The effect of water stress on the vacuole-extravacuole compartmentation of proline in potato cell suspension cultures

Solute compartmentation in cells is an important component of metabolic regulation. There is only little information on how stress treatment of cells effects this component. Therefore, the effect of water stress [10% (w/v) PEG 6000] on the vacuolar-extravacuolar proline compartmentation was studied in a cell suspension culture of Svlanum tuberosum L, cv, HH258, In non-stressed cells 34% of the total cellular proline was located in the vacuole. After 20 h of water stress the proline pool of the cells was increased 4-6 fold and only t6% of it was found in the vacuole. A negative correlation between the total cellular proline content and its percentage in the vacuole was observed, irrespective of the culture method (stress or non-stress culture). The stress-induced changes in proline compartmentation are discussed.     

Genotype differences in the metabolism of proline and polyamines under moderate drought in tomato plants

Water stress is one of the most important factors limiting the growth and productivity of crops. The implication of compatible osmolytes such as proline and polyamines in osmotic adjustment has been widely described in numerous plants species under stress conditions. In the present study, we investigated the response of five cherry tomato cultivars (Solanum lycopersicum L.) subjected to moderate water stress in order to shed light on the involvement of proline and polyamine metabolism in the mechanisms of tolerance to moderate water stress. Our results indicate that the most water stress‐resistant cultivar (Zarina) had increased degradation of proline associated with increased polyamine synthesis, with a higher concentration of spermidine and spermine under stress conditions. In contrast, Josefina, the cultivar most sensitive to water stress, showed a proline accumulation associated with increased synthesis after being subjected to stress. In turn, in this cultivar, no rise in polyamine synthesis was detected. Therefore, all the data appear to indicate that polyamine metabolism is more involved in the tolerance response to moderate water stress.     

Biochemical and molecular determinance of resistance and susceptibility in Solanum tuberosum (potato) plants challenged with Phytopthora infestans

Determinance of resistance was studied in four different varieties of Solanum tuberosum using biochemical and molecular parameters. It was clearly evident that due to infection of P. infestans, the total protein, total phenol and lignin were induced in all four verities; however, the induction was found more in resistant varieties compared to susceptible varieties. Induction of proteins was also determined by SDS-PAGE analysis. Deposition of lignin was showed by histological comparison using Phloroglucinol HCl staining. Higher deposition of lignin in resistant varieties could be considered as reliable characters related to disease resistance and could be used as biochemical markers for late blight resistance. The RAPD profile generated using eight different decameric primers showed both, polymorphic as well as monomorphic bands. There were many unique bands found only in resistant varieties. These polymorphic bands could also be served as molecular markers for screening of LB resistance potato varieties.     

Responses of potato genotypes to drought. II. Leaf area index, growth and yield

Canopy expansion, growth and yield were examined in 19 genotypes of potato (Solanurn tuberosum L.) grown either with irrigation or droughted from the time of plant emergence. In the irrigated treatment, genotypes differed both in the maximum leaf area index (L_ai) achieved and in the duration the canopy was maintained. Drought reduced both the rate of canopy expansion and the maximum L_ai achieved. In the droughted treatment, biomass production was correlated with leaf area duration. The relation between the effect of drought on the growth of individual leaves and biomass or yield is examined. It is concluded that selection for early leaf appearance combined with the ability to sustain leaf growth with increasing soil moisture deficit (SMD) would improve productivity in the presence of drought.     

Susceptibility of eight potato cultivars to tuber infection by Phytophthora erythroseptica and Pythium ultimum and its relationship to mefenoxam-mediated control of pink rot and leak

