining differential growth rates.
- Published
- 2017
- Full Text
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13. Studying Transcription Factor Binding to Specific Genomic Loci by Chromatin Immunoprecipitation (ChIP).
- Author
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Kumar SV and Lucyshyn D
- Subjects
- Binding Sites physiology, Chromatin Immunoprecipitation methods, Gene Expression Regulation physiology, Genomics methods, Chromatin metabolism, DNA metabolism, Plant Growth Regulators metabolism, Plant Proteins metabolism, Protein Binding physiology, Transcription Factors metabolism
- Abstract
Plant hormone signaling involves complex transcriptional networks, where transcription factors orchestrate the control of specific gene expression. These networks include cross talk between hormone signaling pathways, and the integration of environmental signals and the developmental program. Understanding how particular transcription factors respond and integrate specific signals is crucial in order to understand the basic mechanisms of hormonal signaling and cross talk. Studying transcription factor binding at specific genomic loci by chromatin immunoprecipitation (ChIP) is therefore a valuable technique in order to analyze transcriptional regulation. The method is based on cross-linking proteins to DNA, the isolation of chromatin, and immunoprecipitation of a transcription factor of interest. The attached DNA is then recovered and analyzed by quantitative real-time PCR in order to establish binding sites of the respective transcription factor. Here, we present a relatively simple and short protocol for ChIP on single loci.
- Published
- 2017
- Full Text
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14. PPP1, a plant-specific regulator of transcription controls Arabidopsis development and PIN expression.
- Author
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Benjamins R, Barbez E, Ortbauer M, Terpstra I, Lucyshyn D, Moulinier-Anzola J, Khan MA, Leitner J, Malenica N, Butt H, Korbei B, Scheres B, Kleine-Vehn J, and Luschnig C
- Subjects
- Arabidopsis Proteins chemistry, Binding Sites, Cell Nucleus metabolism, Computer Simulation, Cytoplasm metabolism, DNA-Binding Proteins chemistry, Gene Expression Regulation, Plant, Meristem physiology, Phylogeny, Plants, Genetically Modified, Promoter Regions, Genetic, Protein Domains, RNA-Binding Proteins chemistry, Arabidopsis genetics, Arabidopsis growth & development, Arabidopsis Proteins genetics, Arabidopsis Proteins metabolism, DNA-Binding Proteins genetics, DNA-Binding Proteins metabolism, RNA-Binding Proteins genetics, RNA-Binding Proteins metabolism
- Abstract
Directional transport of auxin is essential for plant development, with PIN auxin transport proteins representing an integral part of the machinery that controls hormone distribution. However, unlike the rapidly emerging framework of molecular determinants regulating PIN protein abundance and subcellular localization, insights into mechanisms controlling PIN transcription are still limited. Here we describe PIN2 PROMOTER BINDING PROTEIN 1 (PPP1), an evolutionary conserved plant-specific DNA binding protein that acts on transcription of PIN genes. Consistent with PPP1 DNA-binding activity, PPP1 reporter proteins are nuclear localized and analysis of PPP1 null alleles and knockdown lines indicated a function as a positive regulator of PIN expression. Furthermore, we show that ppp1 pleiotropic mutant phenotypes are partially reverted by PIN overexpression, and results are presented that underline a role of PPP1-PIN promoter interaction in PIN expression control. Collectively, our findings identify an elementary, thus far unknown, plant-specific DNA-binding protein required for post-embryonic plant development, in general, and correct expression of PIN genes, in particular.
- Published
- 2016
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15. Spring loading a pre-cleavage intermediate for hairpin telomere formation.
