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Biomedical subjects

D Inzé

Publications and source records attributed to D Inzé.

At least 109 records · Page 6Linked to original sources
PubMed1995

Differential in vitro DNA binding activity to a promoter element of the gn1 beta-1,3-glucanase gene in hypersensitively reacting tobacco plants.

In a hypersensitive reaction to pathogen infection, expression of the beta-1,3-glucanase gn1 gene is induced in cells surrounding the necrotic lesions. The 5'-flanking sequence of gn1 was examined to investigate the molecular basis controlling activation of gene expression during this plant defense response. Studies on transgenic tobacco plants containing gn1 promoter deletions fused to the beta-glucuronidase reporter gene revealed the presence of negative and positive regulatory sequences mediating both the level and the spatial distribution of gn1 expression. Promoter sequences to -138 bp were sufficient to confer increased gene expression around the necrotic lesions produced in response to Pseudomonas syringae pv. syringae inoculation. It is demonstrated by electrophoretic mobility shift assays that nuclear proteins in both healthy and hypersensitively reacting tobacco leaves interact with DNA sequences within the regulatory elements identified. Among the binding sequences characterized, the promoter region extending from -250 to -217 bp contained the DNA motif -GGCGGC- found to be conserved in most if not all promoters of genes encoding pathogenesis-related basic proteins. The activity bound by this promoter sequence was stronger in hypersensitively responding tissues than in healthy untreated tobacco leaves.

Base Sequence↗
PubMed1995

Superroot, a recessive mutation in Arabidopsis, confers auxin overproduction.

We have isolated seven allelic recessive Arabidopsis mutants, designated superroot (sur1-1 to sur1-7), displaying several abnormalities reminiscent of auxin effects. These characteristics include small and epinastic cotyledons, an elongated hypocotyl in which the connection between the stele and cortical and epidermal cells disintegrates, the development of excess adventitious and lateral roots, a reduced number of leaves, and the absence of an inflorescence. When germinated in the dark, sur1 mutants did not develop the apical hook characteristic of etiolated seedlings. We were able to phenocopy the Sur1- phenotype by supplying auxin to wild-type seedlings, to propagate sur1 explants on phytohormone-deficient medium, and to regenerate shoots from these explants by the addition of cytokinins alone to the culture medium. Analysis by gas chromatography coupled to mass spectrometry indicated increased levels of both free and conjugated indole-3-acetic acid. sur1 was crossed to the mutant axr2 and the altered-auxin response mutant ctr1. The phenotype of both double mutants was additive. The sur1 gene was mapped on chromosome 2 at 0.5 centimorgans from the gene encoding phytochrome B.

Arabidopsis↗
PubMed1994

Three discrete classes of Arabidopsis cyclins are expressed during different intervals of the cell cycle.

We have isolated cDNAs encoding four additional mitotic-like cyclins from Arabidopsis: cyc2aAt, cyc2bAt, cyc3aAt, and cyc3bAt. Examination of amino acid sequences deduced from plant cyclin cDNAs isolated so far showed that they can be grouped into three distinct classes. The members of each plant cyclin family are more related to each other than to any animal or yeast cyclin. Reverse transcription-PCR analysis demonstrated that cyc2aAt was expressed in all plant organs, whereas cyc2bAt mRNAs were found only in roots; cyc3aAt was not expressed in leaves and was barely expressed in flowers. On the other hand, cyc3bAt transcripts were observed in all organs. Whole-mount in situ hybridizations on roots showed that the cyclin mRNAs were confined to parts of the roots with mitotic activity. Furthermore, results of whole-mount in situ hybridizations on roots treated with either oryzalin or hydroxyurea suggest that the different cyclin classes have distinct functions in the cell cycle.

Amino Acid Sequence↗
PubMed1994

Differential expression of catalase genes in Nicotiana plumbaginifolia (L.).

We have analyzed the expression of three catalase (Cat; EC 1.11.1.6) genes from Nicotiana plumbaginifolia by means of RNA blot and in situ hybridizations. Our data demonstrate that the expression of each catalase is associated with a particular H2O2-producing process. Cat1 appears to be specifically involved in the scavenging of photorespiratory H2O2 and is under control of a circadian rhythm, Cat2 is uniformly expressed in different organs with a cellular preference for vascular tissues, and the expression profile of Cat3 points to a role in glyoxysomal processes. Differential expression of these catalases is also manifested in response to temperature changes. DNA sequence comparison with other dicotyledonous catalases led to the identification of at least three distinct classes, which indicates that the functional organization of catalases is generally conserved in dicotyledonous plants.

Catalase↗
PubMed1994

Molecular identification of catalases from Nicotiana plumbaginifolia (L.).

