Expression Patterns of Genes Involved in Sugar Metabolism and Accumulation during Apple Fruit Development

Both sorbitol and sucrose are imported into apple fruit from leaves. The metabolism of sorbitol and sucrose fuels fruit growth and development, and accumulation of sugars in fruit is central to the edible quality of apple. However, our understanding of the mechanisms controlling sugar metabolism and accumulation in apple remains quite limited. We identified members of various gene families encoding key enzymes or transporters involved in sugar metabolism and accumulation in apple fruit using homology searches and comparison of their expression patterns in different tissues, and analyzed the relationship of their transcripts with enzyme activities and sugar accumulation during fruit development. At the early stage of fruit development, the transcript levels of sorbitol dehydrogenase, cell wall invertase, neutral invertase, sucrose synthase, fructokinase and hexokinase are high, and the resulting high enzyme activities are responsible for the rapid utilization of the imported sorbitol and sucrose for fruit growth, with low levels of sugar accumulation. As the fruit continues to grow due to cell expansion, the transcript levels and activities of these enzymes are down-regulated, with concomitant accumulation of fructose and elevated transcript levels of tonoplast monosaccharide transporters (TMTs), MdTMT1 and MdTMT2; the excess carbon is converted into starch. At the late stage of fruit development, sucrose accumulation is enhanced, consistent with the elevated expression of sucrose-phosphate synthase (SPS), MdSPS5 and MdSPS6, and an increase in its total activity. Our data indicate that sugar metabolism and accumulation in apple fruit is developmentally regulated. This represents a comprehensive analysis of the genes involved in sugar metabolism and accumulation in apple, which will serve as a platform for further studies on the functions of these genes and subsequent manipulation of sugar metabolism and fruit quality traits related to carbohydrates

Aconitase B Is Required for Optimal Growth of Xanthomonas campestris pv. vesicatoria in Pepper Plants

The aerobic plant pathogenic bacterium Xanthomonas campestris pv. vesicatoria (Xcv) colonizes the intercellular spaces of pepper and tomato. One enzyme that might contribute to the successful proliferation of Xcv in the host is the iron-sulfur protein aconitase, which catalyzes the conversion of citrate to isocitrate in the tricarboxylic acid (TCA) cycle and might also sense reactive oxygen species (ROS) and changes in cellular iron levels. Xcv contains three putative aconitases, two of which, acnA and acnB, are encoded by a single chromosomal locus. The focus of this study is aconitase B (AcnB). acnB is co-transcribed with two genes, XCV1925 and XCV1926, encoding putative nucleic acid-binding proteins. In vitro growth of acnB mutants was like wild type, whereas in planta growth and symptom formation in pepper plants were impaired. While acnA, XCV1925 or XCV1926 mutants showed a wild-type phenotype with respect to bacterial growth and in planta symptom formation, proliferation of the acnB mutant in susceptible pepper plants was significantly impaired. Furthermore, the deletion of acnB led to reduced HR induction in resistant pepper plants and an increased susceptibility to the superoxide-generating compound menadione. As AcnB complemented the growth deficiency of an Escherichia coli aconitase mutant, it is likely to be an active aconitase. We therefore propose that optimal growth and survival of Xcv in pepper plants depends on AcnB, which might be required for the utilization of citrate as carbon source and could also help protect the bacterium against oxidative stress

A Bacterial Acetyltransferase Destroys Plant Microtubule Networks and Blocks Secretion

The eukaryotic cytoskeleton is essential for structural support and intracellular transport, and is therefore a common target of animal pathogens. However, no phytopathogenic effector has yet been demonstrated to specifically target the plant cytoskeleton. Here we show that the Pseudomonas syringae type III secreted effector HopZ1a interacts with tubulin and polymerized microtubules. We demonstrate that HopZ1a is an acetyltransferase activated by the eukaryotic co-factor phytic acid. Activated HopZ1a acetylates itself and tubulin. The conserved autoacetylation site of the YopJ / HopZ superfamily, K289, plays a critical role in both the avirulence and virulence function of HopZ1a. Furthermore, HopZ1a requires its acetyltransferase activity to cause a dramatic decrease in Arabidopsis thaliana microtubule networks, disrupt the plant secretory pathway and suppress cell wall-mediated defense. Together, this study supports the hypothesis that HopZ1a promotes virulence through cytoskeletal and secretory disruption

