Publications by authors named "Jacques Hille"

25 Publications

  • Page 1 of 1

A novel seed plants gene regulates oxidative stress tolerance in Arabidopsis thaliana.

Cell Mol Life Sci 2020 Feb 27;77(4):705-718. Epub 2019 Jun 27.

Center of Plant Systems Biology and Biotechnology, 139 Ruski Blvd, 4000, Plovdiv, Bulgaria.

Oxidative stress can lead to plant growth retardation, yield loss, and death. The atr7 mutant of Arabidopsis thaliana exhibits pronounced tolerance to oxidative stress. Using positional cloning, confirmed by knockout and RNA interference (RNAi) lines, we identified the atr7 mutation and revealed that ATR7 is a previously uncharacterized gene with orthologs in other seed plants but with no homology to genes in lower plants, fungi or animals. Expression of ATR7-GFP fusion shows that ATR7 is a nuclear-localized protein. RNA-seq analysis reveals that transcript levels of genes encoding abiotic- and oxidative stress-related transcription factors (DREB19, HSFA2, ZAT10), chromatin remodelers (CHR34), and unknown or uncharacterized proteins (AT5G59390, AT1G30170, AT1G21520) are elevated in atr7. This indicates that atr7 is primed for an upcoming oxidative stress via pathways involving genes of unknown functions. Collectively, the data reveal ATR7 as a novel seed plants-specific nuclear regulator of oxidative stress response.
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http://dx.doi.org/10.1007/s00018-019-03202-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7040063PMC
February 2020

Occurrence, biochemistry and biological effects of host-selective plant mycotoxins.

Food Chem Toxicol 2018 Feb 27;112:251-264. Epub 2017 Dec 27.

Center of Plant Systems Biology and Biotechnology, 139 Ruski blvd., Plovdiv 4000, Bulgaria; Institute of Molecular Biology and Biotechnology, 105 Ruski blvd., Plovdiv 4000, Bulgaria; Department of Plant Physiology and Molecular Biology, Plovdiv University, 24 Tsar Assen str., Plovdiv 4000, Bulgaria. Electronic address:

Host-selective mycotoxins (HSTs) are various secondary metabolites or proteinaceous compounds secreted by pathogenic necrotrophic fungi that feed off on dead tissues of certain plants. Research on the HSTs has not only fundamental but also practical importance. On one hand they are implicated in the onset of devastating crop diseases. On the other hand, they have been studied as a good model for revealing the intricate mechanisms of plant-pathogen interactions. At the cellular level, HSTs target different compartments and in most instances induce programmed cell death (PCD) by a wide range of mechanisms. Often the responses provoked by HSTs resemble the effector-triggered immunity used by plant cells to combat biotrophic pathogens, which suggests that HST-producing fungi exploit the plants' own defensive systems to derive benefits. Although by definition HSTs are active only in tissues of susceptible plant genotypes, it has been demonstrated that some of them are able to influence animal cells as well. The possible effects, like cytotoxicity or cytostasis, can be harmful or beneficial and thus HSTs may either pose a health risk for humans and livestock, or be of prospective use in the fields of pharmacology, medicine and agriculture.
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http://dx.doi.org/10.1016/j.fct.2017.12.047DOI Listing
February 2018

ROS-mediated abiotic stress-induced programmed cell death in plants.

Front Plant Sci 2015 18;6:69. Epub 2015 Feb 18.

Institute of Molecular Biology and Biotechnology, Plovdiv Bulgaria ; Department of Molecular Biology, Institute of Biochemistry and Biology, University of Potsdam, Potsdam-Golm Germany.

During the course of their ontogenesis plants are continuously exposed to a large variety of abiotic stress factors which can damage tissues and jeopardize the survival of the organism unless properly countered. While animals can simply escape and thus evade stressors, plants as sessile organisms have developed complex strategies to withstand them. When the intensity of a detrimental factor is high, one of the defense programs employed by plants is the induction of programmed cell death (PCD). This is an active, genetically controlled process which is initiated to isolate and remove damaged tissues thereby ensuring the survival of the organism. The mechanism of PCD induction usually includes an increase in the levels of reactive oxygen species (ROS) which are utilized as mediators of the stress signal. Abiotic stress-induced PCD is not only a process of fundamental biological importance, but also of considerable interest to agricultural practice as it has the potential to significantly influence crop yield. Therefore, numerous scientific enterprises have focused on elucidating the mechanisms leading to and controlling PCD in response to adverse conditions in plants. This knowledge may help develop novel strategies to obtain more resilient crop varieties with improved tolerance and enhanced productivity. The aim of the present review is to summarize the recent advances in research on ROS-induced PCD related to abiotic stress and the role of the organelles in the process.
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http://dx.doi.org/10.3389/fpls.2015.00069DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4332301PMC
March 2015

Natural products from resurrection plants: potential for medical applications.

Biotechnol Adv 2014 Nov 26;32(6):1091-101. Epub 2014 Mar 26.

