Search Results - (Author, Cooperation:A. R. Fernie)

2011-12-14

American Association for the Advancement of Science (AAAS)

0036-8075

1095-9203

Biology

Chemistry and Pharmacology

Computer Science

Medicine

Natural Sciences in General

Physics

Animals ; Arabidopsis/genetics/growth & development/*metabolism ; Arabidopsis Proteins/genetics/*metabolism ; Biological Transport ; Cell Membrane/metabolism ; Fluorescence Resonance Energy Transfer ; HEK293 Cells ; Humans ; Membrane Transport Proteins/genetics/*metabolism ; Mutant Proteins/metabolism ; Oryza/metabolism ; Phloem/*metabolism ; Plant Leaves/metabolism ; Plant Proteins/genetics/metabolism ; Plant Roots/growth & development ; Promoter Regions, Genetic ; Sucrose/*metabolism

2011-05-13

Nature Publishing Group (NPG)

0028-0836

1476-4687

Biology

Chemistry and Pharmacology

Medicine

Natural Sciences in General

Physics

Carbamoyl-Phosphate Synthase (Ammonia)/metabolism ; Diatoms/*classification/enzymology/genetics/growth & development/*metabolism ; Gene Expression Regulation ; Gene Knockdown Techniques ; Nitrates/metabolism ; *Photosynthesis ; *Phylogeny ; RNA Interference ; Urea/*metabolism

2011-10-07

Nature Publishing Group (NPG)

0028-0836

1476-4687

Biology

Chemistry and Pharmacology

Medicine

Natural Sciences in General

Physics

Chorismate Mutase/*metabolism ; Cytoplasm/enzymology ; Gene Expression Regulation, Plant ; Genetic Complementation Test ; Host-Pathogen Interactions ; Metabolome ; Models, Biological ; Plant Proteins/metabolism ; Plastids/enzymology ; Protein Multimerization ; Saccharomyces cerevisiae/genetics ; Salicylic Acid/metabolism ; Two-Hybrid System Techniques ; Ustilago/*enzymology/*pathogenicity ; Virulence Factors/genetics/*metabolism ; Zea mays/*metabolism/*microbiology

1365-3040

Blackwell Publishing Journal Backfiles 1879-2005

Biology

Sugars are not only metabolic substrates: they also act as signals that regulate the metabolism of plants. Previously, we found that glycolysis is induced in transgenic tubers expressing a yeast invertase in the cytosol but not in those expressing invertase in the apoplast. This suggests that either the low level of sucrose, the increased formation of cytosolic glucose or the increased levels of metabolites downstream of the sucrose cleavage is responsible for the induction of glycolysis in storage organs. In order to discriminate between these possibilities, we cloned and expressed a bacterial sucrose phosphorylase gene from Pseudomonas saccharophila in potato tubers. Due to the phosphorolytic cleavage of sucrose, formation of glucose was circumvented, thus allowing assessment of the importance of cytosolic glucose – and, by implication, flux through hexokinase – in glycolytic induction. Expression of sucrose phosphorylase led to: (i) a decrease in sucrose content, but no decrease in glucose or fructose; (ii) a decrease in both starch accumulation and tuber yield; (iii) increased levels of glycolytic metabolites; (iv) an induction of the activities of key enzymes of glycolysis; and (v) increased respiratory activity. We conclude that the induction of glycolysis in heterotrophic tissues such as potato tubers occurs via a glucose-independent mechanism.

