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D Inzé

Publications and source records attributed to D Inzé.

At least 37 records · Page 2Linked to original sources
PubMed2000

CEO1, a new protein from Arabidopsis thaliana, protects yeast against oxidative damage.

The Saccharomyces cerevisiae strain WYT, deficient in the YAP1 transcription factor, was used in a molecular screen to identify genes from Arabidopsis thaliana that could overcome the oxidative stress-sensitive phenotype of these yeast cells. A cDNA named CEO1 increased the tolerance to oxidative damage caused by tert-butylhydroperoxide of both the Yap1(-) mutant and the wild-type yeast. Additionally, in Yap1(-) yeast, CEO1 also induced cross-tolerance to oxidative damage caused by hydrogen peroxide and diamide. CEO1 was assigned as being part of a small gene family that, until now, is exclusively restricted to plants. In Arabidopsis, CEO1 was produced in all organs, especially in roots and stems. By using the yeast two-hybrid system, proteins that specifically interact with CEO1 in yeast were identified, and putative DNA-binding proteins were consistently recovered.

Amino Acid Sequence↗
PubMed2000

Cloning and molecular analysis of two new sesquiterpene cyclases from Artemisia annua L.

Artemisia annua L. is the only source of artemisinin, a new promising antimalarial drug (Qinghaosu Antimalarial Coordinating Research Group, Chin. Med. J. 92 (1979) 811). Our efforts are focused on the overproduction of this valuable medicine by genetic engineered A. annua plants. Therefore, we decided to isolate the gene(s) encoding sesquiterpene cyclase(s) in A. annua as a first step in improving artemisinin yield. Four partial genomic clones, gASC21, gASC22, gASC23 and gASC24, were isolated through polymerase chain reaction (PCR) with degenerated primers based on homologous boxes present in sesquiterpene cyclases from divergent sources. Intron-exon organisation of those partial genomic clones was analysed and it was shown that A. annua contains a gene family for sesquiterpene cyclases. Based on gASC21, gASC22, gASC23 and gASC24 sequences, the full-length cDNA clones cASC34 and cASC125 were subsequently isolated by rapid amplification of cDNA ends PCR. The derived amino acid sequences of both full-length clones show high homology with sesquiterpene cyclases from plants. Reverse transcription-PCR analysis revealed transient and tissue specific expression patterns for cASC34 and cASC125, in contrast to the constitutively expressed 8-epicedrol synthase, a previously reported sesquiterpene cyclase from A. annua. Both cASC34 and cASC125 could only be detected in flowering plants when artemisinin concentration is at highest.

Journal Article↗
PubMed2000

Crystallization and preliminary X-ray crystallographic analysis of NADPH: azodicarbonyl/quinone oxidoreductase, a plant zeta-crystallin.

Arabidopsis thaliana P1 protein was crystallized using the hanging drop vapor-diffusion method in 0.1 M piperazine-1, 4-bis(2-ethanesulfonic acid) buffer, containing 14% polyethylene glycol 6000 and 0.2 M magnesium acetate at pH 6.5 and 20 degrees C. The crystals are orthorhombic and belong to the space group P2(1)2(1)2(1) with unit cell dimensions of a=49.8, b=122.4 and c=149. 9 A. The diffraction data up to 2.9 A were collected by a multiwire area detector.

Alkanesulfonic Acids↗
PubMed2000

Different expression of an S-adenosylmethionine synthetase gene in transgenic tobacco callus modifies alkaloid biosynthesis.

Transformed callus cultures of Nicotiana tabacum were generated in which the SAM-1 gene from Arabidopsis thaliana encoding S-adenosylmethionine synthetase (SAM-S), under the control of the 35S promoter, had been integrated. The presence of the SAM-1 gene was detected in all tested transformants and the SAM-S activity correlated with the accumulation of SAM in the tobacco callus cultures. Three distinct phenotypic classes were identified among the transgenic cell lines in relation to growth of the cells, structure of the calli, and level of SAM. Transgene silencing was observed in several cultivated transgenic calli and this phenomenon was correlated directly with a low level of SAM-1 mRNA accompanied by a decrease of the SAM-S activity. The transgenic calli overexpressing the SAM-1 gene accumulated a high SAM level. The modifications in SAM-S activity were reflected in the pattern of secondary products present in the different cell lines, thereby demonstrating that the flux through the biosynthetic pathway of a plant secondary product can be modified by means of genetic engineering.

Alkaloids↗
PubMed2000

Chemical inhibitors: a tool for plant cell cycle studies.