University of Potsdam, Institute of Biochemistry and Biology, Department of Molecular Biology, Karl-Liebknecht str. 24-25, Haus 20, 14476 Potsdam-Golm, Germany; Max-Planck Institute of Molecular Plant Physiology, Plant Signalling Group, Am Mühlenberg 1, 14476 Potsdam-Golm, Germany.

Resurrection species are a group of land plants that can tolerate extreme desiccation of their vegetative tissues during harsh drought stress, and still quickly - often within hours - regain normal physiological and metabolic functions following rehydration. At the molecular level, this desiccation tolerance is attributed to basal cellular mechanisms including the constitutive expression of stress-associated genes and high levels of protective metabolites present already in the absence of stress, as well as to transcriptome and metabolome reconfigurations rapidly occurring during the initial phases of drought stress. Parts of this response are conferred by unique metabolites, including a diverse array of sugars, phenolic compounds, and polyols, some of which accumulate to high concentrations within the plant cell. In addition to drought stress, these metabolites are proposed to contribute to the protection against other abiotic stresses and to an increased oxidative stress tolerance. Recently, extracts of resurrection species and particular secondary metabolites therein were reported to display biological activities of importance to medicine, with e.g. antibacterial, anticancer, antifungal, and antiviral activities, rendering them possible candidates for the development of novel drug substances as well as for cosmetics. Herein, we provide an overview of the metabolite composition of resurrection species, summarize the latest reports related to the use of natural products from resurrection plants, and outline their potential for medical applications.
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http://dx.doi.org/10.1016/j.biotechadv.2014.03.005DOI Listing
November 2014

Towards in vivo mutation analysis: knock-out of specific chlorophylls bound to the light-harvesting complexes of Arabidopsis thaliana - the case of CP24 (Lhcb6).

Biochim Biophys Acta 2014 Sep 19;1837(9):1500-6. Epub 2014 Feb 19.

Department of Biophysical chemistry, Groningen Biological Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 7, 9747 AG Groningen, The Netherlands; Department of Physics and Astronomy, Faculty of Sciences, VU University Amsterdam, De Boelelaan 1081, 1081 HV Amsterdam, The Netherlands. Electronic address:

In the last ten years, a large series of studies have targeted antenna complexes of plants (Lhc) with the aim of understanding the mechanisms of light harvesting and photoprotection. Combining spectroscopy, modeling and mutation analyses, the role of individual pigments in these processes has been highlighted in vitro. In plants, however, these proteins are associated with multiple complexes of the photosystems and function within this framework. In this work, we have envisaged a way to bridge the gap between in vitro and in vivo studies by knocking out in vivo pigments that have been proposed to play an important role in excitation energy transfer between the complexes or in photoprotection. We have complemented a CP24 knock-out mutant of Arabidopsis thaliana with the CP24 (Lhcb6) gene carrying a His-tag and with a mutated version lacking the ligand for chlorophyll 612, a specific pigment that in vitro experiments have indicated as the lowest energy site of the complex. Both complexes efficiently integrated into the thylakoid membrane and assembled into the PSII supercomplexes, indicating that the His-tag does not impair the organization in vivo. The presence of the His-tag allowed the purification of CP24-WT and of CP24-612 mutant in their native states. It is shown that CP24-WT coordinates 10 chlorophylls and 2 carotenoid molecules and has properties identical to those of the reconstituted complex, demonstrating that the complex self-assembled in vitro assumes the same folding as in the plant. The absence of the ligand for chlorophyll 612 leads to the loss of one Chl a and of lutein, again as in vitro, indicating the feasibility of the method. This article is part of a special issue entitled: photosynthesis research for sustainability: keys to produce clean energy.
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http://dx.doi.org/10.1016/j.bbabio.2014.02.012DOI Listing
September 2014

A simple and powerful approach for isolation of Arabidopsis mutants with increased tolerance to H2O2-induced cell death.

Methods Enzymol 2013 ;527:203-20

Department of Plant Physiology and Plant Molecular Biology, University of Plovdiv, Plovdiv, Bulgaria.

A genetic approach is described to isolate mutants more tolerant to oxidative stress. A collection of T-DNA activation tag Arabidopsis thaliana mutant lines was screened for survivors under conditions that trigger H2O2-induced cell death. Oxidative stress was induced by applying the catalase (CAT) inhibitor aminotriazole (AT) in the growth media, which results in decrease in CAT enzyme activity, H2O2 accumulation, and subsequent plant death. One mutant was recovered from the screening and named oxr1 (oxidative stress resistant 1). The location of the T-DNA insertion was identified by TAIL-PCR. Oxr1 exhibited lack of cell death symptoms and more fresh weight and chlorophyll content compared to wild type. The lack of cell death correlated with more prominent induction of anthocyanins synthesis in oxr1. These results demonstrate the feasibility of AT as a screening agent for the isolation of oxidative stress-tolerant mutants and indicate a possible protective role for anthocyanins against AT-induced cell death. The chapter includes protocols for ethyl methanesulfonate mutagenesis, mutant screening using AT, T-DNA identification by TAIL-PCR, CAT activity measurements, and determination of malondialdehyde, chlorophyll, and anthocyanins.
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http://dx.doi.org/10.1016/B978-0-12-405882-8.00011-8DOI Listing
January 2014

Molecular mechanisms of desiccation tolerance in the resurrection glacial relic Haberlea rhodopensis.

