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PubMed1999

The promoter of the strictosidine synthase gene from periwinkle confers elicitor-inducible expression in transgenic tobacco and binds nuclear factors GT-1 and GBF.

Strictosidine synthase (STR) is a key enzyme in the biosynthesis of terpenoid indole alkaloids. This class of secondary metabolites harbours several pharmaceutically important compounds used, among other applications, in cancer treatment. Terpenoid indole alkaloid biosynthesis and expression of biosynthetic genes including Str1 is induced by fungal elicitors. To identify elicitor-responsive regulatory promoter elements and trans-acting factors, the single-copy Str1 gene was isolated from the subtropical plant species Catharanthus roseus (Madagascar periwinkle). Str1 upstream sequences conferred elicitor-responsive expression to the beta-glucuronidase (gusA) reporter gene in transgenic tobacco plants. Main enhancer sequences within the Str1 promoter region studied were shown to be located between -339 and -145. This region and two other regions of the promoter bound the tobacco nuclear protein factor GT-1. A G-box located around position -105 bound nuclear and cloned G-box-binding factors (GBFs). A mutation that knocked out GBF binding had no measurable effect on expression, which indicates that the G-box is not essential for the elicitor responsiveness of the Str1 promoter. No obvious homologies with promoter elements identified in other elicitor-responsive genes were observed, suggesting that the Str1 gene may depend on novel regulatory mechanisms.

Base Sequence↗

PubMed1999

Selectable marker-free transgenic plants without sexual crossing: transient expression of cre recombinase and use of a conditional lethal dominant gene.

Transgenic tobacco plants were produced that contained single-copy pART54 T-DNA, with a 35S-uidA gene linked to loxP-flanked kanamycin resistance (nptII) and cytosine deaminase (codA) genes. Retransformation of these plants with pCre1 (containing 35S transcribed cre recombinase and hygromycin (hpt) resistance genes) resulted in excision of the loxP-flanked genes from the genome. Phenotypes of progeny from selfed-retransformed plants confirmed nptII and codA excision and integration of the cre-linked hpt gene. To avoid integration of the hpt gene, and thereby generate plants totally free of marker genes, we attempted to transiently express the cre recombinase. Agrobacterium tumefaciens (pCre1) was cocultivated with leaf discs of two pART54-transformed lines and shoots were regenerated in the absence of hygromycin selection. Nineteen of 773 (0.25%) shoots showed tolerance to 5-fluorocytosine (5-fc) which is converted to the toxic 5-fluorouracil by cytosine deaminase. 5-fc tolerance in six shoots was found to be due to excision of the loxP-flanked region of the pART54 T-DNA. In four of these shoots excision could be attributed to cre expression from integrated pCre1 T-DNA, whereas in two shoots excision appeared to be a consequence of transient cre expression from pCre1 T-DNA molecules which had been transferred to the plant cells but not integrated into the genome. The absence of selectable marker genes was confirmed by the phenotype of the T1 progeny. Therefore, through transient cre expression, marker-free transgenic plants were produced without sexual crossing. This approach could be applicable to the elimination of marker genes from transgenic crops which must be vegetatively propagated to maintain their elite genotype.

Agrobacterium tumefaciens↗

PubMed1999

Interaction between composite elements in the napA promoter: both the B-box ABA-responsive complex and the RY/G complex are necessary for seed-specific expression.