We have isolated three different catalase cDNAs from Nicotiana plumbaginifolia (cat1, cat2, and cat3) and a partial sequence of a fourth catalase gene (cat4) that shows no discernible expression based on Northern analysis. The catalase sequences were used to determine the similarity with other plant catalases and to study the transcriptional response to paraquat, 3-aminotriazole, and salicylic acid. 3-Aminotriazole induces mRNA levels of cat1, cat2 and cat3, indicating that a reduction in catalase activity positively affects catalase mRNA abundance. Salicylic acid that binds catalase in vitro, had no effect on catalase transcript levels at physiological concentrations. Paraquat resulted in the induction of cat1.

Amino Acid Sequence↗
PubMed1994

The Arabidopsis thaliana apurinic endonuclease Arp reduces human transcription factors Fos and Jun.

An Arabidopsis thaliana cDNA encoding an analogue, referred to as Arp for apurinic endonuclease-redox protein, of the human redox factor REF has been cloned. Arp stimulates in vitro DNA-binding activity of the human transcription factor Jun and Fos by the reduction of a cysteine residue located in the DNA-binding domain. Based on amino acid sequence homology, this redox activity is probably confined to the small internal domain of the Arp protein. In analogy to REF, we show that the Arabidopsis Arp protein also functions as an apurinic/apyrimidinic class II endonuclease. This base-free endonuclease activity resides in the carboxyl-terminal domain, and this part of the protein has significant sequence similarity to bacterial (Escherichia coli exonuclease III and Streptococcus pneumoniae exonuclease A) and animal (Drosophila Rrp1 and human REF/HAP) apurinic/apyrimidinic endonucleases. The amino-terminal domain of the Arp protein is highly charged and apparently increases the affinity of the protein for DNA. Therefore, the Arabidopsis Arp protein is multifunctional and may be involved both in DNA repair and in the regulation of transcription.

Arabidopsis↗
PubMed1994

Control of cell proliferation during plant development.

Knowledge of the control of cell division in eukaryotes has increased tremendously in recent years. The isolation and characterization of the major players from a number of systems and the study of their interactions have led to a comprehensive understanding of how the different components of the cell cycle apparatus are brought together and assembled in a fine-tuned machinery. Many parts of this machine are highly conserved in organisms as evolutionary distant as yeast and animals. Some key regulators of cell division have also been identified in higher plants and have been shown to be functional homologues of the yeast or animal proteins. Although still in its early days, investigations into the regulation of these molecules have provided some clues on how cell division is coupled to plant development.

Animals↗
PubMed1994

Extensin gene expression is induced by mechanical stimuli leading to local cell wall strengthening in Nicotiana plumbaginifolia.

Nicotiana plumbaginifolia Viv. harbors a single extensin gene, although related hydroxyproline-rich sequences are present in the genome. Northern analysis showed that the gene is highly expressed in roots and to a lesser extent in stems. Expression in leaves is low but mRNA levels are increased upon infection with the incompatible bacterium Pseudomonas syringae. Extensin transcript levels in leaves were slightly enhanced after wounding and salicylic acid treatment. In-situ hybridization experiments showed high accumulation of extensin mRNA in cells which, at certain stages of development, require reinforcement of their cell walls. The cortical cells in stem nodes and roots, which are put under severe mechanical stress by adjacent developing tissues, tend to express the gene to high levels. Immunolocalization of the extensin protein in stems and roots demonstrated a close association of the protein with lignin deposition. Mature tissues contained more extensin than younger tissues. The extensin promoter was fused to the beta-glucuronidase gene.

Biomechanical Phenomena↗
PubMed1994

Developmental and environmental regulation of the Nicotiana plumbaginifolia cytosolic Cu/Zn-superoxide dismutase promoter in transgenic tobacco.

Superoxide dismutases (SODs) play a key role in the cellular defense against reactive oxygen species. To study the transcriptional regulation at the cellular level, the promoter of the Nicotiana plumbaginifolia cytosolic gene encoding Cu/ZnSOD (SODCc) was fused to the beta-glucuronidase (GUS) reporter gene (gusA) and analyzed in transgenic tobacco plants. The promoter was highly active in vascular bundles of leaves and stems, where it is confined to phloem cells. In flowers, GUS activity was detected in ovules and pollen grains, in pigmented tissues of petals, and in vascular tissue of ovaries and anthers. In response to treatment with the superoxide-generating herbicide paraquat, very strong GUS staining was observed in photosynthetically active cells of leaves and in some epidermal root cells of seedlings. The expression of the SODCc-gusA was also induced in seedlings after heat shock and chilling and after treatment with sulfhydryl antioxidants such as reduced glutathione and cysteine. It is postulated that SODCc expression is directly linked to a cell-specific production of excess superoxide radicals in the cytosol.

Base Sequence↗
PubMed1994

Distinct phenotypes generated by overexpression and suppression of S-adenosyl-L-methionine synthetase reveal developmental patterns of gene silencing in tobacco.