In addition to cultural practices, the application of the fungicide mefenoxam is an important disease management tactic used to control both pink rot and leak on potato tubers grown in the USA. Mefenoxam resistance has been identified in many of the potato growing regions, and therefore resistance management strategies are very important for retaining this fungicide as a tool to manage these storage rot diseases. The relationship between mefenoxam efficacy and cultivar susceptibility to pink rot and leak was assessed in post‐harvest inoculation studies. Mefenoxam was applied to potato (Solanum tuberosum) cultivars known to express varying levels of susceptibility to pink rot and leak caused by Phytophthora erythroseptica and Pythium ultimum, respectfully. Tubers harvested from plants treated with in‐furrow and foliar applications of mefenoxam were inoculated with isolates sensitive to the fungicide. Incidence and severity of both diseases ranged widely among cultivars. Russet Norkotah was the most susceptible to infection by P. erythroseptica, while cvs Pike and Atlantic were the most resistant. Cultivars Dark Red Norland, Russet Norkotah, Goldrush and Russet Burbank were most susceptible to infection by P. ultimum whereas Snowden was most resistant. Control of pink rot differed significantly among cultivars following mefenoxam treatment, ranging from 28% (cv. Goldrush) to 67% (cv. Snowden) and generally provided the greatest level of disease control on susceptible and moderately susceptible cultivars such as Russet Norkotah and Snowden, respectively. In contrast, the impact of mefenoxam on leak development was minimal and disease control did not differ significantly among the cultivars. The fungicide failed to control leak in the susceptible cvs Atlantic and Pike and control ranged from 1.7% to 5.2% in cvs Goldrush, Russet Norkotah, Dark Red Norland, Russet Burbank and Kennebec. The greatest level of leak control was achieved with the moderately resistant cv., Snowden, at 12.7%. Cultivars most likely to benefit from mefenoxam treatments should be targeted as part of a pink rot management programme. Judicious use of the fungicide, when matched with the level of cultivar susceptibility, may prove to be an efficient and effective approach to reduce infection rates and possibly manage mefenoxam resistance thereby maintaining longevity of the compound.     

In vitro recurrent selection of potato: production and characterization of salt tolerant cell lines and plants

A stable salt-tolerant potato cell line, able to grow on media containing 60–450 mM NaCl (i.e. low to high salinity) was selected. Callus grown on 120 or 150 mM NaCl showed higher fresh weights than the rest of the treatments. Replacing NaCl by KCl or Na2SO4 showed that reductions in fresh weight were mainly due to the presence of Na+ ions. When PEG 6000 was added to the medium instead of salt, the salt tolerant cell lines were unable to overcome the PEG-induced water stress. Whole plants, regenerated from salt tolerant callus, exhibited salt stress tolerance as evidenced by their higher fresh and dry weights when watered with 90 mM NaCl, and they also produced more tubers per plant under salt stress. Salt-tolerant plants differed phenotypically from control plants both in terms of leaf shape, tuber flesh and skin colour, which was reddish. In addition, DNA fingerprinting by RAPDs, with 70 different primers, confirmed that the salt tolerant regenerants also differed genotypically from the control, salt sensitive Kennebec potato plants from which they had been selected. This revised version was published online in June 2006 with corrections to the Cover Date.     

Dry matter production and partitioning in potato plants subjected to combined deficiencies of nitrogen, phosphorus and potassium

Three experiments examined effects on growth, dry matter partitioning and nutrient uptake in potato plants grown in large pots under different combinations of adequate and deficient levels of nitrogen, phosphorus and potassium. N supply affected the growth of all leaves, with low N reducing both the size of individual leaves and the extent of branch growth. P and K availability affected the growth of later formed leaves and only when both were deficient was branch growth substantially reduced. At later stages of growth, total green leaf area was significantly reduced by deficiency of each of the nutrients. Partitioning of dry matter to tubers was markedly reduced by K deficiency and increased in one experiment by P deficiency. When both P and K were deficient, partitioning approximated that under non‐limiting conditions. Leaf weight ratio (LWR) was higher under K deficiency, but not when P was also deficient, and was consistently higher when the ratio of K : P in dry matter was less than approximately five. In these experiments, LWR was not consistently related to shoot N% and N supply had relatively little effect on partitioning. There were large treatment effects on tuber dry matter percentage, characterised by significant interactions especially between N and K. Deficiency of one nutrient increased the concentration of others but uptake was highly regulated as crop content of all three nutrients was reduced when the supply of any one was deficient. The results show that the response of potatoes to single deficiencies may be influenced greatly by the levels of other nutrients.     

Characteristics of oxidative stress in potato plants with modified carbohydrate metabolism

Effects of sugars on the development of hypothermia-induced oxidative stress were studied in leaves of two potato genotypes (Solanum tuberosum L., cv. Désirée): with normal carbohydrate metabolism and a genotype with increased sugar content modified by insertion of yeast-derived invertase gene. It was found that generation of proceeds more actively in transformed plants than in control plants. On the contrary H₂O₂ concentration and the catalese and peroxidase activities were lower. At the same time, the activities of superoxide dismutase were similar in plants of both genotypes. A short-term incubation of plants at ?7°C confirmed that a higher freezing tolerance of transformed plants was due to low-molecular-weight components of antioxidant protection system rather than to enzymatic component. Literature data and experimental results suggest that the protective effect of sugars is caused by their ability to scavenge ROS nonspecifically under stress conditions     

Linkage analysis in tetraploid potato and association of markers with quantitative resistance to late blight ( Phytophthora infestans )