Cell Mol Life Sci 2013 Feb 21;70(4):689-709. Epub 2012 Sep 21.

Department of Plant Physiology and Plant Molecular Biology, University of Plovdiv, 24 Tsar Assen Str., Plovdiv, 4000, Bulgaria.

Haberlea rhodopensis is a resurrection plant with remarkable tolerance to desiccation. Haberlea exposed to drought stress, desiccation, and subsequent rehydration showed no signs of damage or severe oxidative stress compared to untreated control plants. Transcriptome analysis by next-generation sequencing revealed a drought-induced reprogramming, which redirected resources from growth towards cell protection. Repression of photosynthetic and growth-related genes during water deficiency was concomitant with induction of transcription factors (members of the NAC, NF-YA, MADS box, HSF, GRAS, and WRKY families) presumably acting as master switches of the genetic reprogramming, as well as with an upregulation of genes related to sugar metabolism, signaling, and genes encoding early light-inducible (ELIP), late embryogenesis abundant (LEA), and heat shock (HSP) proteins. At the same time, genes encoding other LEA, HSP, and stress protective proteins were constitutively expressed at high levels even in unstressed controls. Genes normally involved in tolerance to salinity, chilling, and pathogens were also highly induced, suggesting a possible cross-tolerance against a number of abiotic and biotic stress factors. A notable percentage of the genes highly regulated in dehydration and subsequent rehydration were novel, with no sequence homology to genes from other plant genomes. Additionally, an extensive antioxidant gene network was identified with several gene families possessing a greater number of antioxidant genes than most other species with sequenced genomes. Two of the transcripts most abundant during all conditions encoded catalases and five more catalases were induced in water-deficient samples. Using the pharmacological inhibitor 3-aminotriazole (AT) to compromise catalase activity resulted in increased sensitivity to desiccation. Metabolome analysis by GC or LC-MS revealed accumulation of sucrose, verbascose, spermidine, and γ-aminobutyric acid during drought, as well as particular secondary metabolites accumulating during rehydration. This observation, together with the complex antioxidant system and the constitutive expression of stress protective genes suggests that both constitutive and inducible mechanisms contribute to the extreme desiccation tolerance of H. rhodopensis.
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http://dx.doi.org/10.1007/s00018-012-1155-6DOI Listing
February 2013

Molecular basis of plant stress.

Cell Mol Life Sci 2012 Oct 25;69(19):3161-3. Epub 2012 Jul 25.

Department of Plant Physiology and Plant Molecular Biology, University of Plovdiv, Bulgaria.

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http://dx.doi.org/10.1007/s00018-012-1086-2DOI Listing
October 2012

In vitro regeneration of wild chervil (Anthriscus sylvestris L.).

In Vitro Cell Dev Biol Plant 2012 Jun 25;48(3):355-361. Epub 2011 Oct 25.

Anthriscus sylvestris (L.) Hoffm. (Apiaceae) is a common wild plant that accumulates the lignan deoxypodophyllotoxin. Deoxypodophyllotoxin can be hydroxylated at the C-7 position in recombinant organisms yielding podophyllotoxin, which is used as a semi-synthetic precursor for the anticancer drugs, etoposide phosphate and teniposide. As in vitro regeneration of A. sylvestris has not yet been reported, development of a regeneration protocol for A. sylvestris would be useful as a micropropagation tool and for metabolic engineering of the plant. Calli were induced from hypocotyl explants and transferred to shoot induction medium containing zeatin riboside. Regenerated shoots were obtained within 6 mo and were transferred onto growth regulator-free root induction medium containing 1% sucrose. Regenerated plants transferred to soil and acclimatized in a greenhouse. Plants were transferred to the field with a 100% survival rate. Regenerated plants flowered and were fully fertile. This is the first report of complete regeneration of A. sylvestris via shoot organogenesis from callus.
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http://dx.doi.org/10.1007/s11627-011-9410-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3375414PMC
June 2012

Oxidative stress provokes distinct transcriptional responses in the stress-tolerant atr7 and stress-sensitive loh2 Arabidopsis thaliana mutants as revealed by multi-parallel quantitative real-time PCR analysis of ROS marker and antioxidant genes.

Plant Physiol Biochem 2012 Oct 4;59:20-9. Epub 2012 Jun 4.

Department of Plant Physiology and Plant Molecular Biology, University of Plovdiv, 24 Tsar Assen Str., Plovdiv 4000, Bulgaria.