Synchrony provides a large number of cells at defined points of the cell cycle. Highly synchronised cells are powerful and effective tools for molecular analyses and for studying the biochemical events of the cell cycle in plants. Usually, plant cell suspensions can be synchronised by chemical agents, which arrest the cell cycle by acting on the driving forces of the cell cycle engine such as cyclin-dependent kinase activity, enzymes involved in DNA synthesis or proteolysis of cell cycle regulators or by acting on the cell cycle apparatus (mitotic spindle). The specificity, reversibility and efficiency of each type of cell cycle inhibitor are described and related to their mode of action.

Cell Cycle↗
PubMed2000

The Arabidopsis thaliana PIN1At gene encodes a single-domain phosphorylation-dependent peptidyl prolyl cis/trans isomerase.

A homologue of the human site-specific prolyl cis/trans isomerase PIN1 was identified in Arabidopsis thaliana. The PIN1At gene encodes a protein of 119 amino acids that is 53% identical with the catalytic domain of the human PIN1 parvulin. Steady-state PIN1At mRNA is found in all plant tissues tested. We show by two-dimensional NMR spectroscopy that the PIN1At is a prolyl cis/trans isomerase with specificity for phosphoserine-proline bonds. PIN1At is the first example of an eukaryotic parvulin without N- or C-terminal extensions. The N-terminal WW domain of 40 amino acids, typical of all the phosphorylation-dependent eukaryotic parvulins, is absent. However, triple-resonance NMR experiments showed that PIN1At contained a hydrophobic helix similar to the alpha1 helix observed in PIN1 that could mediate the protein-protein interactions.

Amino Acid Sequence↗
PubMed2000

The Rhodococcus fascians-plant interaction: morphological traits and biotechnological applications.

Rhodococcus fascians is a Gram-positive bacterium that infects dicotyledonous and monocotyledonous plants, leading to an alteration in the normal growth process of the host. The disease results from the modulation of the plant hormone balances, and cytokinins are thought to play an important role in the induction of symptoms. Generally, on the aerial parts of the plants, existing meristems were found to be most sensitive to the action of R. fascians, but, depending on the infection procedure, differentiated tissues as well gave rise to shoots. Similarly, in roots not only actively dividing cells, but also cells with a high competence to divide were strongly affected by R. fascians. The observed symptoms, together with the determined hormone levels in infected plant tissue, suggest that auxins and molecules of bacterial origin are also involved in leafy gall formation. The complexity of symptom development is furthermore illustrated by the necessary and continuous presence of the bacteria for symptom persistence. Indeed, elimination of the bacteria from a leafy gall results in the further development of the multiple embryonic buds of which it consists. This interesting characteristic offers novel biotechnological applications: a leafy gall can be used for germplasm storage and for plant propagation. The presented procedure proves to be routinely applicable to a very wide range of plants, encompassing several recalcitrant species.

Arabidopsis↗
PubMed2000

Dual action of the active oxygen species during plant stress responses.

Adaptation to environmental changes is crucial for plant growth and survival. However, the molecular and biochemical mechanisms of adaptation are still poorly understood and the signaling pathways involved remain elusive. Active oxygen species (AOS) have been proposed as a central component of plant adaptation to both biotic and abiotic stresses. Under such conditions, AOS may play two very different roles: exacerbating damage or signaling the activation of defense responses. Such a dual function was first described in pathogenesis but has also recently been demonstrated during several abiotic stress responses. To allow for these different roles, cellular levels of AOS must be tightly controlled. The numerous AOS sources and a complex system of oxidant scavengers provide the flexibility necessary for these functions. This review discusses the dual action of AOS during plant stress responses.

Air Pollutants↗
PubMed2000

Developmental expression of the Arabidopsis thaliana CycA2;1 gene.

The associations of cyclins with highly conserved cyclin-dependent kinases are key events in the regulation of cell cycle progression. The spatio-temporal expression of an Arabidopsis thaliana (L.) Heynh. mitotic cyclin, Arath;CycA2;1, was studied by histochemical beta-glucuronidase (GUS) analysis and in-situ hybridizations. The CycA2,1] promoter was active in the egg apparatus before fertilization. During embryogenesis, CycA2;1:gus expression was found in the embryo and the developing endosperm. Throughout plant development, CycA2;1 transcripts were found in both dividing and non-dividing cells, indicating that the expression of this cyclin is not a limiting factor for cell division. In the pericycle and stelar parenchyma, CycA2;1 transcripts were located at the xylem poles, a position that can be correlated with competence for lateral root formation. In addition, CycA2;1:gus expression was upregulated in roots by auxins and in the shoot apex by cytokinins. Transcription of CycA2;1 was shown by reverse transcription-polymerase chain reaction to be strongly induced by sucrose in A. thaliana cell suspensions.

