NN Plants Research Articles

Omics Profiles of Non-GM Tubers from Transgrafted Potato with a GM Scion.

"Transgrafting" is a grafting procedure whereby a transgenic plant body is grafted to a non-transgenic plant body. It is a novel plant breeding technology that allows non-transgenic plants to obtain benefits usually conferred to transgenic plants. Many plants regulate flowering by perceiving the day-length cycle via expression of FLOWERING LOCUS T (FT) in the leaves. The resulting FT protein is translocated to the shoot apical meristem via the phloem. In potato plants, FT is involved in the promotion of tuber formation. Here we investigated the effects of a genetically modified (GM) scion on the edible parts of the non-GM rootstock by using potato plants transformed with StSP6A, a novel potato homolog of the FT gene. Scions prepared from GM or control (wild-type) potato plants were grafted to non-GM potato rootstocks; these were designated as TN and NN plants, respectively. After tuber harvest, we observed no significant differences in potato yield between TN and NN plants. Transcriptomic analysis revealed that only one gene-with unknown function-was differentially expressed between TN and NN plants. Subsequent proteomic analysis indicated that several members of protease inhibitor families, known as anti-nutritional factors in potato, were slightly more abundant in TN plants. Metabolomic analysis revealed a slight increase in metabolite abundance in NN plants, but we observed no difference in the accumulation of steroid glycoalkaloids, toxic metabolites found in potato. Finally, we found that TN and NN plants did not differ in nutrient composition. Taken together, these results indicate that FT expression in scions had a limited effect on the metabolism of non-transgenic potato tubers.

Proteomic Analysis of Alfalfa (Medicago sativa L.) Roots in Response to Rhizobium Nodulation and Salt Stress.

(1) Background: Alfalfa is an important legume forage throughout the world. Although alfalfa is considered moderately tolerant to salinity, its production and nitrogen-fixing activity are greatly limited by salt stress. (2) Methods: We examined the physiological changes and proteomic profiles of alfalfa with active nodules (NA) and without nodules (NN) under NaCl treatment. (3) Results: Our data suggested that NA roots showed upregulation of the pathways of abiotic and biotic stress responses (e.g., heat shock proteins and pathogenesis-related proteins), antioxidant enzyme synthesis, protein synthesis and degradation, cell wall degradation and modification, acid phosphatases, and porin transport when compared with NN plants under salt stress conditions. NA roots also upregulated the processes or proteins of lipid metabolism, heat shock proteins, protein degradation and folding, and cell cytoskeleton, downregulated the DNA and protein synthesis process, and vacuolar H+-ATPase proteins under salt stress. Besides, NA roots displayed a net H+ influx and low level of K+ efflux under salt stress, which may enhance the salt tolerance of NA plants. (4) Conclusions: The rhizobium symbiosis conferred the host plant salt tolerance by regulating a series of physiological processes to enhance stress response, improve antioxidant ability and energy use efficiency, and maintain ion homeostasis.

Differential effects of the recombinant type 1 ribosome-inactivating protein, OsRIP1, on growth of PSB-D and BY-2 cells.

Plant suspension cells were treated with recombinant OsRIP1, a type 1 ribosome-inactivating protein (RIP) from rice (Oryza sativa L.). OsRIP1 triggered cell death in tobacco BY-2 cells but not in Arabidopsis PSB-D cells. Phenotypic changes in BY-2 cells exposed to OsRIP1, included loss of growth capacity, loss of integrity of the plasma membrane and vacuolar collapse. These effects were also accompanied by RNA degradation and DNA fragmentation. Targeting of exogenous OsRIP1 to plant vacuoles and OsRIP1-induced accumulation of transcripts for vacuolar processing enzymes (VPEs) indicated that OsRIP1 provoked plant cell death in tobacco BY-2 cells through the activation of VPEs and subsequent vacuolar disruption, which was probably independent of its N-glycosylase activity on cytosolic ribosomes. Necrosis with limited production of H2O2 was observed after infiltration of high concentrations of OsRIP1 in epidermal cells of Nicotiana tabacum cv. Samsun NN plants. Our study provides the first evidence that OsRIP1 exerts differential effects on the growth of PSB-D and BY-2 cells. The vacuole-dependent cell death pathway is associated with the lethal effect of the exogenously applied OsRIP1 on BY-2 cells.