- Author
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Lucyshyn D, Huang SH, and Kobryn K
- Subjects
- Bacterial Proteins chemistry, Bacterial Proteins genetics, Catalytic Domain genetics, Cold Temperature, DNA chemistry, DNA metabolism, DNA Cleavage, DNA Replication, Endodeoxyribonucleases chemistry, Endodeoxyribonucleases genetics, Models, Genetic, Mutation, Nucleic Acid Conformation, Recombinases chemistry, Recombinases genetics, Bacterial Proteins metabolism, Endodeoxyribonucleases metabolism, Recombinases metabolism, Telomere chemistry, Telomere metabolism
- Abstract
The Borrelia telomere resolvase, ResT, forms the unusual hairpin telomeres of the linear Borrelia replicons in a process referred to as telomere resolution. Telomere resolution is a DNA cleavage and rejoining reaction that proceeds from a replicated telomere intermediate in a reaction with mechanistic similarities to that catalyzed by type IB topoisomerases. Previous reports have implicated the hairpin-binding module, at the end of the N-terminal domain of ResT, in distorting the DNA between the scissile phosphates so as to promote DNA cleavage and hairpin formation by the catalytic domain. We report that unwinding the DNA between the scissile phosphates, prior to DNA cleavage, is a key cold-sensitive step in telomere resolution. Through the analysis of ResT mutants, rescued by substrate modifications that mimic DNA unwinding between the cleavage sites, we show that formation and/or stabilization of an underwound pre-cleavage intermediate depends upon cooperation of the hairpin-binding module and catalytic domain. The phenotype of the mutants argues that the pre-cleavage intermediate promotes strand ejection to favor the forward reaction and that subsequent hairpin capture is a reversible reaction step. These reaction features are proposed to promote hairpin formation over strand resealing while allowing reversal back to substrate of aborted reactions., (© The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.)
- Published
- 2015
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16. Meta-regulation of Arabidopsis auxin responses depends on tRNA maturation.
- Author
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Leitner J, Retzer K, Malenica N, Bartkeviciute R, Lucyshyn D, Jäger G, Korbei B, Byström A, and Luschnig C
- Subjects
- Arabidopsis genetics, Arabidopsis growth & development, Arabidopsis Proteins genetics, Membrane Transport Proteins genetics, Membrane Transport Proteins metabolism, RNA-Binding Proteins genetics, Arabidopsis metabolism, Arabidopsis Proteins metabolism, Gene Expression Regulation, Developmental, Gene Expression Regulation, Plant, Indoleacetic Acids metabolism, RNA Processing, Post-Transcriptional, RNA, Transfer metabolism, RNA-Binding Proteins metabolism
- Abstract
Polar transport of the phytohormone auxin throughout plants shapes morphogenesis and is subject to stringent and specific control. Here, we identify basic cellular activities connected to translational control of gene expression as sufficient to specify auxin-mediated development. Mutants in subunits of Arabidopsis Elongator, a protein complex modulating translational efficiency via maturation of tRNAs, exhibit defects in auxin-controlled developmental processes, associated with reduced abundance of PIN-formed (PIN) auxin transport proteins. Similar anomalies are observed upon interference with tRNA splicing by downregulation of RNA ligase (AtRNL), pointing to a general role of tRNA maturation in auxin signaling. Elongator Protein 6 (ELP6) and AtRNL expression patterns underline an involvement in adjusting PIN protein levels, whereas rescue of mutant defects by auxin indicates rate-limiting activities in auxin-controlled organogenesis. This emphasizes mechanisms in which auxin serves as a bottleneck for plant morphogenesis, translating common cellular activities into defined developmental readouts., (Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.)
- Published
- 2015
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17. Arabidopsis TOL proteins act as gatekeepers for vacuolar sorting of PIN2 plasma membrane protein.
- Author
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Korbei B, Moulinier-Anzola J, De-Araujo L, Lucyshyn D, Retzer K, Khan MA, and Luschnig C
- Subjects
- Arabidopsis Proteins genetics, Arabidopsis Proteins metabolism, Protein Transport, Arabidopsis metabolism, Arabidopsis Proteins physiology, Vacuoles metabolism
- Abstract
Controlling variations in plasma membrane (PM) protein abundance is of utmost importance for development in higher plants. For modulating PM protein activity, endocytosed proteins can be either cycled between PM and endosomes or sorted for their irreversible inactivation to lysosomes/vacuoles. Cargo ubiquitination triggers vacuolar delivery for degradation, which is controlled by Endosomal Sorting Complex Required for Transport (ESCRT). Essential parts of this machinery are conserved across kingdoms, but determinants liable for initial recognition and concentration of ubiquitinated cargo have not been identified in plants. Here, we describe members of an Arabidopsis TOL (TOM1-LIKE) family as ubiquitin binding proteins that act redundantly in control of plant morphogenesis. Specifically, tol mutant combinations exhibit defects that reflect alterations in responses mediated by the phytohormone auxin. Consistently, we provide evidence for a role of TOLs in recognition and further endocytic sorting of a PIN-FORMED (PIN)-type auxin carrier protein at the PM, modulating dynamic auxin distribution and associated growth responses. Such TOL-dependent vacuolar sorting depends on cargo ubiquitination and coincides with dynamic rearrangements in TOL distribution. Collectively, these findings lead us to suggest a function for TOLs early in the passage of endocytosed ubiquitinated PM cargo, acting as gatekeepers for degradative protein sorting to the vacuole., (Copyright © 2013 Elsevier Ltd. All rights reserved.)