Electronic Resource

1365-3040

Blackwell Publishing Journal Backfiles 1879-2005

Biology

As reported in a previous paper (Plant, Cell and Environment 24, 357–365, 2001), introduction of sucrose phosphorylase into the cytosol of potato results in increased respiration, an inhibition of starch accumulation and decreased tuber yield. Herein a more detailed investigation into the effect of sucrose phosphorylase expression on tuber metabolism, in order to understand why storage and growth are impaired is described. (1) Although the activity of the introduced sucrose phosphorylase was low and accounted for less than 10% of that of sucrose synthase its expression led to a decrease in the activities of enzymes of starch synthesis relative to enzymes of glycolysis and relative to total amylolytic activity. (2) Incubation of tuber discs in [14C]glucose revealed that the transformants display a two-fold increase of the unidirectional rate of sucrose breakdown. However this was largely compensated by a large stimulation of sucrose re-synthesis and therefore the net rate of sucrose breakdown was not greatly affected. Despite this fact major shifts in tuber metabolism, including depletion of sucrose to very low levels, higher rates of glycolysis, and larger pools of amino acids were observed in these lines. (3) Expression of sucrose phosphorylase led to a decrease of the cellular ATP/ADP ratio and energy charge in intact growing tubers. It was estimated that at least 30% of the ATP formed during respiration is consumed as a result of the large acceleration of the cycle of sucrose breakdown and re-synthesis in the transformants. Although the absolute rate of starch synthesis in short-term labelling experiments with discs rose, starch synthesis fell relative to other fluxes including respiration, and the overall starch content of the tubers was lower than in wild-type tubers. (4) External supply of amino acids to replace sucrose as an osmoticum led to a feed-back inhibition of glycolysis, but did not restore allocation to starch. (5) However, an external supply of the non-metabolizable sucrose analogue palatinose – but not sucrose itself – stimulated flux to starch in the transformants. (6) The results indicate that the impaired performance of sucrose phosphorylase-expressing tubers is attributable to decreased levels of sucrose and increased energy consumption during sucrose futile cycling, and imply that sucrose degradation via sucrose synthase is important to maintain a relatively large sucrose pool and to minimize the ATP consumption required for normal metabolic function in the wild type.

Electronic Resource

1365-3040

Blackwell Publishing Journal Backfiles 1879-2005

Biology

Starch is of great importance both as a carbon storage reserve in plants and as a biotechnologically important product. The potato tuber is an attractive model system for the study of starch metabolism, because it is a relatively homogenous tissue in which conversion of sucrose to starch represents the dominant metabolic flux. All the major genes of the potato tuber sucrose to starch pathway have been cloned in recent years, allowing the generation of a suite of antisense transgenic lines to be produced in which the activity of each individual enzyme in the pathway is progressively decreased. Investigations of these plants have provided a complete picture of the distribution of control in this important pathway. Sucrose synthase, UGPase, hexokinase, cytosolic phosphoglucomutase, plastidial phosphoglucomutase, the amyloplastidial adenylate translocator, AGPase, starch synthase and starch branching enzyme have flux control coefficients (FCCs) of 0.10, approximating 0.00, approximating 0.00, 0.15, 0.23, 0.98, 0.35, 0.12 and approximating 0.00 for starch accumulation. These results show that the majority of the control on starch accumulation in potato tubers resides in the transfer of adenylate between the cytosol and the amyloplast, with a minor contribution being made by the first two steps of the plastidial starch synthesis pathway (the reactions catalysed by plastidial phosphoglucomutase and AGPase). This contrasts with leaves, in which the majority of the control has been found to reside in the reactions catalysed by plastidial phosphoglucomutase and AGPase. In leaves, ATP for starch synthesis is generated within the plastid via photophosphorylation. Several studies have attempted to increase the rate of starch synthesis by overexpressing pathway enzymes in tubers. The results of these studies and the role of other ATP producers in the starch synthetic process are reviewed. In the same time period methods of non-aqueous fractionation have been adapted to potato tuber tissue in order to ascertain subcellular metabolite levels. Results obtained from these studies allow the calculation of mass action ratios of the constitutive enzymes of the sucrose to starch transition. When taken together with the known regulatory properties of these enzymes the combination of broad control analysis studies and assessment of the mass action ratios of the respective enzymes allows a comprehensive description of this important metabolic network. Some illustrative examples of how this network responds to environmental change are presented. Finally implications of this whole pathway evaluation for more general studies of plant metabolic pathways and networks are discussed.

Electronic Resource