Transcription profiling by array of wild type pea seeds and seeds with embryo-specific expression of a bacterial trehalose 6-phosphate phosphatase (TPP), encoded by the otsB gene to investigate the influence of decreased T6P content on gene expression in developing cotyledons.
Ontology highlight
ABSTRACT:
ORGANISM(S): Pisum sativum
SUBMITTER: Tobias Meitzel
PROVIDER: E-MTAB-6659 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:During seed growth, sugar and nitrogen compounds confer regulatory control on storage activities. Thus, seed storage production could be regulated by the supply of nutrients. In order to improve nitrogen flux into the embryo, transgenic pea lines were created where ADP-glucose pyrophosphorylase (AGP) from Pisum sativum has been repressed by RNAi approach in the seeds under control of the seed-specific LeB4 promotor (Bäumlein et al. Cis-analysis of a seed protein gene promoter: the conservative RY repeat CATGCATG within legumin box is essential for tissue-specific expression of a legumin gene. Plant J 1992 2: 233-239). The plastidial enzyme AGP catalyzes the reaction of glucose-1-phosphate and ATP to pyrophosphate and ADP-glucose, which is the substrate for starch synthase. The AGP activity and transcript levels were strongly decreased in three independent transgenic lines. Repression of AGP results in a wrinkled seed phenotype obviously due to transient accumulation of free sugars during maturation. Mature seeds have reduced starch content whereas the protein concentration is higher due to increased fractions of albumins and globulins. Repression of AGP interferes with storage protein metabolism and alters fluxes of nitrogen during seed growth. The influence of decreased AGP on altered gene expression in developing cotyledons was analysed using a 6k-Oligo-microarray. Ps6kOLI1 microarray hybridization were performed using three independent biological replicates of four developmental stages (20, 25, 30 and 35 DAP) from seeds of the transgenic line iAGP-3.
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Project description:Nitrogen application to legume seeds regulates seed metabolism and composition. In order to improve nitrogen flux into the embryo, the Vicia faba amino acid permease VfAAP1 (Miranda et al. Amino acid permeases in developing seeds of Vicia faba L.: expression precedes storage protein synthesis and is regulated by amino acid supply. Plant J 2001 28: 61-72) was expressed in pea under control of the seed-specific LeB4 promotor (Bäumlein et al. Cis-analysis of a seed protein gene promoter: the conservative RY repeat CATGCATG within legumin box is essential for tissue-specific expression of a legumin gene. Plant J 1992 2: 233-239). Several transgenic lines have been generated. Mature seeds of the homozygous marker gene-free line AAP14/10 showed an increase in amino acid supply, seed nitrogen and protein content due to higher globulin fraction. The effect of VfAAP1 was tested under field conditions in two growing periods, 2005 and 2006, and the data could be confirmed. Over-expression of VfAAP1 interferes with storage protein metabolism and alters fluxes of nitrogen during seed growth. The influence of VfAAP1 on altered gene expression in developing cotyledons was analysed using a 6k-Oligo-microarray. Four developmental stages (18, 22, 26 and 30 DAP) from seeds, grown 2006, of the transgenic line AAP14/10 were hybridized against wildtype control.
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Project description:ABA regulates in plants a wide range of developmental events, mediates responses to environmental stress and is necessary to proceed through seed maturation and to acquire desiccation tolerance and dormancy. Immuno-modulation is a suitable means to study ABA functions during seed maturation. Anti-ABA single chain antibody was expressed in pea seed driving LeB4-promoter (Saalbach et al., High-level expression of a single chain Fv fragment (scFv) antibody in transgenic pea seeds J. Plant Physiol. 2001 158: 529-533), which produced only a weak phenotype with slightly decreased seed weight, globulin/albumin and total nitrogen content (aABA line 16 cultivar Erbi). In another approach with a stronger, improved USP-promoter used to express the anti-ABA antibody in pea seeds a different phenotype emerged (aABA line 7, cultivar Eifel). In this line individual seed weight increased by 20 to 30% together with higher globulin and albumin content. To dissect the aABA phenotype at the molecular level, a search for genes with differential expression patterns in transgenic plant versus wild type seeds has been performed using 6k-oligo microarray analysis. cDNA probes were prepared from RNA isolated from embryo of developing seeds of wild type (12, 18, and 22 DAP) and transgenic aABA plants (12, 18, and 22 DAP), which correspond to the transition phase of seed development, and 6k-oligo microarray.