- Published
- 2013
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18. Transcription factor PIF4 controls the thermosensory activation of flowering.
- Author
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Kumar SV, Lucyshyn D, Jaeger KE, Alós E, Alvey E, Harberd NP, and Wigge PA
- Subjects
- Arabidopsis Proteins genetics, Basic Helix-Loop-Helix Transcription Factors genetics, Gene Expression Regulation, Plant, Photoperiod, Plant Leaves metabolism, Promoter Regions, Genetic genetics, Signal Transduction, Time Factors, Arabidopsis growth & development, Arabidopsis metabolism, Arabidopsis Proteins metabolism, Basic Helix-Loop-Helix Transcription Factors metabolism, Flowers growth & development, Flowers metabolism, Temperature
- Abstract
Plant growth and development are strongly affected by small differences in temperature. Current climate change has already altered global plant phenology and distribution, and projected increases in temperature pose a significant challenge to agriculture. Despite the important role of temperature on plant development, the underlying pathways are unknown. It has previously been shown that thermal acceleration of flowering is dependent on the florigen, FLOWERING LOCUS T (FT). How this occurs is, however, not understood, because the major pathway known to upregulate FT, the photoperiod pathway, is not required for thermal acceleration of flowering. Here we demonstrate a direct mechanism by which increasing temperature causes the bHLH transcription factor PHYTOCHROME INTERACTING FACTOR4 (PIF4) to activate FT. Our findings provide a new understanding of how plants control their timing of reproduction in response to temperature. Flowering time is an important trait in crops as well as affecting the life cycles of pollinator species. A molecular understanding of how temperature affects flowering will be important for mitigating the effects of climate change.
- Published
- 2012
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19. Plant development: PIF4 integrates diverse environmental signals.
- Author
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Lucyshyn D and Wigge PA
- Subjects
- Acclimatization, Animals, Arabidopsis genetics, Arabidopsis growth & development, Arabidopsis Proteins genetics, Arabidopsis Proteins metabolism, Basic Helix-Loop-Helix Transcription Factors genetics, Basic Helix-Loop-Helix Transcription Factors metabolism, Basic Helix-Loop-Helix Transcription Factors physiology, Circadian Rhythm, Environment, Helix-Loop-Helix Motifs, Homeostasis, Life Cycle Stages, Signal Transduction, Transcription Factors physiology, Arabidopsis physiology
- Abstract
Flexible adaptation to environmental changes is essential for plants. Recent studies suggest that a group of basic helix-loop-helix transcription factors play a central role in the crosstalk between environmental cues and hormone signalling.
- Published
- 2009
- Full Text
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20. Engineered bakers yeast as a sensitive bioassay indicator organism for the trichothecene toxin deoxynivalenol.
- Author
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Abolmaali S, Mitterbauer R, Spadiut O, Peruci M, Weindorfer H, Lucyshyn D, Ellersdorfer G, Lemmens M, Moll WD, and Adam G
- Subjects
- ATP-Binding Cassette Transporters genetics, Acetyltransferases genetics, Bacteria metabolism, Endopeptidases genetics, Gene Deletion, Inhibitory Concentration 50, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae growth & development, Saccharomyces cerevisiae Proteins genetics, Sensitivity and Specificity, Trichothecenes metabolism, Ubiquitin C genetics, Microbiological Techniques, Saccharomyces cerevisiae drug effects, Trichothecenes toxicity
- Abstract
The aim of this study was to increase the sensitivity of Saccharomyces cerevisiae towards trichothecene toxins, in particular to deoxynivalenol (DON), in order to improve the utility of this yeast as a bioassay indicator organism. We report the construction of a strain with inactivated genes (PDR5, PDR10, PDR15) encoding ABC transporter proteins with specificity for the trichothecene deoxynivalenol, with inactivated AYT1 (encoding a trichothecene-3-O-acetyltransferase), and inactivated UBI4 and UBP6 genes. Inactivation of the stress inducible polyubiquitin gene UBI4 or the ubiquitin protease UBP6 increased DON sensitivity, the inactivation of both genes had a synergistic effect. The resulting pdr5 pdr10 pdr15 ayt1 ubp6 ubi4 mutant strain showed 50% growth inhibition at a DON concentration of 5 mg/l under optimal conditions. The development of a simple two step assay for microbial DON degradation in 96 well microtiter format and its testing with the DON detoxifying bacterium BBSH 797 is reported.
