Effects of down-regulating ornithine decarboxylase upon putrescine-associated metabolism and growth in Nicotiana tabacum L.

Transgenic plants of Nicotiana tabacum L. homozygous for an RNAi construct designed to silence ornithine decarboxylase (ODC) had significantly lower concentrations of nicotine and nornicotine, but significantly higher concentrations of anatabine, compared with vector-only controls. Silencing of ODC also led to significantly reduced concentrations of polyamines (putrescine, spermidine and spermine), tyramine and phenolamides (caffeoylputrescine and dicaffeoylspermidine) with concomitant increases in concentrations of amino acids ornithine, arginine, aspartate, glutamate and glutamine. Root transcript levels of S-adenosyl methionine decarboxylase, S-adenosyl methionine synthase and spermidine synthase (polyamine synthesis enzymes) were reduced compared with vector controls, whilst transcript levels of arginine decarboxylase (putrescine synthesis), putrescine methyltransferase (nicotine production) and multi-drug and toxic compound extrusion (alkaloid transport) proteins were elevated. In contrast, expression of two other key proteins required for alkaloid synthesis, quinolinic acid phosphoribosyltransferase (nicotinic acid production) and a PIP-family oxidoreductase (nicotinic acid condensation reactions), were diminished in roots of odc-RNAi plants relative to vector-only controls. Transcriptional and biochemical differences associated with polyamine and alkaloid metabolism were exacerbated in odc-RNAi plants in response to different forms of shoot damage. In general, apex removal had a greater effect than leaf wounding alone, with a combination of these injury treatments producing synergistic responses in some cases. Reduced expression of ODC appeared to have negative effects upon plant growth and vigour with some leaves of odc-RNAi lines being brittle and bleached compared with vector-only controls. Together, results of this study demonstrate that ornithine decarboxylase has important roles in facilitating both primary and secondary metabolism in Nicotiana

Counting the costs:Nitrogen partitioning in <i>Sorghum </i>mutants

Long-standing growth/defence theories state that the production of defence compounds come at a direct cost to primary metabolism when resources are limited. However, such trade-offs are inherently difficult to quantify. We compared the growth and nitrogen partitioning in wild type Sorghum bicolor (L.) Moench, which contains the cyanogenic glucoside dhurrin, with unique mutants that vary in dhurrin production. The totally cyanide deficient 1 (tcd1) mutants do not synthesise dhurrin at all whereas mutants from the adult cyanide deficient class 1 (acdc1) have decreasing concentrations as plants age. Sorghum lines were grown at three different concentrations of nitrogen. Growth, chemical analysis, physiological measurements and expression of key genes in biosynthesis and turnover were determined for leaves, stems and roots at four developmental stages. Nitrogen supply, ontogeny, tissue type and genotype were all important determinants of tissue nitrate and dhurrin concentration and turnover. The higher growth of acdc1 plants strongly supports a growth/defence trade-off. By contrast, tcd1 plants had slower growth early in development, suggesting that dhurrin synthesis and turnover may be beneficial for early seedling growth rather than being a cost. The relatively small trade-off between nitrate and dhurrin suggests these may be independently regulated.</jats:p

Label-free Raman hyperspectral imaging analysis localizes the cyanogenic glucoside dhurrin to the cytoplasm in sorghum cells

Abstract Localisation of metabolites in sorghum coleoptiles using Raman hyperspectral imaging analysis was compared in wild type plants and mutants that lack cyanogenic glucosides. This novel method allows high spatial resolution in situ localization by detecting functional groups associated with cyanogenic glucosides using vibrational spectroscopy. Raman hyperspectral imaging revealed that dhurrin was found mainly surrounding epidermal, cortical and vascular tissue, with the greatest amount in cortical tissue. Numerous “hotspots” demonstrated dhurrin to be located within both cell walls and cytoplasm adpressed towards the plasmamembrane and not in the vacuole as previously reported. The high concentration of dhurrin in the outer cortical and epidermal cell layers is consistent with its role in defence against herbivory. This demonstrates the ability of Raman hyperspectral imaging to locate cyanogenic glucosides in intact tissues, avoiding possible perturbations and imprecision that may accompany methods that rely on bulk tissue extraction methods, such as protoplast isolation