During seed maturation, the transcriptional activity of napin genes is regulated by developmental signals involving the transcriptional activator ABI3 and abscisic acid (ABA). To localize cis elements involved in the seed-specific activity of the napin napA promoter, a systematic analysis was performed focusing on two major element complexes, the B-box and RY/G. Substitution mutation analysis using promoter-reporter gene fusions in stable transgenic tobacco showed synergistic interactions between elements within these complexes. The distal part of the B-box shows similarities to abscisic acid response elements and the proximal portion contains a CA-rich element. In vitro studies involving Exonuclease III protection and electrophoretic mobility shift assays revealed binding by nuclear proteins to elements within the B-box. The distal and proximal parts of the B-box were found to bind distinct nuclear protein complexes. By gain-of-function analysis with a tetramer of the B-box fused to a truncated (-46) cauliflower mosaic virus (CaMV) 35S minimal promoter, it was demonstrated that the B-box mediates strong activity in seeds. Further, it was shown that the elements in the B-box constitute an ABA-responsive complex, since the B-box tetramer mediates ABA-responsiveness in vegetative tissues to a construct containing the CaMV virus 35S enhancer (-343 to -90). Thus, the seed-specific activity of the napA promoter relies on the combinatorial interaction between the RY/G complex and the B-box ABA-responsive complex during the ABA response in seed development.

2S Albumins, Plant↗

PubMed1999

A tobacco cryptic constitutive promoter, tCUP, revealed by T-DNA tagging.

We have isolated a constitutive promoter sequence, tCUP, from tobacco by T-DNA tagging using a promoterless GUS-nos3' reporter gene construct. The T-DNA integration event produced a translational fusion with the GUS gene that is expressed widely in organs, at both the mRNA and enzyme activity levels. In tobacco transformed with a tCUP-GUS-nos3' gene, GUS specific activity in leaves was within a range of values similar to those of plants transformed with the widely used constitutive promoter gene fusion, CaMV 35S promoter-GUS-nos3'. Characteristics of the tCUP promoter sequence differ from those of other plant constitutive promoters; for instance, the tCUP sequence lacks a TATA box. Transcription initiates at a single site within the tCUP sequence which is similar to a transcriptional start site consensus sequence determined for plant genes. The tCUP promoter is cryptic as RNA accumulation at the transcriptional start site is not detected in untransformed tobacco. Thus, tCUP is the first example of a cryptic, constitutive promoter isolated from plants. The tCUP-GUS-nos3' gene fusion produced GUS activity in tissues of all species tested suggesting that tCUP may utilize fundamental transcription mechanisms found in plants.

Amino Acid Sequence↗

PubMed1999

A constitutively expressed Myc-like gene involved in anthocyanin biosynthesis from Perilla frutescens: molecular characterization, heterologous expression in transgenic plants and transactivation in yeast cells.

The coordinate expression of anthocyanin biosynthetic genes in leaves and stems of a red forma of Perilla frutescens is presumably controlled by regulatory gene(s). A Myc-like gene (Myc-rp) was isolated from a cDNA library prepared from the leaves of red P. frutescens, and its deduced amino acid sequence shows 64% identity with that of delila from snapdragon. The Myc-rp gene was expressed in leaves and roots of both red and green P. frutescens equally. Comparison of deduced amino acid sequence of Myc-rp with that of Myc-gp, the second allele isolated from a green forma of P. frutescens, indicates that the 132nd amino acid, alanine, existing in MYC-RP was changed to serine in MYC-GP. The heterologous expression of these two alleles of Myc-like gene in tobacco and tomato resulted in an increase of the anthocyanin contents in flowers of tobacco and vegetative tissues and flowers of tomato. However, the flowers of transgenic tobacco expressing the fragment with a partial deletion (encoding 1-115 amino acids deleted) of Myc(-gp gave no change in anthocyanin accumulation, but some morphological changes of the flower were observed. In yeast, the MYC-RP/GP and Delila protein exhibited transactivation activity on the GAL-1 promoter from yeast and the promoter of dihydroflavonol 4-reductase (DFR) gene from P. frutescens. A transactivation domain of MYC-RP/GP and Delila could be located in the region between the 193rd and the 420th amino acid of MYC-RP/GP proteins. Our data indicate that this Myc-like gene presumably functions in the regulation of anthocyanin biosynthesis similarly in different tissues of dicot plants.

Alcohol Oxidoreductases↗

PubMed1999

VsENBP1 regulates the expression of the early nodulin PsENOD12B.