S-Adenosyl-L-methionine synthetase (SAM-S) catalyzes the conversion of L-methionine and ATP into S-adenosyl-L-methionine. Tobacco plants that were transformed with a construct allowing high transcription levels of an Arabidopsis sam-s gene could be grouped into two main classes based on their morphology. One class developed yellow-green leaves and had high SAM-S activity and transgene mRNA levels, whereas the other class was stunted and had leather-like leaves, very low SAM-S activity, and suppressed mRNA level of the transgene. Because both overexpression and silencing of transgene expression led to distinct, abnormal phenotypes, the developmental pattern of transgene silencing was visualized. In the lower leaves, the suppressed phenotype was associated with the veins. In successive leaves, the area of the suppressed tissue increased until all newly developed leaves displayed the suppressed phenotype. In this study, a hypothesis is presented for this developmental gene silencing. Furthermore, transgenic plants with suppressed SAM-S activity had a characteristic smell, a consequence of the accumulation of L-methionine that is converted into the volatile methanethiol.

Amino Acids↗
PubMed1994

Developmental expression of the arabidopsis cyclin gene cyc1At.

In eukaryotes, the control of cell cycle progression is exercised by heteromeric protein kinase complexes composed of a cell cycle-dependent, kinase-related subunit (Cdc2) and a cyclin subunit. To explore the possibility that cyclin transcription plays a role in the developmental regulation of cell division, we examined the spatial and temporal expression of a cyclin gene (cyc1At) in Arabidopsis. In root and shoot apical meristems and during embryogenesis, cyc1At expression is almost exclusively confined to dividing cells. A cell-specific pattern of cyc1At expression was noticed in root meristems. We examined the effects of induction of cell division of differentiated cells on cyc1At expression. During lateral root formation, induction of cyc1At expression is a very early event and was detected before anatomical modifications were visible. Treatment of roots with oryzalin, which blocks cell division in metaphase, did not inhibit the auxin induction of cyc1At, suggesting that induction of cyc1At expression precedes the completion of the first division cycle after induction of lateral roots. In tobacco protoplasts, an increase in cyc1At expression was observed only when cell division was induced. Together, the results suggest that Cyc1At accumulation in Arabidopsis is transcriptionally regulated and might be one of the limiting factors for the activation of cell division.

Arabidopsis↗
PubMed1993

A negatively light-regulated gene from Arabidopsis thaliana encodes a protein showing high similarity to blue copper-binding proteins.

A negatively photo-regulated gene (bcb) has been isolated by means of differential hybridization of a genomic library of Arabidopsis thaliana. In mature plants, a 20-fold increase in the amount of steady-state bcb mRNA can be detected upon 48 h of dark adaption. The expression level of the gene is also dependent upon the developmental stage of the plant. The 21.5-kDa gene product (BCB) shows extensive similarity with blue Cu(2+)-binding proteins such as plastocyanin and stellacyanin. The protein abundance increases only twofold upon dark adaption, which implies the presence of post-transcriptional control. The isolation of a novel negatively photoregulated gene allows us to investigate the complex expression profile of genes responding to the absence of light.

Amino Acid Sequence↗
PubMed1993

One-step purification and characterization of a lignin-specific O-methyltransferase from poplar.

O-Methyltransferases (OMT; EC 2.1.1.6) play an important role in the synthesis of lignin precursors by catalyzing the O-methylation of o-diphenolic substrates such as caffeic acid (CA) and 5-hydroxyferulic acid (5OH). Here, we report on the purification of a lignin-specific OMT (38 kDa) from poplar (Populus trichocarpa x P. deltoides). The OMT was purified from xylem by a single affinity chromatography step on adenosine agarose. The enzyme uses both CA and 5OH as substrates. We previously have reported the cloning of a corresponding OMT cDNA [Dumas et al., Plant Physiol. 98 (1992) 796-797]. Expression of this OMT cDNA in Escherichia coli further confirmed the identity of the clone. Genomic hybridization demonstrates the presence of one or two OMT genes per haploid poplar genome. RNA gel blot hybridization shows high levels of steady-state OMT mRNA in the xylem of young poplar trees, as compared to the levels in leaves.

Blotting, Northern↗
PubMed1993

Redox-activated expression of the cytosolic copper/zinc superoxide dismutase gene in Nicotiana.

Superoxide dismutases (SODs; superoxide: superoxide oxidoreductase, EC 1.15.1.1) play a key role in protection against oxygen radicals, and SOD gene expression is highly induced during environmental stress. To determine the conditions of SOD induction, the promoter of the cytosolic copper/zinc SOD (Cu/ZnSODcyt) gene was isolated in Nicotiana plumbaginifolia and fused to the beta-glucuronidase reporter gene. Oxidative stress is likely to alter the cellular redox in favor of the oxidized status. Surprisingly, the expression of the Cu/ZnSODcyt gene is induced by sulfhydryl antioxidants such as reduced glutathione, cysteine, and dithiothreitol, whereas the oxidized forms of glutathione and cysteine have no effect. It is therefore possible that reduced glutathione directly acts as an antioxidant and simultaneously activates the Cu/ZnSODcyt gene during oxidative stress.

Amino Acid Sequence↗