Rhizobium Symbiosis Leads to Increased Drought Tolerance in Chinese Milk Vetch (Astragalus sinicus L.)

Chinese milk vetch (Astragalus sinicus L.) is an important leguminous green manure that frequently suffers from seasonal drought. To improve its drought tolerance, the effects of rhizobia inoculation on drought tolerance and the underlying physiological mechanism were investigated. Drought tolerance in combination with nitrogen assimilation, free amino acids, and polyamines was investigated in milk vetch with active nodules (AN), with inactive nodules (LN), or without nodules (NN). AN plants had increased drought tolerance compared to LN and NN plants. Glutamine synthetase (GS), glutamine 2-oxoglutarate amino transferase (GOGAT), and glutamate dehydrogenase (GDH) activities were decreased after drought, with higher levels in AN plants than in LN and NN plants under both control and drought stress conditions. Higher levels of proline (Pro), arginine (Ala), alanine (Ala), and glutamate (Glu) were observed in AN plants compared with LN and NN plants. Putrescine (Put), spermidine (Spd), and spermine (Spm) levels were increased in response to drought, and higher levels of Put and Spd were maintained in AN plants. It is suggested that active nodulation leads to increased drought tolerance in milk vetch, which is associated with improved nitrogen fixation and ammonium assimilation, which in turn lead to the promotion of free amino-acid and polyamine synthesis.

Nutrient Deficiency Affects the Growth and Nitrate Concentration of Hydroponic Radish

In hydroponics, a continuous supply of mineral nutrients is essential for plant growth. However, constitutive nutrient-rich conditions also increase the nitrate content in the plants, which can be harmful to human health. Here, we investigated the effect of nutrient deficiency on the growth and component composition of hydroponic radish by changing the timing of nutrient removal from the hydroponic solution. Radish plants that were 14 days old were transferred to four different nutrient conditions for 14 days: nutrient deficiency for 14 days (WW), full nutrient for 14 days (NN), nutrient deficiency for the last 7 days (NW), and nutrient deficiency for the first 7 days (WN). After the treatments, the NW plants had similar taproot growth to NN plants. In contrast, the WN plants significantly reduced taproot growth. The WW plants reduced the shoot and taproot weight and their water contents. The nitrate content in the taproots was reduced in the NW and WW plants. The WW plants contained lower total phenol and higher ascorbic acid and sugar contents. These results suggest that the uptake of nutrient minerals at the young growth stage is important for the growth of radish taproot. Nutrient deficiency management can be one of the most effective tools for regulating radish growth and composition.

Molecular characterization and pathogenicity of an infectious cDNA clone of tomato brown rugose fruit virus

Tomato brown rugose fruit virus (ToBRFV) is a new member of the genus Tobamovirus, and has the potential to affect the production and marketability of tomatoes and peppers. In this study, we sequenced and analyzed the complete genome of ToBRFV isolates from tomato plants showing mosaic and mottling symptoms in Yunnan Province of China. We constructed a full-length infectious cDNA clone of ToBRFV, which could induce systemic infection with typical symptoms in tomato, Nicotiana benthamiana, and N. tabacum cv. Samsun nn plants through Agrobacterium-mediated inoculation. Further experimental evidence demonstrated that the rod-shaped virions accumulating in agroinfiltrated plants are sap-transmissible. This is the first report on the construction of a biologically active, full-length infectious cDNA clone of ToBRFV. The system developed herein will facilitate further research on functions of ToBRFV-encoded proteins and plant-ToBRFV interactions through reverse genetic approaches.

Plant-derived SAC domain of PAR-4 (Prostate Apoptosis Response 4) exhibits growth inhibitory effects in prostate cancer cells.

