Transcriptomic analysis comparing stay-green and senescent Sorghum bicolor lines identifies a role for proline biosynthesis in the stay-green trait.

Sorghum bicolor is an important cereal crop grown on the arid and semi-arid regions of >98 different countries. These regions are such that this crop is often subjected to low water conditions, which can compromise yields. Stay-green sorghum plants are able to retain green leaf area for longer under drought conditions and as such have higher yields than their senescent counterparts.

A Novel Function for Arabidopsis CYCLASE1 in Programmed Cell Death Revealed by Isobaric Tags for Relative and Absolute Quantitation (iTRAQ) Analysis of Extracellular Matrix Proteins.

Programmed cell death is essential for plant development and stress adaptation. A detailed understanding of the signal transduction pathways that regulate plant programmed cell death requires identification of the underpinning protein networks. Here, we have used a protagonist and antagonist of programmed cell death triggered by fumonisin B1 as probes to identify key cell death regulatory proteins in Arabidopsis.

Transcriptomic analysis of Sorghum bicolor responding to combined heat and drought stress.

Background: Abiotic stresses which include drought and heat are amongst the main limiting factors for plant growth and crop productivity. In the field, these stress types are rarely presented individually and plants are often subjected to a combination of stress types. Sorghum bicolor is a cereal crop which is grown in arid and semi-arid regions and is particularly well adapted to the hot and dry conditions in which it originates and is now grown as a crop.

Light regulation of cadmium-induced cell death in Arabidopsis.

Cadmium is an environmental pollutant with deleterious effects on both prokaryotic and eukaryotic organisms. In plants, the effects of cadmium toxicity are concentration dependent; lower doses destabilize many physiological processes and inhibit cell growth and multiplication, while higher doses evoke a more severe response that triggers activation of cell death. We recently investigated the effects of light on cadmium toxicity in Arabidopsis using a cell suspension culture system.

UDP-glucose pyrophosphorylase is a novel plant cell death regulator.

Programmed cell death (PCD) is an essential process that functions in plant organ sculpture, tissue differentiation, nutrient recycling, and defense against pathogen attack. A full understanding of the mechanism of PCD in plants is hindered by the limited identification of protein components of the complex signaling circuitry that underpins this important physiological process. Here we have used Arabidopsis thaliana and fumonisin B1 (FB1) to identify proteins that constitute part of the PCD signaling network.

Proteomics reveals new insights into the role of light in cadmium response in Arabidopsis cell suspension cultures.

Light plays an important role in plant growth, development, and response to environmental stresses. To investigate the effects of light on the plant responses to cadmium (Cd) stress, we performed a comparative physiological and proteomic analysis of light- and dark-grown Arabidopsis cells after exposure to Cd. Treatment with different concentrations of Cd resulted in stress-related phenotypes such as cell growth inhibition and decline of cell viability.

Metabolic control analysis of developing oilseed rape (Brassica napus cv Westar) embryos shows that lipid assembly exerts significant control over oil accumulation.

Metabolic control analysis allows the study of metabolic regulation. We applied both single- and double-manipulation top-down control analysis to examine the control of lipid accumulation in developing oilseed rape (Brassica napus) embryos. The biosynthetic pathway was conceptually divided into two blocks of reactions (fatty acid biosynthesis (Block A), lipid assembly (Block B)) connected by a single system intermediate, the acyl-coenzyme A (acyl-CoA) pool.

Proteomic analysis of dark response in Arabidopsis cell suspension cultures.

Despite intense research on light responses in plants, the consequences of a simple shift from light to darkness are largely unexplored. In this research, the physiological outcome and proteomic changes in Arabidopsis cell suspension cultures after switching from light to total darkness were examined. Deprivation of light led to a visible loss of chlorophyll and failure to develop functional chloroplasts that are present in light-grown cells.

Tissue-specific whole transcriptome sequencing in castor, directed at understanding triacylglycerol lipid biosynthetic pathways.

Background: Storage triacylglycerols in castor bean seeds are enriched in the hydroxylated fatty acid ricinoleate. Extensive tissue-specific RNA-Seq transcriptome and lipid analysis will help identify components important for its biosynthesis.

Methodology/findings: Storage triacylglycerols (TAGs) in the endosperm of developing castor (Ricinus communis) seeds are highly enriched in ricinoleic acid (18:1-OH).