A DNA-binding protein, VsENBP1, previously isolated from Vicia sativa was shown to bind in a sequence-specific manner to the early nodulin ENOD12 gene promoter from Pisum sativum. Here, the functional importance of the VsENBP1 binding sites on the PsENOD12B promoter has been studied in vivo. A promoter-gusA fusion in which a mutation was introduced at the putative target sequence, AATAA, was inactive in nodules of transgenic Vicia hirsuta roots. Gel retardation assays showed that VsENBP1 does not bind to the mutated promoter segment, suggesting that VsENBP1 activates the PsENOD12B expression in nodules through its interaction with its target sequence. In the presence of the 35S enhancer, an ENOD12 promoter-GUS construct gave expression in root vascular tissue in addition to the root nodules. Overexpression of Vsenbp1 in transgenic V. hirsuta roots reduced the leaky expression in root vascular tissue in contrast to nodules in which a small increase in GUS expression was observed. The results indicate that VsENBP1 acts as a repressor of ENOD12 expression in root tissue.

Base Sequence↗

PubMed1999

Secondary xylem-specific expression of caffeoyl-coenzyme A 3-O-methyltransferase plays an important role in the methylation pathway associated with lignin biosynthesis in loblolly pine.

Two types of structurally distinct O-methyltransferases mediate the methylation of hydroxylated monomeric lignin precursors in angiosperms. Caffeate 3-O-methyltransferase (COMT; EC 2.1.1.68) methylates the free acids and caffeoyl CoA 3-O-methyltransferase (CCoAOMT; EC 2.1.1.104) methylates coenzyme A esters. Recently, we reported a novel hydroxycinnamic acid/hydroxycinnamoyl CoA ester O-methyltransferase (AEOMT) from loblolly pine differentiating xylem that was capable of methylating both acid and ester precursors with similar efficiency. In order to determine the possible existence and role of CCoAOMT in lignin biosynthesis in gymnosperms, a 1.3 kb CCoAOMT cDNA was isolated from loblolly pine that showed 79-82% amino acid sequence identity with many angiosperm CCoAOMTs. The recombinant CCoAOMT expressed in Escherichia coli exhibited a significant methylating activity with hydroxycinnamoyl CoA esters whereas activity with hydroxycinnamic acids was insignificant. Moreover, 3.2 times higher catalytic efficiency for methylating caffeoyl CoA over 5-hydroxyferuloyl CoA was observed which could serve as a driving force towards synthesis of guaiacyl lignin. The secondary xylem-specific expression of CCoAOMT was demonstrated using RNA blot analysis, western blot analysis, and O-methyltransferase enzyme assays. In addition, Southern blot analysis indicated that CCoAOMT may exist as a single-copy gene in loblolly pine genome. The transgenic tobacco plants carrying loblolly pine CCoAOMT promoter-GUS fusion localized the site of GUS activity at the secondary xylem tissues. These data suggest that CCoAOMT, in addition to AEOMT, plays an important role in the methylation pathway associated with lignin biosynthesis in loblolly pine.

Amino Acid Sequence↗

PubMed1999

Overexpression of a cytoplasm-localized allene oxide synthase promotes the wound-induced accumulation of jasmonic acid in transgenic tobacco.

Jasmonic acid (JA) is involved in regulating the expression of certain plant defense genes and response to various stresses. JA biosynthesis is hypothesized to occur both in chloroplasts and the cytoplasm. In order to test whether or not a cytosol-localized allene oxide synthase (AOS) can promote JA biosynthesis, transgenic tobacco plants containing a flax AOS cDNA without a chloroplast transit sequence under the control of a tetracycline-inducible promoter were generated. Induction of the flax AOS gene in transgenic plants with chlor-tetracycline (Tc) led to the expression of the flax AOS mRNA and protein, which resulted in high level of metabolism of 13(S)-hydroperoxyoctadecatrienoic acid (13(S)-HPOT) and formation of 12-oxo-phytodienoic acid (12-O-PDA). Subcellular fractionation demonstrated that the flax AOS protein and activity were associated with the cytosol. Overexpression of the flax AOS in induced transgenic plants did not increase JA levels in healthy, undamaged leaf tissues. However, in wounded tissues overexpressing a flax AOS, levels of JA and the transcript of a pathogenesis-related gene (PR-1) dramatically increased when compared to those not expressing the flax AOS. Analysis of the release of wound-induced C6 volatiles showed that the level of (Z)-3-hexen-1-ol decreased about 30% due to overexpression of the cytoplasm-localized AOS, while (Z)-3-hexenal and (Z)-3-hexenyl acetate appeared not to be significantly altered. The data indicate that cytoplasmic AOS responds to wounding by increasing the levels of the wound-induced JA which in turn directly or indirectly enhances the expression of plant defense genes.