The gene Par-4 (Prostate Apoptosis Response 4) was originally identified in prostate cancer cells undergoing apoptosis and its product Par-4 showed cancer specific pro-apoptotic activity. Particularly, the SAC domain of Par-4 (SAC-Par-4) selectively kills cancer cells leaving normal cells unaffected. The therapeutic significance of bioactive SAC-Par-4 is enormous in cancer biology; however, its large scale production is still a matter of concern. Here we report the production of SAC-Par-4-GFP fusion protein coupled to translational enhancer sequence (5′ AMV) and apoplast signal peptide (aTP) in transgenic Nicotiana tabacum cv. Samsun NN plants under the control of a unique recombinant promoter M24. Transgene integration was confirmed by genomic DNA PCR, Southern and Northern blotting, Real-time PCR, and Nuclear run-on assays. Results of Western blot analysis and ELISA confirmed expression of recombinant SAC-Par-4-GFP protein and it was as high as 0.15% of total soluble protein. In addition, we found that targeting of plant recombinant SAC-Par-4-GFP to the apoplast and endoplasmic reticulum (ER) was essential for the stability of plant recombinant protein in comparison to the bacterial derived SAC-Par-4. Deglycosylation analysis demonstrated that ER-targeted SAC-Par-4-GFP-SEKDEL undergoes O-linked glycosylation unlike apoplast-targeted SAC-Par-4-GFP. Furthermore, various in vitro studies like mammalian cells proliferation assay (MTT), apoptosis induction assays, and NF-κB suppression suggested the cytotoxic and apoptotic properties of plant-derived SAC-Par-4-GFP against multiple prostate cancer cell lines. Additionally, pre-treatment of MAT-LyLu prostate cancer cells with purified SAC-Par-4-GFP significantly delayed the onset of tumor in a syngeneic rat prostate cancer model. Taken altogether, we proclaim that plant made SAC-Par-4 may become a useful alternate therapy for effectively alleviating cancer in the new era.

Control of tobacco mosaic virus by Pseudomonas fluorescens CZ powder in greenhouses and the field

Pseudomonas fluorescens strain CZ has been reported to inhibit tobacco mosaic virus (TMV) by producing an antibiotic protein. In this study the effects of CZ powder on TMV infection were investigated on tobacco grown in greenhouses and in the field. CZ stocks were cultured in a mixture medium (5 g wheat bran, 1 g soybean flour, 1 g corn flour, 1 g peanut flour), and then mixed with silica white at a ratio of 6 ml:1 g to make CZ powder. The effective cell concentration in a 100-fold dilution of CZ powder was 49.8 × 1010 cfu ml−1. In greenhouse experiments where Nicotiana tabacum cv. Samsun NN plants were challenged by mechanical inoculation with a mixture of the 100-fold diluted CZ powder and TMV, the in vitro suppression of the TMV infection was 88.3% compared to the controls in which a mixture of water and TMV was used as inoculum. Similarly when N. tabacum cv. NC89 plants were inoculated by cutting the leaves with virus-contaminated scissors, dipping the scissors in a 100-fold dilution of CZ powder before cutting showed a disinfection effect of 96.3% compared with water dipping controls. Drenching and spraying a 100-fold dilution of CZ promoted growth and inhibited virus infection by 59.2% on N. tabacum cv. NC89 plants in the greenhouse. In field trials, 100-fold dilution of CZ suppressed TMV infection by 58.2% and 47.6% in 2010 and 2011, respectively, which was similar to the effect of Ningnanmycin (antibiotic purified from Streptomyces noursei var. xichangensis), a registered antiviral agent in tobacco. All these results indicated that CZ has a potential to be used as a hand tool disinfector and an antiviral agent against TMV.

Inducible expression of p50 from TMV for increased resistance to bacterial crown gall disease in tobacco

The dominant tobacco mosaic virus (TMV) resistance gene N induces a hypersensitive response upon TMV infection and protects tobacco against systemic spread of the virus. It has been proposed to change disease resistance specificity by reprogramming the expression of resistance genes or their corresponding avirulence genes. To reprogramme the resistance response of N towards bacterial pathogens, the helicase domain (p50) of the TMV replicase, the avirulence gene of N, was linked to synthetic promoters 4D and 2S2D harbouring elicitor-responsive cis-elements. These promoter::p50 constructs induce local necrotic lesions on NN tobacco plants in an Agrobacterium tumefaciens infiltration assay. A tobacco genotype void of N (nn) was transformed with the promoter::p50 constructs and subsequently crossed to NN plants. Nn F1 offspring selected for the T-DNA develop normally under sterile conditions. After transfer to soil, some of the F1 plants expressing the 2S2D::p50 constructs develop spontaneous necrosis. Transgenic Nn F1 plants with 4D::p50 and 2S2D::p50 expressing constructs upregulate p50 transcription and induce local necrotic lesions in an A. tumefaciens infiltration assay. When leaves and stems of Nn F1 offspring harbouring promoter::p50 constructs are infected with oncogenic A. tumefaciens C58, transgenic lines harbouring the 2S2D::p50 construct induce necrosis and completely lack tumor development. These results demonstrate a successful reprogramming of the viral N gene response against bacterial crown gall disease and highlight the importance of achieving tight regulation of avirulence gene expression and the control of necrosis in the presence of the corresponding resistance gene.