The Arabidopsis thaliana atr7 mutant is tolerant to oxidative stress induced by paraquat (PQ) or the catalase inhibitor aminotriazole (AT), while its original background loh2 and wild-type plants are sensitive. Both, AT and PQ, which stimulate the intracellular formation of H₂O₂ or superoxide anions, respectively, trigger cell death in loh2 but do not lead to visible damage in atr7. To study gene expression during oxidative stress and ROS-induced programmed cell death, two platforms for multi-parallel quantitative real-time PCR (qRT-PCR) analysis of 217 antioxidant and 180 ROS marker genes were employed. The qRT-PCR analyses revealed AT- and PQ-induced expression of many ROS-responsive genes mainly in loh2, confirming that an oxidative burst plays a role in the activation of the cell death in this mutant. Some of the genes were specifically regulated by either AT or PQ, serving as markers for particular types of ROS. Genes significantly induced by both AT and PQ in loh2 included transcription factors (ANAC042/JUB1, ANAC102, DREB19, HSFA2, RRTF1, ZAT10, ZAT12, ethylene-responsive factors), signaling compounds, ferritins, alternative oxidases, and antioxidant enzymes. Many of these genes were upregulated in atr7 compared to loh2 under non-stress conditions at the first time point, indicating that higher basal levels of ROS and higher antioxidant capacity in atr7 are responsible for the enhanced tolerance to oxidative stress and suggesting a possible tolerance against multiple stresses of this mutant.
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http://dx.doi.org/10.1016/j.plaphy.2012.05.024DOI Listing
October 2012

Seasonal variations in the deoxypodophyllotoxin content and yield of Anthriscus sylvestris L. (Hoffm.) grown in the field and under controlled conditions.

J Agric Food Chem 2011 Aug 18;59(15):8132-9. Epub 2011 Jul 18.

Department of Pharmaceutical Biology, University of Groningen, Groningen, The Netherlands.

Deoxypodophyllotoxin (DPT) is the main lignan in Anthriscus sylvestris . For this study two sets of experiments with 16 plants and seeds, collected from a wide geographical range, were carried out. The DPT content in roots was significantly lower (p < 0.05) when the plants were cultivated in a non-native environment. For field-grown plants the highest DPT content was found in March (second year): 0.15% w/w (dry weight) in roots; 0.03% w/w in aerial parts. For plants grown in the climate room, the highest concentration (0.14% w/w) was observed in April (second year) in the roots and in July (first year) in the aerial parts (0.05% w/w). For the isolation of DPT, roots are the most suitable part. The best harvest times are March (second year) for outdoor plants and April (second year) for indoor plants when height content and adequate biomass give the optimal DPT yield.
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http://dx.doi.org/10.1021/jf200177qDOI Listing
August 2011

A mutation in the cytosolic O-acetylserine (thiol) lyase induces a genome-dependent early leaf death phenotype in Arabidopsis.

BMC Plant Biol 2010 Apr 29;10:80. Epub 2010 Apr 29.

Molecular Biology of Plants, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Kerklaan 30, Haren, The Netherlands.

Background: Cysteine is a component in organic compounds including glutathione that have been implicated in the adaptation of plants to stresses. O-acetylserine (thiol) lyase (OAS-TL) catalyses the final step of cysteine biosynthesis. OAS-TL enzyme isoforms are localised in the cytoplasm, the plastids and mitochondria but the contribution of individual OAS-TL isoforms to plant sulphur metabolism has not yet been fully clarified.

Results: The seedling lethal phenotype of the Arabidopsis onset of leaf death3-1 (old3-1) mutant is due to a point mutation in the OAS-A1 gene, encoding the cytosolic OAS-TL. The mutation causes a single amino acid substitution from Gly162 to Glu162, abolishing old3-1 OAS-TL activity in vitro. The old3-1 mutation segregates as a monogenic semi-dominant trait when backcrossed to its wild type accession Landsberg erecta (Ler-0) and the Di-2 accession. Consistent with its semi-dominant behaviour, wild type Ler-0 plants transformed with the mutated old3-1 gene, displayed the early leaf death phenotype. However, the old3-1 mutation segregates in an 11:4:1 (wild type: semi-dominant: mutant) ratio when backcrossed to the Colombia-0 and Wassilewskija accessions. Thus, the early leaf death phenotype depends on two semi-dominant loci. The second locus that determines the old3-1 early leaf death phenotype is referred to as odd-ler (for old3 determinant in the Ler accession) and is located on chromosome 3. The early leaf death phenotype is temperature dependent and is associated with increased expression of defence-response and oxidative-stress marker genes. Independent of the presence of the odd-ler gene, OAS-A1 is involved in maintaining sulphur and thiol levels and is required for resistance against cadmium stress.

Conclusions: The cytosolic OAS-TL is involved in maintaining organic sulphur levels. The old3-1 mutation causes genome-dependent and independent phenotypes and uncovers a novel function for the mutated OAS-TL in cell death regulation.
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http://dx.doi.org/10.1186/1471-2229-10-80DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2890954PMC
April 2010

The Arabidopsis onset of leaf death5 mutation of quinolinate synthase affects nicotinamide adenine dinucleotide biosynthesis and causes early ageing.

Plant Cell 2008 Oct 31;20(10):2909-25. Epub 2008 Oct 31.

Molecular Biology of Plants, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, 9751 NN Haren, The Netherlands.