Chloroplasts↗

PubMed1999

Structure and promoter/leader deletion analysis of mirabilis mosaic virus (MMV) full-length transcript promoter in transgenic plants.

A full-length transcript (FLt) promoter fragment was isolated from a genomic clone of mirabilis mosaic virus (MMV), a double-stranded DNA plant pararetrovirus belonging to the caulimovirus family. The boundaries required for maximal promoter expression were defined by 5' and 3' deletion analysis of the MMV promoter fragments coupled to a GUS reporter gene. The expression patterns of these chimeric gene constructs were evaluated both in transgenic Nicotiana tabacum cv. Samsun NN plants and in protoplast transient expression experiments. A 360 bp FLt promoter fragment (sequence -297 to +63 from the transcription start site) was found sufficient for strong promoter activity. The transcription start site (TSS) of the MMV FLt promoter was determined by primer extension analysis using total RNA isolated from transgenic plants containing a MMV promoter:uidA fusion gene. Analysis of the 5' and 3' deletion constructs showed that an upstream region (sequence -248 to -193 from the transcription start site) is required for the MMV FLt promoter activity along with the as-1, TATA box regions. In addition, a 31 bp sequence (+33 to +63 from the transcription start site) located downstream of a TATA box is also essential for the maximum expression of the MMV FLt promoter. Analysis of transcripts (mRNA) from these chimeric constructs also indicated that the MMV FLt promoter fragment (-297 to +63 from the transcription start site) has the highest promoter activity. In a comparative analysis the MMV FLt promoter showed much greater activity than the CaMV 35S promoter.

3' Untranslated Regions↗

PubMed1999

Identification of UV-B light-responsive regions in the promoter of the tryptophan decarboxylase gene from Catharanthus roseus.

The tryptophan decarboxylase (Tdc) gene encodes a key enzyme in the biosynthesis of terpenoid indole alkaloids (TIAs) in Catharanthus roseus. TIAs absorb ultraviolet light (UV) and putative functions in plants include a role as UV protectants. In support of this possible function we demonstrate here that UV light induces accumulation of several TIAs as well as expression of the Tdc gene in C. roseus. In addition, in tobacco a Tdc-gusA construct was found to be specifically induced by UV-B light. Lack of induction by UV-A or other wavelengths of light indicate that Tdc expression is regulated by a specific UV-B receptor and corresponding signal transduction pathway. To identify UV-responsive Tdc promoter elements, a loss-of-function analysis was performed, in which deletion derivatives were fused to the gusA reporter gene and analysed in transgenic tobacco plants. Truncation of the Tdc promoter from -1818 (relative to the start of transcription) to -160 reduced expression levels two-fold without affecting the qualitative UV response. Deletion to -37 further reduced expression levels five-fold, but the delta37 promoter also remained UV-responsive. Subsequently, the -160 to -37 region was further studied by gain-of-function experiments, in which the transcriptional activities of tetramerized subfragments fused to truncated promoters were analysed. Combination of the data identified several functional regions in the -160 to +198 promoter. The - 160 to -99 region acts as the main transcriptional enhancer. UV-responsive elements appeared to be redundant in the -160 Tdc promoter and to reside between -99 and -37 and between -37 and + 198.