Long‐term protection against tobacco mosaic virus induced by the marine alga oligo‐sulphated‐galactan Poly‐Ga in tobacco plants

In order to study the antiviral effect of the oligo-sulphated galactan Poly-Ga, the leaves of tobacco plants Xhanti(NN) were sprayed with water (control), with increasing concentrations of Poly-Ga, for increasing numbers of treatments or cultivated for increasing times after treatment. Control and treated plants were infected with tobacco mosaic virus (TMV) and the numbers of necrotic lesions were measured in infected leaves. The number of necrotic lesions decreased with increasing concentrations of Poly-Ga, with increasing numbers of treatments and with increasing time after treatment, indicating a long-term protection against TMV that mimicks vaccination. In addition, control Xhanti(nn) plants and plants treated with Poly-Ga and cultivated for increasing times after treatment were infected with TMV in the middle part of the plant, and the levels of TMV-capsid protein (CP) transcripts were measured in apical leaves. TMV-CP transcripts decreased in distant leaves, indicating that Poly-Ga induces systemic protection against TMV. The activities of the defence enzymes phenylalanine ammonia lyase (PAL) and lipoxygenase (LOX) and the amounts of several phenylpropanoid compounds (PPCs) were measured in control and treated plants without infection. A progressive increase in PAL activity was observed with increasing time after treatment, together with the accumulation of free and conjugated PPCs. In contrast, LOX activity remained unchanged. Interestingly, the increase in PAL activity showed a linear correlation with the decrease in necrotic lesions and the decrease in TMV-CP transcript level. Thus, Poly-Ga induced systemic and long-term protection against TMV in tobacco plants that is determined, at least in part, by a sustained activation of PAL and the accumulation of PPCs with potential antiviral activity.

English

Tobacco cultivars Nicotiana tabacum cv. Samsun NN plants carrying the N gene contain a multitude of N-related genes. We cloned a few N homologues andisolated two full-length cDNAs of NL-C26 and NL-B69 genes from N. tabacum cv. Samsun NN. Nucleotide sequence analysis showed that the coding regions of NL-C26 (3,498 bp) and NL-B69 (3,510 bp) had 86 and 83% nucleotide identities with the N gene, respectively. Amino acid sequence analysis revealed that NL-C26 and NL-B69 had the Toll-interleukin-1 receptor/nucleotide-binding site/leucine-rich repeat (TIR/NBS/LRR) structure with 78 and 73% identities to N, respectively.These result indicated that NL-C26 was more similar to N than NL-B69. Tobacco mosaic virus (TMV) infection experiments suggest that NL-C26 and NL-B69 could interact with distinct avirulence (Avr) proteins of yet unidentified pathogens and function as potential HR-inducing resistance proteins similar to N.   Key words: N gene, tobacco mosaic virus (TMV), N homologue, Nicotiana tabacum, hypersensitive response.  

Host Plant Resistance to Pathogen with Reference to Induction of Systemic Acquired Resistance (SAR) in Tobacco (Nicotiana Tabacum Var. Samsun) When Infected with Tobacco Mosaic Virus (TMV)

