Transcription profiling by array of potato tubers with altered PME activity
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ABSTRACT: Although processed potato tuber texture is an important trait that influences consumer preference, a detailed understanding of tuber textural properties at the molecular level is lacking. Previous work has identified tuber pectin methyl esterase activity (PME) as a potential factor impacting on textural properties and the expression of a gene encoding an isoform of PME (PEST1) was associated with cooked tuber textural properties. In this study a transgenic approach was undertaken to investigate further the impact of the PEST1 gene. Antisense and over-expressing potato lines were generated. In over-expressing lines tuber PME activity was enhanced by up to 2.3 fold whereas in antisense lines PME activity was decreased by up to 38%. PME isoform analysis indicated that the PEST1 gene encoded one isoform of PME. Analysis of cell walls from tubers from the over-expressing lines indicated that the changes in PME activity resulted in a decrease in pectin methylation. Analysis of processed tuber texture demonstrated that the reduced level of pectin methylation in the over-expressing transgenic lines was associated with a firmer processed texture. Thus there is a clear link between PME activity, pectin methylation and processed tuber textural properties.
Project description:Similar to other plant-parasitic nematodes, root lesion nematodes possess an array of enzymes that are involved in degradation of the plant cell wall. Here we report the identification of a gene encoding a cell wall degrading enzyme, pectin methylesterase PME (EC 3.1.1.11), in the root lesion nematode Pratylenchus penetrans. Both genomic and coding sequences of the gene were cloned for this species, showing the presence of four introns that excluded a potential bacterial contamination. Expression of the Pp-pme gene was localized in the esophageal glands of P. penetrans as determined by in situ hybridization. Temporal expression of Pp-pme in planta was validated for early time points of infection. The possible function and activity of the gene were assessed by transient expression of Pp-pme in N. benthamiana plants via a Potato virus X-based vector. To our knowledge, this is the first report on identification and characterization of a PME gene within the phylum Nematoda.
Project description:During senescence of detached rice leaves, tryptophan (Trp) and Trp-derived secondary metabolites such as serotonin and 4-coumaroylserotonin accumulated in concert with methanol (MeOH) production. This senescence-induced MeOH induction was closely associated with levels of pectin methylesterase (PME)1 mRNA and PME enzyme activity. Exogenous challenge of detached rice leaves with 1% MeOH accelerated Trp and serotonin biosynthesis with induction of the corresponding genes. No other solvents including ethanol resulted in a Trp-inducing effect. This MeOH-induced Trp synthesis was positively regulated by abscisic acid but negatively regulated by cytokinin, suggesting hormonal involvement on the action of MeOH. Endogenous overproduction or suppression of MeOH either by PME1 overexpression or RNAi gene silencing revealed that PME1 overexpressing lines produced twofold higher Trp levels with elevated Trp biosynthetic gene expression, whereas RNAi lines showed twofold reduction in Trp level in healthy control rice leaves, suggesting that MeOH acts as an endogenous elicitor to enhance Trp biosynthesis. Among many transcription factors induced following MeOH treatment, the WRKY family showed significant induction patterns of which WRKY14 appeared to play a key regulatory role in MeOH-induced Trp and Trp-derived secondary metabolite biosynthesis.
Project description:The experiment followed transcriptional changes during potato tuber induction from a stolon tip to a tuber. Samples were taken at stage 1, stage 3, stage 4 and stage 5 according to Kloosterman et al., 2005
Project description:The aim of this study was to conduct a genome-wide analysis for constituent tuber carotenoid QTL. Using a genetical genomics approach samples from clones with similar carotenoid traits were bulked and patterns of gene expression were measured for each bulk by microarray analysis. Variation of gene expression within these bulks may be due to either polymorphisms located near to or within the gene (cis-eQTL) or indirectly from a distant location on the genome (trans-eQTL). Differentially expressed clones from bulks with contrasting carotenoid traits were genetically mapped in order to re-enforce the QTL analysis, and provide a rapid means of developing gene markers closely associated with the target traits.
Project description:Pectins localize in the primary cell wall and consist of multiblock co?polymers among which homogalacturonan (HG) is the simplest and most abundant form. Methylesterification pattern of HG is tuned by pectin methylesterases (PMEs), which activity is itself controlled by specific inhibitors or PMEIs. PME?mediated regulation of HG methylesterification plays a major role in controlling several developmental processes, including seed germination and dark grown hypocotyl elongation. Arabidopsis PME36 is preferentially expressed during the late stages of silique development and, using the knock?out mutant pme36-1, we showed that PME36 is required to implement the characteristic pattern of demethylesterified pectins in mature seed. However, although this pattern was strongly impaired in pme36-1 mature seed, no phenotypical change was observed in the mutant during seed germination and seedling dark growth, suggesting the existence of a compensatory mechanism overcoming the defect in pectin demethylesterification. To get insight into this mechanism, a transcriptomic analysis by microarrays was performed on pme36-1 and Col0 wild-type, in seed at two early stages of germination, preceding tegument rupture: 16 hours and 24 hours after transfer from cold room (stratification) to dark growth condition.