Plant extracellular ATP signalling: new insight from proteomics.

Complex signalling systems have evolved in multicellular organisms to enable cell-to-cell communication during growth and development. In plants, cell communication via the extracellular matrix (apoplast) controls many processes vital for plant survival. Secretion of ATP into the extracellular matrix is now recognised as a previously unknown stimulus for cell signalling with a role in many aspects of plant physiology.

Proteomics reveals a role for the RNA helicase crhR in the modulation of multiple metabolic pathways during cold acclimation of Synechocystis sp. PCC6803.

One of the earliest and largest transcriptional responses that occur during exposure of Synechocystis sp. PCC6803 to cold is the induction of the crhR RNA helicase transcript. We show that crhR deletion results in failure to cold acclimate: there is reduced growth at 24 °C and marked impairment of growth at 20 °C.

Components of complex lipid biosynthetic pathways in developing castor (Ricinus communis) seeds identified by MudPIT analysis of enriched endoplasmic reticulum.

Ricinoleic acid is a feedstock for nylon-11 (N11) synthesis which is currently obtained from castor (Ricinus communis) oil. Production of this fatty acid in a temperate oilseed crop is of great commercial interest, but the highest reported level in transgenic plant oils is 30%, below the 90% observed in castor and insufficient for commercial exploitation. To identify castor oil-biosynthetic enzymes and inform strategies to improve ricinoleic acid yields, we performed MudPIT analysis on endoplasmic reticulum (ER) purified from developing castor bean endosperm.

A phosphatidate phosphatase double mutant provides a new insight into plant membrane lipid homeostasis.

Phospholipids make up the bulk of most eukaryotic cell membranes, but how their synthesis is regulated remains relatively poorly understood in plants. In our article1 we provide evidence that two Mg ( 2+) -dependent phosphatidic acid phosphatase enzymes, called PAH1 and PAH2, are capable of repressing phospholipid biosynthesis at the endoplasmic reticulum in Arabidopsis thaliana. The precise mechanism of repression remains unclear and it does appear to vary in several respects from that already described in Saccharomyces cerevisiae.

Proteomic analysis of extracellular ATP-regulated proteins identifies ATP synthase beta-subunit as a novel plant cell death regulator.

Extracellular ATP is an important signal molecule required to cue plant growth and developmental programs, interactions with other organisms, and responses to environmental stimuli. The molecular targets mediating the physiological effects of extracellular ATP in plants have not yet been identified. We developed a well characterized experimental system that depletes Arabidopsis cell suspension culture extracellular ATP via treatment with the cell death-inducing mycotoxin fumonisin B1.

Phosphatidic acid phosphohydrolase 1 and 2 regulate phospholipid synthesis at the endoplasmic reticulum in Arabidopsis.

Phospholipid biosynthesis is essential for the construction of most eukaryotic cell membranes, but how this process is regulated in plants remains poorly understood. Here, we show that in Arabidopsis thaliana, two Mg(2+)-dependent phosphatidic acid phosphohydrolases called PAH1 and PAH2 act redundantly to repress phospholipid biosynthesis at the endoplasmic reticulum (ER). Leaves from pah1 pah2 double mutants contain ~1.

Identification of components associated with thermal acclimation of photosystem II in Synechocystis sp. PCC6803.

Background: Photosystem II (PSII) is the most thermally sensitive component of photosynthesis. Thermal acclimation of this complex activity is likely to be critically important to the ability of photosynthetic organisms to tolerate temperature changes in the environment.

Methodology/findings: We have analysed gene expression using whole-genome microarrays and monitored alterations in physiology during acclimation of PSII to elevated growth temperature in Synechocystis sp.

Differential proteomic analysis using iTRAQ reveals changes in thylakoids associated with Photosystem II-acquired thermotolerance in Synechocystis sp. PCC 6803.

Growth temperature has a marked influence on the thermotolerance of photosystem II (PSII), which is the most heat-sensitive component of photosynthesis. Using Synechocystis sp. PCC 6803 we have established that thylakoids isolated from cells grown at 38 degrees C have a greater degree of thermotolerance than those isolated from cells grown at 25 degrees C.