A model study based on induction of SAR via infecting different lines of tobacco (Nicotiana tabacum var. Samsun) with Tobacco mosaic virus (TMV) was carried out at the Department of Applied Biology, University of Helsinki, Finland in December, 2006. The objective was to demonstrate the onset of defence response, spread of SAR in tobacco plants infected with TMV and detection of expression of marker gene, PR protein gene (PR1) using northern blotting and real time polymerease chain reaction (PCR). Localized necrosis was observed at 3 and 10 days post inoculation (dpi) only in TMV-inoculated NN plants in contrast to absence of similar localized symptom in the corresponding inoculated nn plants except systemic yellowing at 10 dpi. The data generated by northern blot analysis as well as real-time PCR showed existence of variation among experimental treatments with respect to PR1 gene expression. Northern blotting and real-time PCR gave similar results except a difference that was observed for sample 10 probably signifying high precision and sensitivity of real-time PCR in indicating PR1 expression compared to northern blotting. The RNA ladder was not visible in agarose gel and it was not possible to observe the actual result. This confirmed the importance of proceeding to the application of northern blotting. The result of this study indicated localized induction of necrotic lesions in infected tobacco var. Samsun (NN) as indicator for the HR programmed cell death and PRl expression as indicator for SAR in systemic leaves. All plants of nn variety are susceptible and showed only systemic mosaic symptoms in contrast to localized lesions expressed in NN plants. The result of this study indicated that the NN gene in TMV-infected tobacco var. Samsun carrying N allele recognized the corresponding Avirulence gene of the virus inducing the SAR in systemic leaves as indicated via expression of PR1 protein and hypersensitive reaction (HR) in infected leaves as visualized by localized necrosis in infected leaves. This is in agreement with the theory of gene-for-gene interaction in plant defence responses. Keywords : Gene-for-Gene; Inoculation; Pathogenesis-Related proteins East African Journal of Sciences Vol. 2 (2) 2008: pp. 184-188

Transglutaminase activity changes during the hypersensitive reaction, a typical defense response of tobacco NN plants to TMV

The occurrence of glutamyl polyamines (PAs) and changes in activity and levels of transglutaminase (TGase, EC 2.3.2.13), the enzyme responsible for their synthesis, are reported during the progression of the hypersensitive reaction (HR) of resistant NN tobacco plants (Nicotiana tabacum L. cv. Samsun) to tobacco mosaic virus (TMV). Mature leaves of tobacco were collected over 0-72 h after inoculation with TMV or phosphate buffer (mock). In vivo synthesis of polyamine glutamyl derivatives (glutamyl PAs), catalyzed by TGase activity, was evaluated after supplying labeled putrescine (Pu, a physiological substrate of TGase) to leaves. Results show that, starting from 24 h, mono-(gamma-glutamyl)-Pu and bis-(gamma-glutamyl)-Sd were recovered in TMV-inoculated samples but not in mock-inoculated ones; 2 days later, in the former, the amount of glutamyl derivatives further increased. An in vitro radiometric assay showed that, in TMV-inoculated leaves, TGase activity increased from 24 h onwards relative to mock controls. An immunoblot analysis with AtPng1p polyclonal antibody detected a 72-kDa protein whose amount increased at 72 h in TMV-inoculated leaves and in the lesion-enriched areas. A biotin-labeled cadaverine incorporation assay showed that TGase activity occurred in S1 (containing soluble proteins), S2 (proteins released by both cell walls and membranes) and S3 (membrane intrinsic proteins) fractions. In S3 fraction, where changes were the most relevant, TGase activity was enhanced in both mock-inoculated and TMV-inoculated samples, but the stimulation persisted only in the latter case. These data are discussed in the light of a possible role of TGase activity and glutamyl PAs in the defense against a viral plant pathogen.

Development of a Screening System for Plant Defense-Inducing Agent using Transgenic Tobacco Plant with PR-1a Promoter and GUS Gene

Pathogenesis-related protein-1a (PR-1a) is strongly induced in tobacco plants by pathogen attack, exogenous salicylic acid (SA) application and by other developmental processes. In order to develop a rapid screening system for the selection of plant defense-inducing compounds originated from various sources, we have transformed tobacco Samsun NN plants with a chimeric construct consisting of GUS <TEX>$(\beta-glucuronidase)$</TEX>. In the <TEX>$T_1$</TEX> generation, three transgenic lines having stable GUS expression were selected for further promoter analysis. Using GUS histochemical assay, we observed strong GUS induction driven by PR-1a promoter in PR1a-GUS transgenic tobacco leaves in response to the exogenous application of SA or benzol (1,2,3) thiadiazole-7-carbothioic acid S-methyl ester (BTH), a SA­derivative compound. In addition, GUS expression was maintained locally or systemically in PR1a-GUS transgenic line <TEX>$\#5\;T_2$</TEX> generation) until after 3 days when they were treated with same chemicals. Our results suggested that the PR1a-GUS reporter gene system in tobacco plants may be applicable for the large-scale screening of defense-inducing substances.

