Project description:ra15-03_oxpmt7-7 - acyltransferase brachypodium-arabidopsis - What biochemical function? What consequence on lignification when overexpressed in Arabidopsis thaliana? - Transcripts of Arabidopsis stem overexpressing an acyl transferase from Brachypodium are compared to wild type plants. The overexpression is under a specific promoter (C4H).
Project description:Numerous proteins synthesized in cells ranging from bacteria to humans have to be posttranslationally acylated to become biologically active. Bacterial Repeats in ToXin (RTX) cytolysins form a prominent group of proteins that are synthesized as inactive protoxins and undergo a posttranslational acylation on ε-amino groups of two internal conserved lysine residues by co-expressed toxin-activating acyltransferases. We investigated how the chemical nature, position and number of bound acyl chains govern the activities of Bordetella pertussis adenylate cyclase toxin (CyaA), Escherichia coli α-hemolysin (HlyA) and Kingella kingae cytotoxin (RtxA). The three protoxins were acylated in the same E. coli cell background by either of the CyaC, HlyC and RtxC acyltransferases. The results revealed that the acyltransferase itself selects from the acyl-ACP pool of the producing bacterium the type of the acyl chain of adapted length to be covalently linked to the protoxin. The acyltransferase also selects whether both or only one of two conserved lysine residues of the protoxin will be posttranslationally acylated. Functional assays then revealed that RtxA has to be modified by 14-carbon fatty acyl chains to be biologically active, while HlyA remains active also when modified by 16-carbon acyl chains and CyaA is activated exclusively by 16-carbon acyl chains. These results suggest a structural adaptation of the toxin molecules to the length of the acyl chains used for modification of their crucial acylated lysine residue in the second, more conserved acylation site
Project description:Ghrelin, an orexigenic gut-derived peptide, is gaining increasing attention due to its multifaceted role in a number of physiological functions, including metabolism, cardiovascular health, stress and reproduction. Ghrelin exists in circulation primarily as des-acylated and acylated ghrelin. Des-acyl ghrelin, until recently considered to be an inactive form ghrelin, is now known to have independent physiological functionality. However, the relative contribution of acyl and des-acyl ghrelin to reproductive development and function is currently unknown. Here we used ghrelin-O-acyltransferase (GOAT) knockout (KO) mice that have no measurable levels of endogenous acyl ghrelin and chronically high levels of des-acyl ghrelin, to characterise how the developmental and life-long absence of acyl ghrelin affects ovarian development and reproductive capacity. We have combined ovarian transcriptome analysis using RNA sequencing with measures of ovarian morphometry, as well as with the assessment of markers of reproductive maturity and the capacity to breed. Our data show pronounced specific changes in the ovarian transcriptome in the juvenile GOAT KO ovary, indicative of advanced ovarian development. These changes corresponded with diminished ovarian reserve in the juvenile and adult ovaries of these mice, due to a continuous reduction in the number of small follicle populations. These changes did not affect the timing of puberty onset or reproductive capacity under optimal conditions. These data suggest that an absence of acyl ghrelin does not prevent reproductive success but that appropriate levels of acyl and des-acyl ghrelin may be necessary for optimal ovarian maturation.
Project description:Anthocyanins are flavonoid compounds responsible for red/purple colours in the leaves, fruit and flowers of many plant species. They are produced through a multistep pathway which is controlled by MYB transcription factors. VvMYBA1 and VvMYBA2 activate anthocyanin biosynthesis in grapevine (Vitis vinifera) and are non-functional in white grapevine cultivars. In this study, transgenic grapevines with altered VvMYBA gene expression were developed, and transcript analysis was carried out on berries using a microarray technique. The results showed that VvMYBA is a positive regulator of the later stages of anthocyanin synthesis, modification and transport in Shiraz. One up-regulated gene ANTHOCYANIN 3- O-GLUCOSIDE-6”-O-ACYLTRANSFERASE (Vv3AT) encodes a BAHD acyltransferase protein, belonging to a clade separate from most anthocyanin acyltransferases. Functional studies (in planta and in vitro) show that Vv3AT has a broad anthocyanin substrate specificity and can also utilise both aliphatic and aromatic acyl donors, a novel activity for this enzyme family found in nature. In V. vinifera cv. Pinot Noir, a red-berried grapevine mutant lacking acylated anthocyanins, Vv3AT contains a nonsense mutation encoding a truncated protein that lacks two motifs required for BAHD protein activity. Promoter activation assays confirm that Vv3AT transcription is activated by VvMYBA1, which adds to the current understanding of the regulation of the BAHD gene family. The flexibility of Vv3AT to use both classes of acyl donors will be useful in the engineering of anthocyanins in planta or in vitro.