Leaf senescence in Arabidopsis thaliana is a strict, genetically controlled nutrient recovery program, which typically progresses in an age-dependent manner. Leaves of the Arabidopsis onset of leaf death5 (old5) mutant exhibit early developmental senescence. Here, we show that OLD5 encodes quinolinate synthase (QS), a key enzyme in the de novo synthesis of NAD. The Arabidopsis QS was previously shown to carry a Cys desulfurase domain that stimulates reconstitution of the oxygen-sensitive Fe-S cluster that is required for QS activity. The old5 lesion in this enzyme does not affect QS activity but it decreases its Cys desulfurase activity and thereby the long-term catalytic competence of the enzyme. The old5 mutation causes increased NAD steady state levels that coincide with increased activity of enzymes in the NAD salvage pathway. NAD plays a key role in cellular redox reactions, including those of the tricarboxylic acid cycle. Broad-range metabolite profiling of the old5 mutant revealed that it contains higher levels of tricarboxylic acid cycle intermediates and nitrogen-containing amino acids. The mutant displays a higher respiration rate concomitant with increased expression of oxidative stress markers. We postulate that the alteration in the oxidative state is integrated into the plant developmental program, causing early ageing of the mutant.
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http://dx.doi.org/10.1105/tpc.107.056341DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2590718PMC
October 2008

Arabidopsis AAL-toxin-resistant mutant atr1 shows enhanced tolerance to programmed cell death induced by reactive oxygen species.

Biochem Biophys Res Commun 2008 Oct 24;375(4):639-44. Epub 2008 Aug 24.

Department of Plant Physiology and Plant Molecular Biology, University of Plovdiv, 24 Tsar Assen street, Plovdiv 4000, Bulgaria.

The fungal AAL-toxin triggers programmed cell death (PCD) through perturbations of sphingolipid metabolism in AAL-toxin-sensitive plants. While Arabidopsis is relatively insensitive to the toxin, the loh2 mutant exhibits increased susceptibility to AAL-toxin due to the knockout of a gene involved in sphingolipid metabolism. Genetic screening of mutagenized loh2 seeds resulted in the isolation of AAL-toxin-resistant mutant atr1.Atr1 displays a wild type phenotype when grown on soil but it develops less biomass than loh2 on media supplemented with 2% and 3% sucrose. Atr1 was also more tolerant to the reactive oxygen species-generating herbicides aminotriazole (AT) and paraquat. Microarray analyses of atr1 and loh2 under AT-treatment conditions that trigger cell death in loh2 and no visible damage in atr1 revealed genes specifically regulated in atr1 or loh2. In addition, most of the genes strongly downregulated in both mutants were related to cell wall extension and cell growth, consistent with the apparent and similar AT-induced cessation of growth in both mutants. This indicates that two different pathways, a first controlling growth inhibition and a second triggering cell death, are associated with AT-induced oxidative stress.
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http://dx.doi.org/10.1016/j.bbrc.2008.08.056DOI Listing
October 2008

Arabidopsis CPR5 is a senescence-regulatory gene with pleiotropic functions as predicted by the evolutionary theory of senescence.

J Exp Bot 2007 22;58(14):3885-94. Epub 2007 Nov 22.

Molecular Biology of Plants, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Kerklaan 30, 9751 NN, Haren, The Netherlands.

Evolutionary theories of senescence predict that genes with pleiotropic functions are important for senescence regulation. In plants there is no direct molecular genetic test for the existence of such senescence-regulatory genes. Arabidopsis cpr5 mutants exhibit multiple phenotypes including hypersensitivity to various signalling molecules, constitutive expression of pathogen-related genes, abnormal trichome development, spontaneous lesion formation, and accelerated leaf senescence. These indicate that CPR5 is a beneficial gene which controls multiple facets of the Arabidopsis life cycle. Ectopic expression of CPR5 restored all the mutant phenotypes. However, in transgenic plants with increased CPR5 transcripts, accelerated leaf senescence was observed in detached leaves and at late development around 50 d after germination, as illustrated by the earlier onset of senescence-associated physiological and molecular markers. Thus, CPR5 has early-life beneficial effects by repressing cell death and insuring normal plant development, but late-life deleterious effects by promoting developmental senescence. As such, CPR5 appears to function as a typical senescence-regulatory gene as predicted by the evolutionary theories of senescence.
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http://dx.doi.org/10.1093/jxb/erm237DOI Listing
April 2008

Ethylene-induced leaf senescence depends on age-related changes and OLD genes in Arabidopsis.

J Exp Bot 2005 Nov 19;56(421):2915-23. Epub 2005 Sep 19.

Molecular Biology of Plants, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Kerklaan 30, 9751 NN, Haren, The Netherlands.