Alkaloids↗

PubMed2000

Anaerobiosis-specific interaction of tobacco nuclear factors with cis-regulatory sequences in the maize GapC4 promoter.

The promoter of the maize glyceraldehyde-3-phosphate dehydrogenase 4 gene (GapC4) confers strong, specific and ubiquitous anaerobic reporter gene expression in tobacco. To identify factors required for heterologous anaerobic gene expression, 19 progressive 5' and 3' promoter deletions were linked to a chimeric GapC4 TATA box-beta-glucuronidase (GUS) reporter gene construct and transformed into tobacco. In all transgenic lines aerobic expression values were in the range obtained for negative controls while histochemical GUS assays reveal some weak expression in roots only. Anaerobic induction of about 100-fold to more than 1000-fold above unspecific background is mediated by a region of about 190 bp of the GapC4 promoter. Anaerobic reporter gene induction strongly decreases upon deletion of a 20 bp fragment from -286 to -266 relative to the transcription start point. This fragment harbours putative cis-acting sequences. Electrophoretic mobility shift assays with a 50 bp fragment harbouring these cis sequences reveal a high-mobility complex that is formed with nuclear extracts from aerobic and anaerobic leaf tissue while an additional low-mobility complex is anaerobiosis-specific. The formation of the high-mobility complex requires the sequence GTGGGCCCG. The 50 bp fragment alone confers weak and orientation-dependent anaerobic induction to a GapC4 TATA box-beta-glucuronidase (GUS) reporter gene.

Anaerobiosis↗

PubMed2001

Simultaneous analysis of the bidirectional African cassava mosaic virus promoter activity using two different luciferase genes.

The expression of geminivirus genes is controlled by bidirectional promoters which are located in the large intergenic region of the circular DNA genomes and specifically regulated by virus encoded proteins. In order to study the simultaneous regulation of both orientations of the DNA A and DNA B promoters of African cassava mosaic virus (ACMV), they were cloned between two different luciferase genes with the firefly luciferase gene in complementary-sense and the Renilla luciferase gene in virion-sense orientation. The regulation of the ACMV promoters by proteins encoded by the complete DNA A, as well as by the individually expressed transactivator (TrAP) or replication-associated (Rep) proteins was assessed in tobacco and cassava protoplasts using dual luciferase assays. In addition, the regulation of the DNA A promoter integrated into tobacco genome was also assessed. The results show that TrAP activates virion-sense expression strongly both in cassava and tobacco protoplasts, but not in transgenic tobacco plants. In contrast to this, DNA A encoded proteins activate virion-sense expression both in protoplasts and in transgenic plants. At the same time they reduce the expression of the complementary-sense Rep gene on DNA A but activate the expression of the complementary-sense movement protein (MPB) gene on DNA B. The degree of MBP activation is higher in cassava than in tobacco protoplasts, indicating that the plant host also influences the promoter strength. Transient transformation experiments using linearized DNA indicate that the different regulation of the ACMV DNA A promoter in protoplasts and transgenic plants could be due to different DNA curvature in free plasmids and in genes integrated in plant genomic DNA.

Geminiviridae↗

PubMed1998

Insect resistance of transgenic tobacco expressing an insect chitinase gene.

Chitinase expression in the insect gut normally occurs only during moulting, where the chitin of the peritrophic membrane is presumably degraded. Thus, insects feeding on plants that constitutively express an insect chitinase gene might be adversely affected, owing to an inappropriately timed exposure to chitinase. This hypothesis was tested by introducing a cDNA encoding a tobacco hornworm (Manduca sexta) chitinase (EC 3.2.1.14) into tobacco via Agrobacterium tumefaciens-mediated transformation. A truncated but enzymatically active chitinase was present in plants expressing the gene. Segregating progeny of high-expressing plants were compared for their ability to support growth of tobacco budworm (Heliothis virescens) larvae and for feeding damage. Both parameters were significantly reduced when budworms fed on transgenic tobacco plants expressing high levels of the chitinase gene. In contrast, hornworm larvae showed no significant growth reduction when fed on the chitinase-expressing transgenics. However, both budworm and hornworm larvae, when fed on chitinase-expressing transgenic plants coated with sublethal concentrations of a Bacillus thuringiensis toxin, were significantly stunted relative to larvae fed on toxin-treated non-transgenic controls. Foliar damage was also reduced. Plants expressing an insect chitinase gene may have agronomic potential for insect control.