Project description:ngs2020_04_hipath-differential expression analysis to hight co2 of the brachypodium distachyon-Analysis response of the Brachypodium distachyon to hight CO2 -treatment hight CO2
Project description:Plants and microalgae have the ability to harness sunlight into energy dense molecules such as triacylglycerols (TAGs). TAGs hold great importance for human and animal nutrition, and are also a source for renewable energy. Acyl-CoA-dependent TAG synthesis by diacylglycerol acyltransferases (DGATs) in oilseeds has been well characterized, but how the ectopic introduction of TAG synthesis in vegetative tissues of plants affects metabolism is largely unknown. Here we report the identification and characterization of several Chlamydomonas reinhardtii Diacylglycerol Acyltransferase Type Two (DGTT) enzymes and their use in engineering TAG biosynthesis in plant vegetative tissues. Focusing on DGTT2 we demonstrate that it can accept a broad range of substrates. Production of DGTT2 in Arabidopsis results in distinct seedling phenotypes and accumulation of TAG with very long chain fatty acids (VLCFA), in both seedlings (23.4-fold increase) and in the leaves of soil-grown plants (13.3-fold increase). Levels of surface lipids, such as waxes and cutins, are decreased in DGTT2-producing lines, consistent with a decreased carbon/acyl flux to the surface lipid pathway. Global transcript analysis showed that very few genes had altered expression. Little to no change in the transcripts of wax/cutin pathway genes was observed in response to altered carbon partitioning into TAGs in the leaves suggesting that the supply of acyl groups for surface lipid biosynthesis is not transcriptionally adjusted in the transgenic lines. As an indication that the DGTT2-producing plants are richer in feed value, the generalist herbivore Spodoptera exigua reared on the transgenic plants gained more weight. Apparently, the substrate specificity of DGTT2 from Chlamydomonas when produced in the Arabidopsis cells leads to diversion of carbon from surface compounds in TAGs and enhances the nutritional value of leaves. 4 biological replicates DGTT2 transgenic plants are compared to 4 biological replicates of wild type
Project description:Anthocyanins are flavonoid compounds responsible for red/purple colours in the leaves, fruit and flowers of many plant species. They are produced through a multistep pathway which is controlled by MYB transcription factors. VvMYBA1 and VvMYBA2 activate anthocyanin biosynthesis in grapevine (Vitis vinifera) and are non-functional in white grapevine cultivars. In this study, transgenic grapevines with altered VvMYBA gene expression were developed, and transcript analysis was carried out on berries using a microarray technique. The results showed that VvMYBA is a positive regulator of the later stages of anthocyanin synthesis, modification and transport in Shiraz. One up-regulated gene ANTHOCYANIN 3- O-GLUCOSIDE-6â-O-ACYLTRANSFERASE (Vv3AT) encodes a BAHD acyltransferase protein, belonging to a clade separate from most anthocyanin acyltransferases. Functional studies (in planta and in vitro) show that Vv3AT has a broad anthocyanin substrate specificity and can also utilise both aliphatic and aromatic acyl donors, a novel activity for this enzyme family found in nature. In V. vinifera cv. Pinot Noir, a red-berried grapevine mutant lacking acylated anthocyanins, Vv3AT contains a nonsense mutation encoding a truncated protein that lacks two motifs required for BAHD protein activity. Promoter activation assays confirm that Vv3AT transcription is activated by VvMYBA1, which adds to the current understanding of the regulation of the BAHD gene family. The flexibility of Vv3AT to use both classes of acyl donors will be useful in the engineering of anthocyanins in planta or in vitro. Transgenic Chardonnay/Shiraz and non-transformed WT controls were all grown in the same glasshouse in ambient light with a night break. Day and night temperatures were approximately 27oC and 22oC respectively. For microarray experiments, whole berries were sampled from independent transgenic lines: three from transgenic Chardonnay and four from transgenic Shiraz, resulting in three and four biological replicates respectively. Bunches were harvested close to ripeness based on average total soluble solids (TSS, measured as oBrix). This was aimed to be between 20 â 24 oBrix and was determined from TTS of a subsamples from each bunch (Table S53). A sample consisted of all remaining berries from a single bunch except when there were <100 berries in which case more than one bunch was used in the one replicate. Whole berries were immediately frozen in liquid nitrogen. For skin samples, the skins were first removed from fresh berries then frozen in liquid nitrogen. All samples were stored at -80oC.
Project description:Due to its small and sequenced genome, short generation time, efficient transformation and increasing genetic resources, Brachypodium distachyon is an emerging model for grasses. Despite this, data capturing gene expression patterns across different organs and developmental stages is missing. We have generated a comprehensive gene expression atlas for Brachypodium, capturing 9 different organs and developmental stages
Project description:We used Brachypodium distachyon (BD21) as a model grass to gain insight into the affected host molecular pathways upon infection of Panicum Mosaic Virus (PMV) together with its satellite virus, Satellite Panicum Mosaic Virus (SPMV). Brachypodium plants at 2-3 leaf stage were either mock inoculated or inoculated with PMV and PMV+SPMV. Total RNA was isolated from shoot tissues of control and treated plants and was subjected to microarray analysis.
Project description:This SuperSeries is composed of the following subset Series: GSE26802: Post-transcriptional generation of miRNA variants by multiple nucleotidyl transferases contributes to miRNA transcriptome complexity (hESC) GSE26803: Post-transcriptional generation of miRNA variants by multiple nucleotidyl transferases contributes to miRNA transcriptome complexity (KD) GSE26916: Post-transcriptional generation of miRNA variants by multiple nucleotidyl transferases contributes to miRNA transcriptome complexity (Validation 1) GSE26919: Post-transcriptional generation of miRNA variants by multiple nucleotidyl transferases contributes to miRNA transcriptome complexity (Validation 2) GSE29904: Post-transcriptional generation of miRNA variants by multiple nucleotidyl transferases contributes to miRNA transcriptome complexity (normal prostate and prostate cells) Refer to individual Series