Ethylene can only induce senescence in leaves that have reached a defined age. Thus, ethylene-induced senescence depends on age-related changes (ARCs) of individual leaves. The relationship between ethylene and age in the induction of leaf senescence was tested in Arabidopsis Ler-0, Col-0, and Ws-0 accessions as well as in eight old (onset of leaf death) mutants, isolated from the Ler-0 background. Plants with a constant final age of 24 d were exposed to ethylene for 3-16 d. The wild-type accessions showed a common response to the ethylene treatment. Increasing ethylene treatments of 3-12 d caused an increase in the number of yellow leaves. However, an ethylene exposure time of 16 d resulted in a decrease in the amount of yellowing. Thus, ethylene can both positively and negatively influence ARCs and the subsequent induction of leaf senescence, depending on the length of the treatment. The old mutants showed altered responses to the ethylene treatments. old1 and old11 were hypersensitive to ethylene in the triple response assay and a 12-d ethylene exposure resulted in a decrease in the amount of yellow leaves. The other six mutants did not show a decrease in yellow leaves with an ethylene treatment of 16 d. The results revealed that the effect of ethylene on the induction of senescence can be modified by at least eight genes.
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http://dx.doi.org/10.1093/jxb/eri287DOI Listing
November 2005

The products of the broken Tm-2 and the durable Tm-2(2) resistance genes from tomato differ in four amino acids.

J Exp Bot 2005 Nov 19;56(421):2925-33. Epub 2005 Sep 19.

Department Molecular Biology of Plants, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Kerklaan 30, 9751 NN, Haren, The Netherlands.

To gain an insight into the processes underlying disease resistance and its durability, the durable Tm-2(2) resistance gene was compared with the broken Tm-2 resistance gene. The Tm-2 gene of tomato could be isolated via PCR with primers based on the Tm-2(2) sequence. The Tm-2 gene, like the Tm-2(2) gene, encodes an 861 amino acid polypeptide, which belongs to the coiled coil/nucleotide binding site/leucine-rich repeat class of resistance proteins. The functionality and the nature of the isolated Tm-2 gene were confirmed by introducing the gene under the control of the 35S promoter into tomato mosaic virus-susceptible tobacco. This transgenic tobacco was crossed with transgenic tobacco plants producing the movement protein (MP)-authenticated MP as the Avr protein of the Tm-2 resistance. The Tm-2(2) and Tm-2 open reading frames only differ in seven nucleotides, which on a protein level results in four amino acid differences, of which two are located in the nucleotide binding site and two are located in the leucine-rich repeat domain. The small difference between the two proteins suggests a highly similar interaction of these proteins with the MP, which has major implications for the concept of durability. Comparison of the two resistance-conferring alleles (Tm-2 and Tm-2(2)) with two susceptible alleles (tm-2 and lptm-2) allowed discussion of the structure-function relationship in the Tm-2 proteins. It is proposed that the Tm-2 proteins display a partitioning of the leucine-rich repeat domain, in which the N-terminal and C-terminal parts function in signal transduction and MP recognition, respectively.
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http://dx.doi.org/10.1093/jxb/eri288DOI Listing
November 2005

Arabidopsis RecQI4A suppresses homologous recombination and modulates DNA damage responses.

Plant J 2005 Sep;43(6):789-98

Molecular Biology of Plants, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Kerklaan 30, 9751 NN Haren, The Netherlands.

The DNA damage response and DNA recombination are two interrelated mechanisms involved in maintaining the integrity of the genome, but in plants they are poorly understood. RecQ is a family of genes with conserved roles in the regulation of DNA recombination in eukaryotes; there are seven members in Arabidopsis. Here we report on the functional analysis of the Arabidopsis RecQl4A gene. Ectopic expression of Arabidopsis RecQl4A in yeast RecQ-deficient cells suppressed their hypersensitivity to the DNA-damaging drug methyl methanesulfonate (MMS) and enhanced their rate of homologous recombination (HR). Analysis of three recQl4A mutant alleles revealed no obvious developmental defects or telomere deregulation in plants grown under standard growth conditions. Compared with wild-type Arabidopsis, the recQl4A mutant seedlings were found to be hypersensitive to UV light and MMS, and more resistant to mitomycin C. The average frequency of intrachromosomal HR in recQl4A mutant plants was increased 7.5-fold over that observed in wild-type plants. The data reveal roles for Arabidopsis RecQl4A in maintenance of genome stability by modulation of the DNA damage response and suppression of HR.
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http://dx.doi.org/10.1111/j.1365-313X.2005.02501.xDOI Listing
September 2005

Hydrogen peroxide-induced cell death in Arabidopsis: transcriptional and mutant analysis reveals a role of an oxoglutarate-dependent dioxygenase gene in the cell death process.

IUBMB Life 2005 Mar;57(3):181-8

Department of Plant Physiology and Plant Molecular Biology, University of Plovdiv, Plovdiv, Bulgaria.