Synthesis of fatty acids de novo is required for photosynthetic acclimation of Synechocystis sp. PCC 6803 to high temperature.

The role of fatty acid synthesis in the acclimation of the photosynthetic machinery to high temperature was investigated in a mutant of the cyanobacterium Synechocystis sp. PCC 6803 that had a lower than wild-type level of enoyl-(acyl-carrier-protein) reductase FabI, a key component of the type-II fatty acid synthase system. The mutant exhibited marked impairment in the tolerance and acclimation of cells to high temperature: photoautotrophic growth of the mutant was severely inhibited at 40 degrees C.

The effects of extracellular adenosine 5'-triphosphate on the tobacco proteome.

Extracellular adenosine 5'-triphosphate (eATP) is emerging as an important plant signalling compound capable of mobilising intracellular second messengers such as Ca(2+), nitric oxide, and reactive oxygen species. However, the downstream molecular targets and the spectrum of physiological processes that eATP regulates are largely unknown. We used exogenous ATP and a non-hydrolysable analogue as probes to identify the molecular and physiological effects of eATP-mediated signalling in tobacco.

Extracellular ATP is a regulator of pathogen defence in plants.

In healthy plants extracellular ATP (eATP) regulates the balance between cell viability and death. Here we show an unexpected critical regulatory role of eATP in disease resistance and defensive signalling. In tobacco, enzymatic depletion of eATP or competition with non-hydrolysable ATP analogues induced pathogenesis-related (PR) gene expression and enhanced resistance to tobacco mosaic virus and Pseudomonas syringae pv.

Plant extracellular ATP signalling by plasma membrane NADPH oxidase and Ca2+ channels.

Extracellular ATP regulates higher plant growth and adaptation. The signalling events may be unique to higher plants, as they lack animal purinoceptor homologues. Although it is known that plant cytosolic free Ca2+ can be elevated by extracellular ATP, the mechanism is unknown.

Proof of function of a putative 3-hydroxyacyl-acyl carrier protein dehydratase from higher plants by mass spectrometry of product formation.

The predicted mature portion of a putative 3-hydroxyacyl-ACP dehydratase (DH) from Arabidopsis was linked to an N-terminal poly-histidine-tag and the fusion protein expressed in Escherichia coli. Soluble dehydratase was present on induction at 25 degrees C and pure dehydratase eluted from a nickel-affinity column in 0.2-0.

Isolation and fractionation of the endoplasmic reticulum from castor bean (Ricinus communis) endosperm for proteomic analyses.

This chapter describes the preparation and isolation of highly purified endoplasmic reticulum (ER) from the endosperm of developing and germinating castor bean (Ricinus communis) seeds to provide a purified organelle fraction for differential proteomic analyses. The method uses a two-step ultracentrifugation protocol first described by Coughlan (1) and uses sucrose density gradients and a sucrose flotation step to yield purified ER devoid of other contaminating endomembrane material. Using a combination of one dimensional (1D) and two dimensional (2D) gel electrophoresis the complexity and reproducibility of the protein profile of the purified organelle is evaluated prior to detailed proteomic analyses using mass spectrometry based techniques.

Differential proteomic analysis of the endoplasmic reticulum from developing and germinating seeds of castor (Ricinus communis) identifies seed protein precursors as significant components of the endoplasmic reticulum.

The endoplasmic reticulum is a major compartment of storage protein and lipid biosynthesis. Maximal synthesis of these storage compounds occurs during seed development with breakdown occurring during germination. In this study, we have isolated four independent preparations of ER from both developing and germinating seeds of castor bean (Ricinus communis) and used 2-D DIGE, and a combination of PMF and MS/MS sequencing, to quantify and identify differences in protein complement at both stages.

Antisense expression of 3-oxoacyl-ACP reductase affects whole plant productivity and causes collateral changes in activity of fatty acid synthase components.

Brassica napus cv Westar plants were transformed with 3-oxoacyl-ACP reductase (KR) in antisense orientation, driven by either the cauliflower mosaic virus 35S promoter or a seed-specific acyl carrier protein promoter to determine the effects on plant productivity and on the activity of other fatty acid synthase (FAS) components. In plants with altered KR activity, total seed yield was reduced in all cases. In less severely affected plant lines, seeds had a normal appearance and composition but the yield of seeds was reduced by approximately 50%.