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Project description:Posttranslational lysine acetylation is believed to occur in all taxa and to affect thousands of proteins. In contrast to the hundreds of mitochondrial proteins reported to be lysine-acetylated in non-plant species, only a handful have been reported from the plant taxa previously examined. To investigate whether this reflects a biologically significant difference or merely a peculiarity of the samples thus far examined, we immunoenriched and analyzed acetylated peptides from highly purified pea seedling mitochondria using mass spectrometry. Our results indicate that a multitude of mitochondrial proteins, involved in a variety of processes, are acetylated in pea seedlings.
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Project description:Aphid adaptation to harsh winter conditions is illustrated by an alternation of their reproductive mode. Aphids detect photoperiod shortening by sensing the length of the night and switch from viviparous parthenogenesis in spring and summer, to oviparous sexual reproduction in autumn. The photoperiodic signal is transduced from the head to the reproductive tract to change the fate of the future oocytes from mitotic diploid embryogenesis to haploid formation of gametes. Because of viviparous parthenogenesis, the whole process takes place in three consecutive generations. To understand the molecular basis of the switch in the reproductive mode, a transcriptomic approach was used to detect significantly regulated transcripts in the heads of the pea aphid Acyrthosiphon pisum. The transcriptomic profiles of the heads of the first generation were slightly affected by photoperiod shortening. This suggests that trans-generation signaling does not occur between the grand-mothers and the viviparous embryos they contain. By analogy, many of the genes regulated in the heads of the second generation are implicated in visual functions, photoreception and cuticle structure. The modification of the cuticle could decrease the storage of N-β-alanyldopamine and provoke an increase in free dopamine concentration. Based in results in Drosophila, modification of the insulin pathway could cause a decrease of juvenile hormones in short-day reared aphids. Biological material for microarray experiments was prepared under two dayly photoperiodic regimes both at constant temperature of 18°C: i) âShort Nightâ (SN) at 16h of light and ii) âLong Nightâ (LN) at 12h of light to induce the production of sexual morphs. To initiate the experiment, two groups of 105 L3 larvae were placed either under SN or LN condition. This corresponds to generation G0. At the middle of the photophase, 25 individual were frozen when they had reached both the L4 and the wingless adult (WA) stages, in the two photoperiod conditions. The 55 remaining WA individuals (still divided in two groups) were left on 55 plants to lay their offspring: one larva of the 1st stage (L1) was kept per WA. This larva was selected among the 20 first born larvae. This is the generation G1. At the middle of the photophase, 25 individual were frozen when they had reached both the L2 and the L4 stages, in the two photoperiodic conditions. Thus, 25 individuals from 4 different stages (L4-G0, WA-G0, L2-G1 and L4-G1) were collected in the two photoperiod conditions with 3 biological replicates, forming the 24 samples used for microarray experiments. RNAs from heads of aphids from the two photoperiodic conditions were hybridized one against the other for each stage with a dye-swap.The experimental design is thus 24 arrays which corresponds to the described samples of that series.
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Project description:About 475 million years ago plants originated from an ancestral green alga and evolved first as non-vascular and later as vascular plants, becoming the primary producers of biomass on lands. During that time, the light harvesting complex II (LHCII), responsible for sunlight absorption and excitation energy transfer to the Photosystem II (PSII) core, underwent extensive differentiation. Lhcb4 is an ancestral ubiquitous LHCII that during plant diversification continued to change, finally differentiating in flowering plants in up to three isoforms, Lhcb4.1, Lhcb4.2 and Lhcb4.3. The pivotal position of Lhcb4 in the PSII-LHCII supercomplex (PSII-LHCIIsc), as linker for S- and M-trimers of LHCII to the PSII core, suggests its major role in light harvesting modulation. The increased accumulation of Lhcb4.3 observed in PSII-LHCIIsc of plants acclimated to moderate and high light intensities induced us to investigate whether this isoform is present in a specific PSII-LHCIIsc conformation that might explain its light-dependent accumulation. In this work, by combining an improved method for separation of different forms of PSII-LHCIIsc from thylakoids of Pisum sativum grown at increasing irradiances with quantitative proteomics, we assessed that Lhcb4.3 is abundant in PSII-LHCIIsc of type C2S2, irrespective of the plant growth irradiance. Phylogenetic comparative analysis on different taxa of the Viridiplantae lineage and structural modelling further pointed out to an effect of the evolution of different Lhcb4 isoforms on the light-dependent modulation of the PSII-LHCIIsc organization. This information provides new insight on the properties of the Lhcb4 and its isoforms and their role on the structure, function and regulation of PSII.
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