Crop wild relatives as a genetic resource for generating low-cyanide, drought-tolerant Sorghum

Using a Sorghum bicolor cultivar and seven wild Sorghum species endemic to Australia as our experimental system, we monitored their different responses to drought by assessing growth and morphological, physiological and biochemical parameters. Drought stress significantly decreased height, biomass, the maximum potential quantum efficiency of photosystem II, photosynthetic rate and relative water content in S. bicolor, while several of the wild species were much more tolerant. Drought significantly increased dhurrin concentration in aboveground tissue in S. bicolor but not in the wild species. Root dhurrin content was unaffected by drought in S. bicolor, in contrast to the varied responses observed in the wild species. Sorghum macrospermum and S. brachypodum maintained relatively high growth and photosynthetic performance under drought, with negligible aboveground dhurrin content. These wild species are promising candidates for sorghum crop improvement

Sporobolus stapfianus, a model desiccation-tolerant grass

Sporobolus stapfianus Gandoger, one of ~40 known &lsquo;anabiotic&rsquo;grass species (i.e. &lsquo;able to regain vital activity from a state of latent life&rsquo;), is the most versatile tool for research into desiccation tolerance in vegetative grass tissue. Current knowledge on this species is presented, including the features that suit it for investigations into the plant&rsquo;s ability to survive dehydration of its leaf protoplasm. The main contributors to desiccation tolerance in S. stapfianus leaves appear to be: accumulation during dehydration of protectants of membranes and proteins; mechanisms limiting oxidative damage; a retention of protein synthetic activity in late stages of drying that is linked with changes in gene expression and in the proteomic array; and an ability to retain net synthesis of ATP during drying. S. stapfianus exemplifies an advanced stage of an evolutionary trend in desiccation tolerant plants towards increased importance of the dehydration phase (for induction of tolerance, for synthesis of protectants and for proteomic changes)

The resurrection plant Sporobolus stapfianus: An unlikely model for engineering enhanced plant biomass?

The resurrection grass Sporobolus stapfianus Gandoger can rapidly recover from extended periods of time in the desiccated state (water potential equilibrated to 2% relative humidity) (Gaff and Ellis, Bothalia 11:305&ndash;308 1974; Gaff and Loveys, Transactions of the Malaysian Society of Plant Physiology 3:286&ndash;287 1993). Physiological studies have been conducted in S. stapfianus to investigate the responses utilised by these desiccation-tolerant plants to cope with severe water-deficit. In a number of instances, more recent gene expression analyses in S. stapfianus have shed light on the molecular and cellular mechanisms mediating these responses. S. stapfianus is a versatile research tool for investigating desiccation-tolerance in vegetative grass tissue, with several useful characteristics for differentiating desiccation-tolerance adaptive genes from the many dehydration-responsive genes present in plants. A number of genes orthologous to those isolated from dehydrating S. stapfianus have been successfully used to enhance drought and salt tolerance in model plants as well as important crop species. In addition to the ability to desiccate and rehydrate successfully, the survival of resurrection plants in regions experiencing short sporadic rainfall events may depend substantially on the ability to tightly down-regulate cell division and cell wall loosening activities with decreasing water availability and then grow rapidly after rainfall while water is plentiful. Hence, an analysis of gene transcripts present in the desiccated tissue of resurrection plants may reveal important growth-related genes. Recent findings support the proposition that, as well as being a versatile model for devising strategies for protecting plants from water-loss, resurrection plants may be a very useful tool for pinpointing genes to target for enhancing growth rate and biomass production