Arabidopsis↗
PubMed2000

Expression of cell cycle regulatory genes and morphological alterations in response to salt stress in Arabidopsis thaliana.

Hyperosmotic stress severely affects plant growth and development. To examine the effect of salt stress on cell cycle activity in Arabidopsis thaliana (L.) Heynh., the transcriptional regulation of a cyclin-dependent kinase, CDC2aAt, and two mitotic cyclins, Arath;CycB1;1 and Arath;CycA2;1, was studied by using the beta-glucuronidase (gus) reporter gene. Moreover, the mRNA abundance of these cell cycle genes as well as CDC2bAt were monitored during salt stress. Upon NaCl treatment, the promoter activities and transcript levels of all cell cycle genes diminished initially in the shoot apex and were subsequently induced during salt-stress adaptation. Additionally, the promoter activities of CDC2aAt and CycA2;1 decreased in the vascular cylinder of the root in correlation with reduced lateral root formation. In the root tips, a regression of CDC2aAt, CycA2;1, and CycB1;1:gus expression was observed, concomitant with a shrinkage of the root meristem and inhibition of root growth. Our data indicate that salt stress interferes with cell cycle regulation at the transcriptional level, resulting in an adaptive growth response.

Arabidopsis↗
PubMed2000

Cloning of beta-1,3-glucanases expressed during Cichorium somatic embryogenesis.

Three different beta-1,3-glucanase cDNA fragments, CG1, CG2 and CG3, were obtained by RT-PCR from RNA isolated from Cichorium hybrid '474' leaf fragments cultured for 11 days under somatic embryogenesis-inducing conditions. When expressed in Escherichia coli the proteins encoded by the three cDNAs were recognized by antibodies raised against 38 kDa extracellular beta-1,3-glucanases studied previously (Helleboid et al., Planta 205 (1998) 56-63). The CG2 and CG3 cDNAs may represent expressed alleles of one gene because their sequences showed a very high identity (98.5%) and are only 70% identical with CG1. Southern blot analysis revealed the presence of 3-4 genes coding for beta-1,3-glucanases in the Cichorium genome. Expression analysis of the genes corresponding to the three clones analysed by semi-quantitative RT-PCR indicated that CG1 mRNAs were only detectable in Cichorium hybrid '474' leaf fragments from day 3 of somatic embryogenesis induction, whereas CG2-CG3 mRNAs were already present in non-induced leaf tissue of both the embryogenic hybrid '474' and a non-embryogenic genotype. The level of CG1 mRNAs was particularly high when embryogenic cells were dividing to produce embryos, and when the amount of callose deposited in cell walls surrounding embryogenic cells and young embryos decreased. These results indicate that expression of the CG1 gene is correlated to the somatic embryogenesis process and that it encodes a 38 kDa beta-1,3-glucanase protein that may be involved in the degradation of callose localized around embryogenic cells and young embryos. A full-length CG1 cDNA clone was obtained using 3' and 5' RACE-PCR, and its sequence revealed that it encodes a beta-1,3-glucanase that is equally homologous to both class III and class IV plant beta-1,3-glucanases.

Amino Acid Sequence↗
PubMed2000

Regulation of cyclin-dependent kinases in Arabidopsis thaliana.

In plants, different families of cyclin-dependent kinases (CDKs) and cyclins have been identified, indicating that also in plants the progression through the cell cycle is regulated by CDKs. In all eukaryotes, CDKs exert their activity through well-controlled phosphorylations of specific substrates on serine/threonine residues. Such post-translational modifications are universal mechanisms in signal transduction pathways. They allow the organism to differentiate, regulate growth and/or adapt to environmental changes, the latter being crucial for plants because of their sedentary life-style. This adaptation might explain the occurrence of a special CDK type with plant-specific features. This review focuses on the involvement of plant CDKs in different phases of the cell cycle in Arabidopsis thaliana and outlines their regulation by binding to other proteins, and by phosphorylation and dephosphorylation.

Arabidopsis↗
PubMed2000

CDK-related protein kinases in plants.

Cyclin-dependent kinases (CDK) form a conserved superfamily of eukaryotic serine-threonine protein kinases, which require binding to a cyclin protein for activity. CDK are involved in different aspects of cell biology and notably in cell cycle regulation. The comparison of nearly 50 plant CDK-related cDNAs with a selected set of their animal and yeast counterparts reveals five classes of these genes in plants. These are described here with respect to their phylogenetic, structural and functional properties. A plant-wide nomenclature of CDK-related genes is proposed, using a system similar to that of the plant cyclin genes. The most numerous class, CDKA, includes genes coding for CDK with the PSTAIRE canonical motif. CDKB makes up a class of plant-specific CDK divided into two groups: CDKB1 and CDKB2. CDKC, CDKD and CDKE form less numerous classes. The CDKD class includes the plant orthologues of metazoan CDK7, which correspond to the CDK-activating kinase (CAK). At present, no functional information is available in plants for CDKC and CDKE.