We have constructed a partial linkage map in tetraploid potato which integrates simplex, duplex and double-simplex AFLP markers. The map consists of 231 maternal and 106 paternal markers with total map lengths of 990.9 cM and 484.6 cM. The longer of the two cumulative map lengths represents approximately 25% coverage of the genome. In tetraploids, much of the polymorphism between parental clones is masked by `dosage' which significantly reduces the number of individual markers that can be scored in a population. Consequently, the major advantage of using AFLPs – their high multiplex ratio – is reduced to the point where the use of alternative multi-allelic marker types would be significantly more efficient. The segregation data and map information have been used in a QTL analysis of late blight resistance, and a multi-allelic locus at the proximal end of chromosome VIII has been identified which contributes significantly to the expression of resistance. No late blight resistance genes or QTLs have previously been mapped to this location. Received: 1 October 1997 / Accepted: 18 March 1998     

Molecular changes associated with tuberisation in Solanum tuberosum. Differential expression of α-tubulin isotypes

Changes in gene expression that occur in the stolon tips of potato ( Solanum tuberosum L.) cv. Record during tuberisation were investigated. Protein extracts from stolon tips at various stages in the tuberisation process were analysed by two-dimensional gel electrophoresis. A number of quantitative and qualitative changes in polypeptide composition accompanied the very early stages of tuberisation. In vitro translation of RNA extracted from stolon tips also revealed quantitative and qualitative changes associated with tuberisation. Immunoblotting of protein extracts with monoclonal antibodies raised against α- and β-tubulin showed quantitative changes in the relative level of β-tubulin, but not α-tubulin, as the stolon tips tuberised. Changes in the pattern of α-tubulin isotype expression were shown to occur at early stages in the tuberisation process.     

Patterns of pinitol accumulation in soybean plants and relationships to drought tolerance

Previous studies indicate that methylated cyclitols are potentially important osmolytes in plants. In a search for genetic diversity for pinitol (D -3-O-methyl-chiro-inositol) accumulation in soybean (Glycine max (L.) Merr.), two- to three-fold differences in pinitol accumulation in leaf blades were found among Chinese plant introductions. Furthermore, it was found that genotypes that accumulated high concentrations of pinitol, when grown under well-watered conditions, had been selected for performance in regions of China having low rainfall. Among the carbohydrates analysed, pinitol accumulation was uniquely associated with adaptation to dry areas of China. A detailed study of pinitol accumulation in the soybean plant showed two- to three-fold gradients in pinitol concentration from the bottom to the top of the plant. The gradient shifted during plant development, with consistently higher concentrations of pinitol in the uppermost leaves. Pinitol accumulation was not correlated with activity of the key biosynthetic enzyme, inositol methyl transferase. This result and other lines of evidence indicated that shifting patterns of pinitol accumulation were due to translocation of the cyclitol from lower to upper nodes. Pinitol, proline, and sugars accumulated in leaf blades on soybean plants subjected to drought, but the molar concentration of pinitol in stressed plants was greater than the concentrations of proline or sugars. Although the mechanism by which pinitol participates in drought tolerance is not fully known, our results provide additional correlative evidence linking pinitol and drought tolerance in soybean.     

Trehalose accumulation in Azospirillum brasilense improves drought tolerance and biomass in maize plants

Bacteria of the genus Azospirillum increase the grain yield of several grass crops. In this work the effect of inoculating maize plants with genetically engineered Azospirillum brasilense for trehalose biosynthesis was determined. Transformed bacteria with a plasmid harboring a trehalose biosynthesis gene-fusion from Saccharomyces cerevisiae were able to grow up to 0.5 M NaCl and to accumulate trehalose, whereas wild-type A. brasilense did not tolerate osmotic stress or accumulate significant levels of the disaccharide. Moreover, 85% of maize plants inoculated with transformed A. brasilense survived drought stress, in contrast with only 55% of plants inoculated with the wild-type strain. A 73% increase in biomass of maize plants inoculated with transformed A. brasilense compared with inoculation with the wild-type strain was found. In addition, there was a significant increase of leaf and root length in maize plants inoculated with transformed A. brasilense . Therefore, inoculation of maize plants with A. brasilense containing higher levels of trehalose confers drought tolerance and a significant increase in leaf and root biomass. This work opens the possibility that A. brasilense modified with a chimeric trehalose biosynthetic gene from yeast could increase the biomass, grain yield and stress tolerance in other relevant crops.     