Hydrogen peroxide is a major regulator of plant programmed cell death (PCD) but little is known about the downstream genes from the H(2)O(2)-signaling network that mediate the cell death. To address this question, a novel system for studying H(2)O(2)-induced programmed cell death in Arabidopsis thaliana was used. The catalase inhibitor aminotriazole (AT) reduced the catalase activity and caused endogenous accumulation of hydrogen peroxide that eventually triggered cell death. Microarray analysis with a DNA chip representing 21500 genes and subsequent comparison with other PCD-related expression studies revealed a set of new H(2)O(2)-responsive genes that were highly regulated in a common fashion during different types of PCD. These included an oxoglutarate-dependent dioxygenase and various oxidoreductases, the transcription factors Zat11, WRKY75 and NAM, proteasomal components, a heterologous group of genes with diverse functions, and genes encoding proteins with unknown functions. Knockout lines of the oxoglutarate-dependent dioxygenase exhibited significantly reduced death symptoms and chlorophyll loss upon H(2)O(2)-induced cell death, indicating a role for this gene in the cell death network.
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http://dx.doi.org/10.1080/15216540500090793DOI Listing
March 2005

Hydrogen peroxide as a signal controlling plant programmed cell death.

J Cell Biol 2005 Jan;168(1):17-20

Department Molecular Biology of Plants, Researchschool GBB, University of Groningen, 9751 NN, Haren, Netherlands.

Hydrogen peroxide (H2O2) has established itself as a key player in stress and programmed cell death responses, but little is known about the signaling pathways leading from H2O2 to programmed cell death in plants. Recently, identification of key regulatory mutants and near-full genome coverage microarray analysis of H2O2-induced cell death have begun to unravel the complexity of the H2O2 network. This review also describes a novel link between H2O2 and sphingolipids, two signals that can interplay and regulate plant cell death.
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http://dx.doi.org/10.1083/jcb.200409170DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2171664PMC
January 2005

Cloning and characterization of the durable tomato mosaic virus resistance gene Tm-2(2) from Lycopersicon esculentum.

Plant Mol Biol 2003 Jul;52(5):1037-49

Department Molecular Biology of Plants, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Kerklaan 30, 9751 NN Haren, The Netherlands.

In tomato, infections by tomato mosaic virus are controlled by durable Tm-2(2) resistance. In order to gain insight into the processes underlying disease resistance and its durability, we cloned and analysed the Tm-2(2) resistance gene and the susceptible allele, tm-2. The Tm-2(20 gene was isolated by transposon tagging using a screen in which plants with a destroyed Tm-2(2) gene survive. The Tm-2(2) locus consists of a single gene that encodes an 861 amino acid polypeptide, which belongs to the CC-NBS-LRR class of resistance proteins. The putative tm-2 allele was cloned from susceptible tomato lines via PCR with primers based on the Tm-2(2) sequence. Interestingly, the tm-2 gene has an open reading frame that is comparable to the Tm-2(2) allele. Between the tm-2 and the Tm-2(2) polypeptide 38 amino acid differences are present of which 26 are located in the second half of the LRR-domain. Susceptible tomato plants, which were transformed with the Tm-2(2) gene, displayed resistance against ToMV infection. In addition, virus specificity, displayed by the Tm-2(2) resistance was conserved in these transgenic lines. To explain the durability of this resistance, it is proposed that the Tm-2(2)-encoded resistance is aimed at the Achilles' heel of the virus.
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http://dx.doi.org/10.1023/a:1025434519282DOI Listing
July 2003

Arabidopsis RecQsim, a plant-specific member of the RecQ helicase family, can suppress the MMS hypersensitivity of the yeast sgs1 mutant.

Plant Mol Biol 2003 May;52(2):273-84

Molecular Biology of Plants, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Kerklaan 30, 9751 NN Haren, The Netherlands.

The Arabidopsis genome contains seven genes that belong to the RecQ family of ATP-dependent DNA helicases. RecQ members in Saccharomyces cerevisiae (SGS1) and man (WRN, BLM and RecQL4) are involved in DNA recombination, repair and genome stability maintenance, but little is known about the function of their plant counterparts. The Arabidopsis thaliana RecQsim gene is remarkably different from the other RecQ-like genes due to an insertion in its helicase domain. We isolated the AtRecQsim orthologues from rice and rape and established the presence of a similar insertion in their helicase domain, which suggests a plant specific function for the insert. The expression pattern of the AtRecQsim gene was compared with the other Arabidopsis RecQ-like members in different tissues and in response to stress. The transcripts of the AtRecQsim gene were found in all plant organs and its accumulation was higher in roots and seedlings, as compared to the other AtRecQ-like members. In contrast to most AtRecQ-like genes, the examined environmental cues did not have a detectable effect on the accumulation of the AtRecQsim transcripts. The budding yeast sgs1 mutant, which is known to be hypersensitive to the DNA-damaging drug MMS, was transformed with the AtRecQsim cDNA. The AtRecQsim gene suppressed the MMS hypersensitivity phenotype of the sgs1 cells. We propose that the Arabidopsis RecQsim gene, despite its unusual structure, exhibits an evolutionary conserved function.
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http://dx.doi.org/10.1023/a:1023968429220DOI Listing
May 2003

The plant disease resistance gene Asc-1 prevents disruption of sphingolipid metabolism during AAL-toxin-induced programmed cell death.

Plant J 2002 Nov;32(4):561-72

Department of Molecular Biology of Plants, Research School GBB, University of Groningen, Kerklaan 30, 9751NN Haren, the Netherlands.