- Published
- 2008
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21. Cloning and characterization of the ribosomal protein L3 (RPL3) gene family from Triticum aestivum.
- Author
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Lucyshyn D, Busch BL, Abolmaali S, Steiner B, Chandler E, Sanjarian F, Mousavi A, Nicholson P, Buerstmayr H, and Adam G
- Subjects
- Chromosome Mapping, Chromosomes, Plant, Cloning, Molecular, Drug Resistance genetics, Molecular Sequence Data, Plant Proteins metabolism, Polymorphism, Single Nucleotide, Polymorphism, Single-Stranded Conformational, Quantitative Trait Loci, Ribosomal Protein L3, Ribosomal Proteins metabolism, Trichothecenes toxicity, Triticum drug effects, Plant Proteins genetics, Ribosomal Proteins genetics, Triticum genetics
- Abstract
Plant pathogenic fungi of the genus Fusarium can cause severe diseases on small grain cereals and maize. The contamination of harvested grain with Fusarium mycotoxins is a threat to human and animal health. In wheat production of the toxin deoxynivalenol (DON), which inhibits eukaryotic protein biosynthesis, is a virulence factor of Fusarium, and resistance against DON is considered to be part of Fusarium resistance. Previously, single amino acid changes in RPL3 (ribosomal protein L3) conferring DON resistance have been described in yeast. The goal of this work was to characterize the RPL3 gene family from wheat and to investigate the potential role of naturally existing RPL3 alleles in DON resistance by comparing Fusarium-resistant and susceptible cultivars. The gene family consists of three homoeologous alleles of both RPL3A and RPL3B, which are located on chromosomes 4A (RPL3-B2), 4B (RPL3-B1), 4D (RPL3-B3), 5A (RPL3-A3), 5B (RPL3-A2) and 5D (RPL3-A1). Alternative splicing was detected in the TaRPL3-A2 gene. Sequence comparison revealed no amino acid differences between cultivars differing in Fusarium resistance. While using developed SNP markers we nevertheless found that one of the genes, namely, TaRPL3-A3 mapped close to a Fusarium resistance QTL (Qfhs.ifa-5A). The potential role of the RPL3 gene family in DON resistance of wheat is discussed.
- Published
- 2007
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22. Heterologous expression of Arabidopsis UDP-glucosyltransferases in Saccharomyces cerevisiae for production of zearalenone-4-O-glucoside.
- Author
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Poppenberger B, Berthiller F, Bachmann H, Lucyshyn D, Peterbauer C, Mitterbauer R, Schuhmacher R, Krska R, Glössl J, and Adam G
- Subjects
- Arabidopsis Proteins genetics, Base Sequence, Cloning, Molecular, DNA Primers, Glucosyltransferases genetics, Recombinant Proteins metabolism, Zearalenone biosynthesis, Arabidopsis genetics, Arabidopsis Proteins metabolism, Glucosides biosynthesis, Glucosyltransferases metabolism, Saccharomyces cerevisiae genetics, Zearalenone analogs & derivatives
- Abstract
Zearalenone, a secondary metabolite produced by several plant-pathogenic fungi of the genus Fusarium, has high estrogenic activity in vertebrates. We developed a Saccharomyces cerevisiae bioassay strain that we used to identify plant genes encoding UDP-glucosyltransferases that can convert zearalenone into zearalenone-4-O-glucoside (ZON-4-O-Glc). Attachment of the glucose moiety to zearalenone prevented the interaction of the mycotoxin with the human estrogen receptor. We found that two of six clustered, similar UGT73C genes of Arabidopsis thaliana encode glucosyltransferases that can inactivate zearalenone in the yeast bioassay. The formation of glucose conjugates seems to be an important plant mechanism for coping with zearalenone but may result in significant amounts of "masked" zearalenone in Fusarium-infected plant products. Due to the unavailability of an analytical standard, the ZON-4-O-Glc is not measured in routine analytical procedures, even though it can be converted back to active zearalenone in the digestive tracts of animals. Zearalenone added to yeast transformed with UGT73C6 was converted rapidly and efficiently to ZON-4-O-Glc, suggesting that the cloned UDP-glucosyltransferase could be used to produce reference glucosides of zearalenone and its derivatives.