Freezing tolerance, cold acclimation and oxidative stress in potato. Paraquat tolerance is related to acclimation but is a poor indicator of freezing tolerance

Potato is a species commonly cultivated in temperate areas where the growing season may be interrupted by frosts, resulting in loss of yield. Cultivated potato, Solanum tuberosum, is freezing sensitive, but it has several freezing-tolerant wild potato relatives, one of which is S. commersonii. Our study was aimed to resolve the relationship between enhanced freezing tolerance, acclimation capacity and capacity to tolerate active oxygen species. To be able to characterize freezing tolerant ideotypes, a potato population (S1), which segregates in freezing tolerance, acclimation capacity and capacity to tolerate superoxide radicals, was produced by selfing a somatic hybrid between a freezing-tolerant Solanum commersonii (LT₅₀=-4.6°C) and -sensitive S. tuberosum (LT₅₀=-3.0°C). The distribution of non-acclimated freezing tolerance (NA-freezing tolerance) of the S1 population varied between the parental lines and we were able to identify genotypes, having significantly high or low NA-freezing tolerance. When a population of 25 genotypes was tested both for NA-freezing and paraquat (PQ) tolerance, no correlation was found between these two traits (R = 0.02). However, the most NA-freezing tolerant genotypes were also among the most PQ tolerant plants. Simultaneously, one of the NA-freezing sensitive genotypes (2022) (LT₅₀=-3.0°C) was observed to be PQ tolerant. These conflicting results may reflect a significant, but not obligatory, role of superoxide scavenging mechanisms in the NA-freezing tolerance of S. commersonii. The freezing tolerance after cold acclimation (CA-freezing tolerance) and the acclimation capacity (AC) was measured after acclimation for 7 days at 4/2°C. Lack of correlation between NA-freezing tolerance and AC (R =-0.05) in the S1 population points to independent genetic control of NA-freezing tolerance and AC in Solanum sp. Increased freezing tolerance after cold acclimation was clearly related to PQ tolerance of all S1 genotypes, especially those having good acclimation capacity. The rapid loss of improved PQ tolerance under deacclimation conditions confirmed the close relationship between the process of cold acclimation and enhanced PQ tolerance. Here, we report an increased PQ tolerance in cold-acclimated plants compared to non-acclimated controls. However, we concluded that high PQ tolerance is not a good indicator of actual freezing tolerance and should not be used as a selectable marker for the identification of a freezing-tolerant genotype.     

The changes in invertase activity and the content of sugars in the course of adaptation of potato plants to hypothermia

The pattern of changes in the activity of various forms of invertase (acid soluble, alkaline, and acid insoluble) and the content of sugars (glucose, fructose, and sucrose) in the course of plant adaptation to prolonged (6 days) hypothermia (5°C) was investigated in the leaves of potato plants (Solanum tuberosum L., cv. Desiree) produced in vitro. We used the wild-type plants as a control and transformed plants with carbohydrate metabolism modified by inserting the yeast gene for invertase (apoplastic enzyme). In the course of adaptation to hypothermia, the activity of acid invertase was shown to rise and the content of sucrose and glucose to increase in the leaves of both genotypes. The greatest activity of acid invertases by the third day of cold acclimation corresponded to the peak level of sugars; in transformed plants, these characteristics exceeded those in the control plants. The transformed plants were more cold resistant than the control plants as suggested by the lack of disturbance of ion permeability of their membranes. It was concluded that owing to accumulation of low-molecular carbohydrates in the course of cold acclimation caused by activation of acid invertase cold resistant plants better adapt to temperature drop.     

The effect of NaCl on proline accumulation in potato seedlings and calli

The effects of salt stress were studied on the accumulation and metabolism of proline and its correlation with Na⁺ and K⁺ content in shoots and callus tissue of four potato cultivars, viz., Agria, Kennebec (relatively salt tolerant), Diamant and Ajax (relatively salt sensitive). Na⁺ and proline contents increased in all cultivars under salt stress. However, K⁺ and protein contents decreased in response to NaCl treatments. The activities of enzymes involved in proline metabolism, Δ¹-pyrroline-5-carboxylate synthetase (P5CS) and proline dehydrogenase (ProDH) increased and decreased, respectively, in response to elevated NaCl concentrations. The changes of P5CS and ProDH activities in more salt sensitive cultivars (Diamant, Ajax) were more than those in the tolerant ones. Then the stimulation of synthesis in combination with a partially increase of protein proteolysis, a decrease in proline utilization and inhibition of oxidation resulted in high proline contents in seedlings and calli under salt stress. In callus tissue, reduced growth and cell size may be partially responsible for high proline accumulation in response to high NaCl levels. However, although the basic proline contents in the seedlings of more salt tolerant cultivars were higher than the sensitive ones, a clear relationship was not generally observed between accumulation of proline and salt tolerance in potato.