Despite the fact that the organic acid content of a fruit is regarded as one of its most commercially important quality traits when assessed by the consumer, relatively little is known concerning the physiological importance of organic acid metabolism for the fruit itself. Here, we evaluate the effect of modifying malate metabolism in a fruit-specific manner, by reduction of the activities of either mitochondrial malate dehydrogenase or fumarase, via targeted antisense approaches in tomato (Solanum lycopersicum). While these genetic perturbations had relatively little effect on the total fruit yield, they had dramatic consequences for fruit metabolism, as well as unanticipated changes in postharvest shelf life and susceptibility to bacterial infection. Detailed characterization suggested that the rate of ripening was essentially unaltered but that lines containing higher malate were characterized by lower levels of transitory starch and a lower soluble sugars content at harvest, whereas those with lower malate contained higher levels of these carbohydrates. Analysis of the activation state of ADP-glucose pyrophosphorylase revealed that it correlated with the accumulation of transitory starch. Taken together with the altered activation state of the plastidial malate dehydrogenase and the modified pigment biosynthesis of the transgenic lines, these results suggest that the phenotypes are due to an altered cellular redox status. The combined data reveal the importance of malate metabolism in tomato fruit metabolism and development and confirm the importance of transitory starch in the determination of agronomic yield in this species.

Recent advances in genomics and proteomics have provided an excellent opportunity to understand complex biological processes such as wood formation at the gene and protein levels. The aim of this work was to describe the proteins participating in the processes involved in juvenile wood formation by isolating proteins from the cambial region of Eucalyptus grandis, at three ages of growth (6-month-old seedlings, 3- and 6-year-old trees), and also to identify proteins differentially expressed. Using a 2-D-LC-MS/MS strategy we identified a total of 240 proteins, with 54 corresponding spots being present in at least two ages. Overall, nine proteins classified into the functional categories of metabolism, cellular processes, and macromolecular metabolism showed significant changes in expression. Proteins were classified into seven main functional categories, with metabolism representing 35.2% of the total proteins identified. The comparison of the reference maps showed not only differences in the expression pattern of individual proteins at each age, but also among isoforms. The results described in this paper provide a dynamic view of the proteins involved in the formation of juvenile wood in E. grandis.

SummaryTomato (Solanum lycopersicum) fruit ripening is regulated co‐operatively by the action of ethylene and a hierarchy of transcription factors, including RIPENING INHIBITOR (RIN) and NON‐RIPENING (NOR). Mutations in these two genes have been adopted commercially to delay ripening, and accompanying textural deterioration, as a means to prolong shelf life. However, these mutations also affect desirable traits associated with colour and nutritional value, although the extent of this trade‐off has not been assessed in detail. Here, we evaluated changes in tomato fruit pericarp primary metabolite and carotenoid pigment profiles, as well as the dynamics of specific associated transcripts, in the rin and nor mutants during late development and postharvest storage, as well of those of the partially ripening delayed fruit ripening (dfd) tomato genotype. These profiles were compared with those of the wild‐type tomato cultivars Ailsa Craig (AC) and M82. We also evaluated the metabolic composition of M82 fruit ripened on or off the vine over a similar period. In general, the dfd mutation resulted in prolonged firmness and maintenance of quality traits without compromising key metabolites (sucrose, glucose/fructose and glucose) and sectors of intermediary metabolism, including tricarboxylic acid cycle intermediates. Our analysis also provided insights into the regulation of carotenoid formation and highlighted the importance of the polyamine, putrescine, in extending fruit shelf life. Finally, the metabolic composition analysis of M82 fruit ripened on or off the vine provided insights into the import into fruit of compounds, such as sucrose, during ripening.

The rhizosphere is an ecosystem exploited by a variety of organisms involved in plant health and environmental sustainability. Abiotic factors influence microorganism-plant interactions, but the microbial community is also affected by expression of heterologous genes from host plants. In the present work, we assessed the community shifts of Alphaproteobacteria phylogenetically related to the Rhizobiales order (Rhizobiales-like community) in rhizoplane and rhizosphere soils of wild-type and transgenic eucalyptus. A greenhouse experiment was performed and the bacterial communities associated with two wild-type (WT17 and WT18) and four transgenic (TR-9, TR-15, TR-22, and TR-23) eucalyptus plant lines were evaluated. The culture-independent approach consisted of the quantification, by real-time polymerase chain reaction (PCR), of a targeted subset of Alphaproteobacteria and the assessment of its diversity using PCR-denaturing gradient gel electrophoresis (DGGE) and 16S rRNA gene clone libraries. Real-time quantification revealed a lesser density of the targeted community in TR-9 and TR-15 plants and diversity analysis by principal components analysis, based on PCR-DGGE, revealed differences between bacterial communities, not only between transgenic and nontransgenic plants, but also among wild-type plants. The comparison between clone libraries obtained from the transgenic plant TR-15 and wild-type WT17 revealed distinct bacterial communities associated with these plants. In addition, a culturable approach was used to quantify the Methylobacterium spp. in the samples where the identification of isolates, based on 16S rRNA gene sequences, showed similarities to the species Methylobacterium nodulans, Methylobacterium isbiliense, Methylobacterium variable, Methylobacterium fujisawaense, and Methylobacterium radiotolerans. Colonies classified into this genus were not isolated from the rhizosphere but brought in culture from rhizoplane samples, except for one line of the transgenic plants (TR-15). In general, the data suggested that, in most cases, shifts in bacterial communities due to cultivation of transgenic plants are similar to those observed when different wild-type cultivars are compared, although shifts directly correlated to transgenic plant cultivation may be found.

Despite the importance of Eucalyptus spp. in the pulp and paper industry, functional genomic approaches have only recently been applied to understand wood formation in this genus. We attempted to establish a global view of gene expression in the juvenile cambial region of Eucalyptus grandis Hill ex Maiden. The expression profile was obtained from serial analysis of gene expression (SAGE) library data produced from 3- and 6-year-old trees. Fourteen-base expressed sequence tags (ESTs) were searched against public Eucalyptus ESTs and annotated with GenBank. Altogether 43,304 tags were generated producing 3066 unigenes with three or more copies each, 445 with a putative identity, 215 with unknown function and 2406 without an EST match. The expression profile of the juvenile cambial region revealed the presence of highly frequent transcripts related to general metabolism and energy metabolism, cellular processes, transport, structural components and information pathways. We made a quantitative analysis of a large number of genes involved in the biosynthesis of cellulose, pectin, hemicellulose and lignin. Our findings provide insight into the expression of functionally related genes involved in juvenile wood formation in young fast-growing E. grandis trees.