Amino Acid Motifs↗
PubMed2000

Cell division events are essential for embryo patterning and morphogenesis: studies on dominant-negative cdc2aAt mutants of arabidopsis.

During plant development, cell division events are coordinately regulated, leading to specific growth patterns. Experimental evidence indicates that the morphogenetic controls that act at the vegetative plant growth stage are flexible and tolerate distortions in patterns and frequencies of cell division. To address questions concerning the relationship between cell division and embryo formation, a novel experimental approach was used. The frequencies of cell division were reduced exclusively during embryo development of Arabidopsis by the expression of a dominant cdc2a mutant. The five independent transgenic lines with the highest levels of the mutant cdc2a affected embryo formation. In the C13 line, seeds failed to germinate. The C1, C5 and C12 lines displayed a range of distortions on the apical-basal embryo pattern. In the C3 line, the shoot apical meristem of the seedlings produced leaves defective in growth and with an incorrect phyllotactic pattern. The results demonstrate that rates of cell division do not dictate cellular differentiation of embryos. Nevertheless, whereas cell divisions are uncoupled from vegetative development, they are instrumental in elaborating embryo structures and modulating embryo and seedling morphogenesis.

Arabidopsis↗
PubMed2000

A novel NADPH:diamide oxidoreductase activity in arabidopsis thaliana P1 zeta-crystallin.

The zeta-crystallin (ZCr) gene P1 of Arabidopsis thaliana, known to confer tolerance toward the oxidizing drug 1,1'-azobis(N, N-dimethylformamide) (diamide) to yeast [Babiychuk, E., Kushnir, S., Belles-Boix, E., Van Montagu, M. & Inzé, D. (1995) J. Biol. Chem. 270, 26224], was expressed in Escherichia coli to characterize biochemical properties of the P1-zeta-crystallin (P1-ZCr). Recombinant P1-ZCr, a noncovalent dimer, showed NADPH:quinone oxidoreductase activity with specificity to quinones similar to that of guinea-pig ZCr. P1-ZCr also catalyzed the divalent reduction of diamide to 1,2-bis(N,N-dimethylcarbamoyl)hydrazine, with a kcat comparable with that for quinones. Two other azodicarbonyl compounds also served as substrates of P1-ZCr. Guinea-pig ZCr, however, did not catalyze the azodicarbonyl reduction. Hence, plant ZCr is distinct from mammalian ZCr, and can be referred to as NADPH:azodicarbonyl/quinone reductase. The quinone-reducing reaction was accompanied by radical chain reactions to produce superoxide radicals, while the azodicarbonyl-reducing reaction was not. Specificity to NADPH, as judged by kcat/Km, was > 1000-fold higher than that to NADH both for quinones and diamide. N-Ethylmaleimide and p-chloromercuribenzoic acid inhibited both quinone-reducing and diamide-reducing activities. Both NADPH and NADP+ suppressed the inhibition, but NADH did not, suggesting that sulfhydryl groups reside in the binding site for the phosphate group on the adenosine moiety of NADPH. The diamide-reducing activity of P1-ZCr accounts for the tolerance of P1-overexpressing yeast to diamide. Other possible physiological functions of P1-ZCr in plants are discussed.

Amino Acid Sequence↗
PubMed2000

Increased leakiness of the tetracycline-inducible Triple-Op promoter in dividing cells renders it unsuitable for high inducible levels of a dominant negative CDC2aAt gene.

A tetracycline-inducible promoter system was used to generate transgenic tobacco plants that confer inducible expression of the wild type or a dominant negative allele of the gene coding for the cyclin-dependent kinase (CDK) of Arabidopsis thaliana CDC2aAt. Although the total extractable CDK activity was doubled, the induced expression of the wild-type CDC2aAt did not correlate with any change of the cell cycle kinetics. An increase of CDK activity upon CDC2aAt expression was only seen in dividing cell populations, demonstrating that CDC2aAt expression itself is not sufficient to induce CDK activation. Induced expression of the dominant negative CDC2aAt.N146 correlated with a reduction of CDK activity to 66% of the level found in non-induced cells. This decrease was not sufficient to block cell division. The isolation of plants showing only low inducible levels of CDC2aAt.N146 suggests that a counterselection against strong inducible lines had occurred. Accordingly, Triple-Op promoter activity was found in dividing cells in the absence of tetracycline.

Arabidopsis↗