Molecular Chaperone Hsp90 Associates with Resistance Protein N and Its Signaling Proteins SGT1 and Rar1 to Modulate an Innate Immune Response in Plants

SGT1 and Rar1 are important signaling components of resistance (R) gene-mediated plant innate immune responses. Here we report that SGT1 and Rar1 associate with the molecular chaperone Hsp90. In addition, we show that Hsp90 associates with the resistance protein N that confers resistance to tobacco mosaic virus. This suggests that Hsp90-SGT1-Rar1 and R proteins might exist in one complex. Suppression of Hsp90 in Nicotiana benthamiana plants shows that it plays an important role in plant growth and development. In addition, Hsp90 suppression in NN plants compromises N-mediated resistance to tobacco mosaic virus. Our results reveal a new role for SGT1- and Rar1-associated chaperone machinery in R gene-mediated defense signaling.

Identification and characterization of a novel tobacco mosaic virus resistance N gene homologue in Nicotiana tabacum plants.

Nicotiana tabacum cv. Xanthi nn plants are susceptible to infection by most tobamoviruses (TMV). However, such plants display a partial hypersensitive resistance response (HR-like response) to TMV-Cg. The genetic mechanism of the HR-like response has yet not been determined, but it may involve a gene with a function similar to that of a resistance gene, responsible for HR in resistant plants. We have cloned a gene homologous to the resistance N gene, named NH, from Nicotiana Xanthi nn plants. The coding region of NH is 5.028 base pairs (bp) long and has 82.6% nucleotide identity with the N gene. In contrast to the N gene, the NH gene lacks intron 4 and does not have sites for alternative splicing of intron 3. Analysis of its sequence revealed that NH belongs to the TIR / NSB / LRR gene class. We were able to detect stable levels of NH-transcript in Nicotiana Xanthi nn plants from 0 to 18 h post-inoculation (hpi) with TMV-Cg. Transcript levels increased slightly at 24 hpi and dropped below basal values at 48 hpi. The NH transcript was also detected in a range of resistant Nicotiana plants (N. tabacum Xanthi NN, N. glutinosa, N. glauca and N. rustica) suggesting that NH is a homologue of the N gene, rather than an allele. We have cloned and characterised the NH gene (GenBank acc. no. bankit598573 AY535010) from nn susceptible plants and postulate that this gene might be involved in the HR-like response seen in these plants.

Expression in plants and immunogenicity of plant virus-based experimental rabies vaccine

A new approach to the production and delivery of vaccine antigens is the use of engineered amino virus-based vectors. A chimeric peptide containing antigenic determinants from rabies virus glycoprotein (G protein) (amino acids 253–275) and nucleoprotein (N protein) (amino acids 404–418) was PCR-amplified and cloned as a translational fusion product with the alfalfa mosaic virus (AlMV) coat protein (CP). This recombinant CP was expressed in two plant virus-based expression systems. The first one utilized transgenic Nicotiana tabacum cv. Samsun NN plants providing replicative functions in trans for full-length infectious RNA3 of AlMV (NF1-g24). The second one utilized Nicotiana benthamiana and spinach ( Spinacia oleracea) plants using autonomously replicating tobacco mosaic virus (TMV) lacking native CP (Av/A4-g24). Recombinant virus containing the chimeric rabies virus epitope was isolated from infected transgenic N. tabacum cv. Samsun NN plants and used for parenteral immunization of mice. Mice immunized with recombinant virus were protected against challenge infection. Based on the previously demonstrated efficacy of this plant virus-based experimental rabies vaccine when orally administered to mice in virus-infected unprocessed raw spinach leaves, we assessed its efficacy in human volunteers. Three of five volunteers who had previously been immunized against rabies virus with a conventional vaccine specifically responded against the peptide antigen after ingesting spinach leaves infected with the recombinant virus. When rabies virus non-immune individuals were fed the same material, 5/9 demonstrated significant antibody responses to either rabies virus or AlMV. Following a single dose of conventional rabies virus vaccine, three of these individuals showed detectable levels of rabies virus-neutralizing antibodies, whereas none of five controls revealed these antibodies. These findings provide clear indication of the potential of the plant virus-based expression systems as supplementary oral booster for rabies vaccinations.