Animals↗

PubMed2000

Expression of Acidothermus cellulolyticus endoglucanase E1 in transgenic tobacco: biochemical characteristics and physiological effects.

The expression of the Acidothermus cellulolyticus endoglucanase E1 gene in transgenic tobacco (Nicotiana tabacum) was examined in this study, where E1 coding sequence was transcribed under the control of a leaf specific Rubisco small subunit promoter (tomato RbcS-3C). Targeting the E1 protein to the chloroplast was established using a chloroplast transit peptide of Rubisco small subunit protein (tomato RbcS-2A) and confirmed by immunocytochemistry. The E1 produced in transgenic tobacco plants was found to be biologically active, and to accumulate in leaves at levels of up to 1.35% of total soluble protein. Optimum temperature and pH for E1 enzyme activity in leaf extracts were 81 degrees C and 5.25, respectively. E1 activity remained constant on a gram fresh leaf weight basis, but dramatically increased on a total leaf soluble protein basis as leaves aged, or when leaf discs were dehydrated. E1 protein in old leaves, or after 5 h dehydration, was partially degraded although E1 activity remained constant. Transgenic plants exhibited normal growth and developmental characteristics with photosynthetic rates similar to those of untransformed SR1 tobacco plants. Results from these biochemical and physiological analyses suggest that the chloroplast is a suitable cellular compartment for accumulation of the hydrolytic E1 enzyme.

Actinomycetales↗

PubMed2000

The restoration of fertility in male sterile tobacco demonstrates that transgene silencing can be mediated by T-DNA that has no DNA homology to the silenced transgene.

Male sterile tobacco plants expressing a pathogenesis-related (PR) beta-1,3-glucanase gene driven by the Arabidopsis thaliana A3 or A9 tapetum-specific promoter, were partially restored to fertility by retransformation with a range of pA9-driven sense and antisense PR glucanase fragments. The restored plants exhibited improved seed set. PR glucanase protein was undetectable in the anthers of these plants and there was an associated increase in microsporocyte callose, the structural target of the A3 and A9-driven PR glucanase. This phenotype was not solely dependent on interactions between sense and antisense PR glucanase transcripts since a pA9-driven restorer was also capable of down regulating a pA3-GUS construct in the absence of extensive promoter, coding region, or terminator sequence homology. Since the A3 and A9 promoters have similar temporal and spatial expression patterns, it is possible that trans-acting factors common to both promoters become limiting in the PR glucanase double transformants resulting in improved levels of fertility. An alternative hypothesis is that additional sequences present in both the silencing and target T-DNAs can mediate the silencing of adjacent non-homologous transgenes.

Arabidopsis↗

PubMed2001

The poplar bark storage protein gene (Bspa) promoter is responsive to photoperiod and nitrogen in transgenic poplar and active in floral tissues, immature seeds and germinating seeds of transgenic tobacco.