The nectrotrophic fungus Alternaria alternata f.sp. lycopersici infects tomato plants of the genotype asc/asc by utilizing a host-selective toxin, AAL-toxin, that kills the host cells by inducing programmed cell death. Asc-1 is homologous to genes found in most eukaryotes from yeast to humans, suggesting a conserved function. A yeast strain with deletions in the homologous genes LAG1 and LAC1 was functionally complemented by Asc-1, indicating that Asc-1 functions in an analogous manner to the yeast homologues. Examination of the yeast sphingolipids, which are almost absent in the lag1Deltalac1Delta mutant, showed that Asc-1 was able to restore the synthesis of sphingolipids. We therefore examined the biosynthesis of sphingolipids in tomato by labeling leaf discs with l-[3-3H]serine. In the absence of AAL-toxin, there was no detectable difference in sphingolipid labeling between leaf discs from Asc/Asc or asc/asc leaves. In the presence of pathologically significant concentrations of AAL-toxin however, asc/asc leaf discs showed severely reduced labeling of sphingolipids and increased label in dihydrosphingosine (DHS) and 3-ketodihydrosphingosine (3-KDHS). Leaf discs from Asc/Asc leaves responded to AAL-toxin treatment by incorporating label into different sphingolipid species. The effects of AAL-toxin on asc/asc leaflets could be partially blocked by the simultaneous application of AAL-toxin and myriocin. Leaf discs simultaneously treated with AAL-toxin and myriocin showed no incorporation of label into sphingolipids or long-chain bases as expected. These results indicate that the presence of Asc-1 is able to relieve an AAL-toxin-induced block on sphingolipid synthesis that would otherwise lead to programmed cell death.
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http://dx.doi.org/10.1046/j.1365-313x.2002.01444.xDOI Listing
November 2002

Arabidopsis onset of leaf death mutants identify a regulatory pathway controlling leaf senescence.

Plant J 2002 Oct;32(1):51-63

Molecular Biology of Plants, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Kerklaan 30, 9751 NN, Haren, The Netherlands.

The onset of leaf senescence is controlled by leaf age and ethylene can promote leaf senescence within a specific age window. We exploited the interaction between leaf age and ethylene and isolated mutants with altered leaf senescence that are named as onset of leaf death (old) mutants. Early leaf senescence mutants representing three genetic loci were selected and their senescence syndromes were characterised using phenotypical, physiological and molecular markers. old1 is represented by three recessive alleles and displayed earlier senescence both in air and upon ethylene exposure. The etiolated old1 seedlings exhibited a hypersensitive triple response. old2 is a dominant trait and the mutant plants were indistinguishable from the wild-type when grown in air but showed an earlier senescence syndrome upon ethylene treatment. old3 is a semi-dominant trait and its earlier onset of senescence is independent of ethylene treatment. Analyses of the chlorophyll degradation, ion leakage and SAG expression showed that leaf senescence was advanced in ethylene-treated old2 plants and in both air-grown and ethylene-treated old1 and old3 plants. Epistatic analysis indicated that OLD1 might act downstream of OLD2 and upstream of OLD3 and mediate the interaction between leaf age and ethylene. A genetic model was proposed that links the three OLD genes and ethylene into a regulatory pathway controlling the onset of leaf senescence.
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http://dx.doi.org/10.1046/j.1365-313x.2002.01400.xDOI Listing
October 2002

Overexpression of the tomato Asc-1 gene mediates high insensitivity to AAL toxins and fumonisin B1 in tomato hairy roots and confers resistance to Alternaria alternata f. sp. lycopersici in Nicotiana umbratica plants.

Mol Plant Microbe Interact 2002 Jan;15(1):35-42

Department of Genetics, Free University, BioCentrum Amsterdam, The Netherlands.

The sphinganine-analog mycotoxins (SAMs) fumonisin B1 and AAL toxins are inhibitors of eukaryotic sphinganine N-acyltransferase in vitro. Treatment of eukaryotes with SAMs generally results in an accumulation of sphingoid base precursors and a depletion of complex sphingolipids. The asc,asc genotypes of tomato (Lycopersicon esculentum) and Nicotiana umbratica are sensitive to SAMs and host of the AAL toxin-producing fungus Alternaria alternata f. sp. lycopersici. Codominant insensitivity to SAMs in tomato is mediated by the Asc-1 gene, and sensitivity is associated with a frame-shift mutation present in asc-1. We investigated the function of Asc-1 in mediating insensitivity to SAMs and resistance to the fungus by overexpression of asc-1 and Asc-1. In this study, it is shown that overexpression of these genes did not lead to visual symptoms in tomato hairy roots and N. umbratica plants. Overexpression of asc-1 did not influence the (in)sensitivity to SAMs. Overexpression of Asc-1 in SAM-sensitive hairy roots and N. umbratica plants, however, mediated a high insensitivity to SAMs and resistance to plant infection by Alternaria alternata f. sp. lycopersici.
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http://dx.doi.org/10.1094/MPMI.2002.15.1.35DOI Listing
January 2002
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