Effects of replacing the movement protein gene of Tobacco mosaic virus by that of Tomato mosaic virus

The broad bean strain of Tobacco mosaic virus (TMV-B) infects Nicotiana tabacum White Burley systemically whereas the tomato strain of T. mosaic virus (ToMV-S1) induces necrotic local lesions and is restricted to inoculated leaves. To examine the possible role of the viral movement protein (MP) in these symptom differences, a chimaeric virus (T/OMP) was produced in which the TMV-B MP gene was replaced by the ToMV-S1 MP gene. T/OMP induced the same symptoms as TMV-B in N. tabacum White Burley . However, in N. tabacum Samsun NN and other plants containing the N resistance gene, T/OMP caused necrotic lesions that were smaller than those produced by TMV-B but similar in size to those of ToMV-S1. We conclude that ToMV MP gene can substitute functionally for the TMV-B MP gene, and that the MP gene influences the size of necrotic local lesions on N-containing hosts.

Effects of supplied nitrogen form on growth and water uptake of French bean ( Phaseolus vulgaris L.) plants

In order to investigate the effect of N form on dry matter (DM) formation and water uptake rate, French bean (Phaseolus vulgaris L. `Sotaxa') plants were grown with a split-root system. Three treatments were compared: sole nitrate (NO− 3) supply (NN), sole ammonium (NH+ 4) supply (AA) and spatially separated supply of NO− 3 and NH+ 4 (NA). The pH of the nutrient solutions was kept constant at 6.3 using a pH-stat system. 9 days after onset of the treatments, NN plants had higher root (36%) and shoot dry matter (11%) than AA plants. N form drastically influenced partitioning of assimilates: in the NA treatment, the root half exposed to NO− 3 revealed a 170% higher DM than the root half exposed to NH+ 4. N form affected stable carbon-isotope discrimination (δ) of leaf tissue. In leaves of plants which were supplied with NH+ 4 (AA; NA) δ was significantly more negative (–29.4‰, –29.6‰) than in NN treatment (–28.2‰). We explain this effect by differences in stomatal conductance. We suppose that the significantly less negative δ of root tissue under NH+ 4 supply is most probably related to higher PEP-case activity. The water uptake rate was higher in NN than in AA grown plants. This effect was found in both, short- and long-term experiments. In case of NA plants, the water uptake in the root part being exposed to NO− 3 was 104% higher than in those receiving NH+ 4. At least in the case of the NA treatment we can exclude shoot growth effects as being responsible for differences in water uptake. We therefore assume that differences in root hydraulic conductivity are responsible for the observed effects.

Polyamine metabolism is upregulated in response to tobacco mosaic virus in hypersensitive, but not in susceptible, tobacco.

• Change is reported in the biosynthetic and oxidative activity of hypersensitive (NN) and susceptible (nn) tobacco (Nicotiana tabacum) plants in response to tobacco mosaic virus (TMV). • Mature leaves of nn and NN tobacco were collected over 0-72 h as uninoculated controls or after inoculation with TMV or phosphate buffer (mock-inoculation). The polyamine response to inoculation was analysed by measuring activity and gene expression of S-adenosylmethionine decarboxylase (SAMDC), arginine-(ADC) and ornithine decarboxylases (ODC); incorporation of labelled putrescine; and activity of diamine oxidase (DAO). • In NN leaves SAMDC activity and transcript levels, and DAO activity increased in the TMV-inoculated plants but not in the other treatments; a two-fold increase in DAO activity was seen after 72 h. Both ADC and ODC activity increased in NN leaves at 72 h in TMV-inoculated plants; ADC mRNA increased with activity. The increase in SAMDC mRNA (24 h) preceded the rise in activity (72 h). [3 H]putrescine added to NN leaves led to enhanced label recovery and incorporation into spermidine and spermine in TMV-inoculated plants. No significant changes in biosynthetic or oxidative activity occurred in nn plants. • After TMV inoculation, NN, unlike nn, tobacco plants upgrade polyamine synthesis and oxidation; this leads to changes in cellular components which might induce programmed cell death.