Regulation of the Fruit-Specific PEP Carboxylase SlPPC2 Promoter at Early Stages of Tomato Fruit Development

The SlPPC2 phosphoenolpyruvate carboxylase (PEPC; EC 4.1.1.31) gene from tomato (Solanum lycopersicum) is differentially and specifically expressed in expanding tissues of developing tomato fruit. We recently showed that a 1966 bp DNA fragment located upstream of the ATG codon of the SlPPC2 gene (GenBank AJ313434) confers appropriate fruit-specificity in transgenic tomato. In this study, we further investigated the regulation of the SlPPC2 promoter gene by analysing the SlPPC2 cis-regulating region fused to either the firefly luciferase (LUC) or the β-glucuronidase (GUS) reporter gene, using stable genetic transformation and biolistic transient expression assays in the fruit. Biolistic analyses of 5′ SlPPC2 promoter deletions fused to LUC in fruits at the 8th day after anthesis revealed that positive regulatory regions are mostly located in the distal region of the promoter. In addition, a 5′ UTR leader intron present in the 1966 bp fragment contributes to the proper temporal regulation of LUC activity during fruit development. Interestingly, the SlPPC2 promoter responds to hormones (ethylene) and metabolites (sugars) regulating fruit growth and metabolism. When tested by transient expression assays, the chimeric promoter:LUC fusion constructs allowed gene expression in both fruit and leaf, suggesting that integration into the chromatin is required for fruit-specificity. These results clearly demonstrate that SlPPC2 gene is under tight transcriptional regulation in the developing fruit and that its promoter can be employed to drive transgene expression specifically during the cell expansion stage of tomato fruit. Taken together, the SlPPC2 promoter offers great potential as a candidate for driving transgene expression specifically in developing tomato fruit from various tomato cultivars

Determination of nutrient salts by automatic methods both in seawater and brackish water: the phosphate blank

9 páginas, 2 tablas, 2 figurasThe main inconvenience in determining nutrients in seawater by automatic methods is simply solved: the preparation of a suitable blank which corrects the effect of the refractive index change on the recorded signal. Two procedures are proposed, one physical (a simple equation to estimate the effect) and the other chemical (removal of the dissolved phosphorus with ferric hydroxide).Support for this work came from CICYT (MAR88-0245 project) and Conselleria de Pesca de la Xunta de GaliciaPeer reviewe

The proteasome acts as a hub for plant immunity and is targeted by pseudomonas type III effectors

Recent evidence suggests that the ubiquitin-proteasome system is involved in several aspects of plant immunity and that a range of plant pathogens subvert the ubiquitin-proteasome system to enhance their virulence. Here, we show that proteasome activity is strongly induced during basal defense in Arabidopsis (Arabidopsis thaliana). Mutant lines of the proteasome subunits RPT2a and RPN12a support increased bacterial growth of virulent Pseudomonas syringae pv tomato DC3000 (Pst) and Pseudomonas syringae pv maculicola ES4326. Both proteasome subunits are required for pathogen-associated molecular pattern-triggered immunity responses. Analysis of bacterial growth after a secondary infection of systemic leaves revealed that the establishment of systemic acquired resistance (SAR) is impaired in proteasome mutants, suggesting that the proteasome also plays an important role in defense priming and SAR. In addition, we show that Pst inhibits proteasome activity in a type III secretion-dependent manner. A screen for type III effector proteins from Pst for their ability to interfere with proteasome activity revealed HopM1, HopAO1, HopA1, and HopG1 as putative proteasome inhibitors. Biochemical characterization of HopM1 by mass spectrometry indicates that HopM1 interacts with several E3 ubiquitin ligases and proteasome subunits. This supports the hypothesis that HopM1 associates with the proteasome, leading to its inhibition. Thus, the proteasome is an essential component of pathogenassociated molecular pattern-triggered immunity and SAR, which is targeted by multiple bacterial effectors.This work was supported by the Federation of the European Biochemical Societies (long-term fellowship to S.Ü.), by the DeutscheForschungsgemeinschaft (grant no. CRC973 to F.B.), by the Royal Society (to V.N.), and by the Biotechnology and Biological Sciences Research Council (grant no. BB/L019345/1 to V.N.).Peer Reviewe