- Published
- 2006
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23. Toxin-dependent utilization of engineered ribosomal protein L3 limits trichothecene resistance in transgenic plants.
- Author
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Mitterbauer R, Poppenberger B, Raditschnig A, Lucyshyn D, Lemmens M, Glössl J, and Adam G
- Abstract
The contamination of agricultural products with Fusarium mycotoxins is a problem of world-wide importance. Fusarium graminearum and related species, which are important pathogens of small grain cereals and maize, produce an economically important and structurally diverse class of toxins designated trichothecenes. Trichothecenes inhibit eukaryotic protein synthesis. Therefore, a proposed role for these fungal toxins in plant disease development is to block or delay the expression of defence-related proteins induced by the plant. Using yeast as a model system, we have identified several mutations in the gene encoding ribosomal protein L3 (Rpl3), which confer semi-dominant resistance to trichothecenes. Expression of an engineered tomato RPL3 (LeRPL3) cDNA, into which one of the amino acid changes identified in yeast was introduced, improved the ability of transgenic tobacco plants to adapt to the trichothecene deoxynivalenol (DON), but did not result in constitutive resistance. We show here that, in the presence of wild-type Rpl3 protein, the engineered Rpl3 protein is not utilized, unless yeast transformants or the transgenic plants are challenged with sublethal amounts of toxin. Our data from yeast two-hybrid experiments suggest that affinity for the ribosome assembly factor Rrb1p could be altered by the toxin resistance-conferring mutation. This toxin-dependent utilization of the resistance-conferring Rpl3 protein could seriously limit efforts to utilize the identified target alterations in transgenic crops to increase trichothecene tolerance and Fusarium resistance.
- Published
- 2004
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24. Detoxification of the Fusarium mycotoxin deoxynivalenol by a UDP-glucosyltransferase from Arabidopsis thaliana.
- Author
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Poppenberger B, Berthiller F, Lucyshyn D, Sieberer T, Schuhmacher R, Krska R, Kuchler K, Glössl J, Luschnig C, and Adam G
- Subjects
- Amino Acid Sequence, Arabidopsis microbiology, Arabidopsis Proteins, Carbon chemistry, Cloning, Molecular, DNA, Complementary metabolism, Escherichia coli metabolism, Fusarium pathogenicity, Gene Library, Genetic Vectors, Glutathione Transferase metabolism, Glycosylation, Mass Spectrometry, Models, Chemical, Molecular Sequence Data, Phenotype, Promoter Regions, Genetic, Protein Conformation, RNA, Messenger metabolism, Reverse Transcriptase Polymerase Chain Reaction, Ribosomes metabolism, Sequence Homology, Amino Acid, Transgenes, Uridine Diphosphate Glucose metabolism, Arabidopsis enzymology, Fusarium metabolism, Glucosyltransferases chemistry, Glucosyltransferases metabolism, Mycotoxins metabolism, Trichothecenes metabolism
- Abstract
Plant pathogenic fungi of the genus Fusarium cause agriculturally important diseases of small grain cereals and maize. Trichothecenes are a class of mycotoxins produced by different Fusarium species that inhibit eukaryotic protein biosynthesis and presumably interfere with the expression of genes induced during the defense response of the plants. One of its members, deoxynivalenol, most likely acts as a virulence factor during fungal pathogenesis and frequently accumulates in grain to levels posing a threat to human and animal health. We report the isolation and characterization of a gene from Arabidopsis thaliana encoding a UDP-glycosyltransferase that is able to detoxify deoxynivalenol. The enzyme, previously assigned the identifier UGT73C5, catalyzes the transfer of glucose from UDP-glucose to the hydroxyl group at carbon 3 of deoxynivalenol. Using a wheat germ extract-coupled transcription/translation system we have shown that this enzymatic reaction inactivates the mycotoxin. This deoxynivalenol-glucosyltransferase (DOGT1) was also found to detoxify the acetylated derivative 15-acetyl-deoxynivalenol, whereas no protective activity was observed against the structurally similar nivalenol. Expression of the glucosyltransferase is developmentally regulated and induced by deoxynivalenol as well as salicylic acid, ethylene, and jasmonic acid. Constitutive overexpression in Arabidopsis leads to enhanced tolerance against deoxynivalenol.
- Published
- 2003
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