In Populus, seasonal nitrogen storage involves the accumulation of a 32 kDa bark storage protein (BSP) in the inner bark parenchyma and xylem rays. Poplar BSPs are encoded by a multigene family and one member, bspA, has been cloned and sequenced. The regulation of bspA was investigated by transforming either hybrid poplar or tobacco with a chimeric gene consisting of the 2.8 kb bspA promoter fused to the coding region of beta-glucuronidase (uidA). In transformed poplar, the bspA 2.8 kb promoter conferred both short-day (SD) and nitrogen (N) inducibility to GUS and activity was localized to the bark (primary and secondary phloem, and cortex) and xylem rays. Night-break treatments inhibited SD induction of GUS. Deletion of the 1.6 kb distal DNA sequences from the bspA promoter eliminated SD induction of GUS while some N induction was retained. These results indicate that although poplar BSP is encoded by a multigene family, transcriptional activation of bspA per se can account for bsp expression in bark and xylem rays in response to either SD or N treatment. These results also show that the elements responsible for SD or N induction are separable. Because of the long generation intervals associated with trees, the developmental regulation of bspA in flowers, developing seeds, and germinating seeds was investigated by transforming the 2.8 kb bspA-promoter::uidA chimeric gene into tobacco. The bspA promoter was active in developing tobacco floral tissues and in seeds during early stages of embryogenesis, decreased progressively during seed maturation and regained activity upon seed germination. Although seed storage proteins of poplar share some similarities to poplar BSP, the observed developmental expression patterns in tobacco are consistent with a role for bspA in vegetative rather than seed storage protein storage.

Gene Expression Regulation, Developmental↗

PubMed2001

Downstream DNA sequences are required to modulate Pvlea-18 gene expression in response to dehydration.

We have previously shown that mRNA and protein encoded by the Pvlea-18 gene from Phaseolus vulgaris L., a member of a new family of late embryogenesis-abundant (LEA) proteins, accumulate in dark-grown bean seedlings not only in response to water deficit but also during optimal irrigation. In this work, we studied Pvlea-18 gene transcriptional regulation by using transgenic Arabidopsis thaliana plants containing a chimeric gene consisting of the Pvlea-18 promoter region and the 3'-nos terminator fused to the GUS gene-coding region. We demonstrate that the chimeric gene is active during Arabidopsis normal development under well-irrigated conditions, and that it is further induced in response to ABA and dehydration treatments. Replacing the 3'-nos terminator with the Pvlea-18 3' region led to an additional increase in expression during development and in response to dehydration, but not in response to exogenous ABA. These results reveal an enhancer effect of the Pvlea-18 3' region, which showed to be higher specifically under dehydration. The small decrease in Pvlea-18 promoter expression observed when transgenic plants treated with fluridone (an ABA biosynthesis inhibitor) were subjected to dehydration suggests that the Pvlea-18 gene dehydration response is predominantly ABA-independent. Finally, we present evidence indicating that Pvlea-18 gene expression is negatively regulated during etiolated growth, particularly in roots, in contrast to the expression pattern observed during normal development.

Abscisic Acid↗

PubMed2001

Inducible expression of bacterio-opsin in transgenic tobacco and tomato plants.

The development of new strategies to enhance resistance of plants to pathogens is instrumental in preventing agricultural losses. Lesion mimic, the spontaneous formation of lesions resembling hypersensitive response lesions in the absence of a pathogen, is a dramatic phenotype occasionally induced upon expression of certain transgenes in plants. These transgenes simulate the presence of a pathogen and, therefore, activate the plant anti-pathogen defense mechanisms and induce a state of systemic resistance. Lesion mimic genes have been successfully used to enhance the resistance of a number of different plants to pathogen attack. However, constitutive expression of these genes in plants is associated with the spontaneous formation of lesions on leaves and stems, reduced growth, and lower yield. We tested the possibility of using a wound-inducible promoter to control the expression of bacterio-opsin (bO), a transgene that confers a lesion mimic phenotype in tobacco and tomato plants when constitutively expressed. We found that plants with inducible expression of bO did not develop spontaneous lesions. Nevertheless. under controlled laboratory conditions, they were found to be resistant to infection by pathogens. The activation of defense mechanisms by the bO gene was not constitutive, and occurred in response to wounding or pathogen infection. Furthermore, wounding of transgenic tobacco plants resulted in the induction of systemic resistance to pathogen attack within 48 h. Our findings provide a promising initial assessment for the use of wound-inducible promoters as a new strategy to enhance pathogen resistance in transgenic crops by means of lesion mimic genes.

Bacteriorhodopsins↗