Genetic diversity of wild population of Pyropia haitanensis based on SSR analysis

Genetic diversity analysis was conducted on 80 individuals of 4 populations of non-bred Pyropia haitanensis by simple sequence repeat (SSR) method. Using 15 pairs of microsatellite primers, 37 polymorphic loci were amplified, representing 94.9% of all loci. At the population level, the percentage of polymorphic bands (P) and polymorphism information content (PIC) were 66.67–84.62% and 0.481–0.488, with average value at 73.72% and 0.483, respectively. Expected heterozygosity (H_e) and Shannon's information index (I) were 0.279 and 0.434, respectively, at the species level, and 0.233 and 0.356 at population level. According to the coefficient of gene differentiation (G_ST), a large proportion of genetic variance (83.6%) of P. haitanensis was among individuals within populations, only 16.4% genetic variance was among populations, which was identified with the moderate gene flow value (N_m = 2.542). UPGMA clustered the 4 populations into 3 groups, and no significant correlation was found between the genetic distance and the corresponding geographic distance among the populations.     

Profiles of glycerolipids in Pyropia haitanensis and their changes responding to agaro-oligosaccharides

A sensitive method based on electrospray ionization tandem mass spectrometry was used to profile glycerolipids in Pyropia haitanensis and their changes responding to agaro-oligosaccharides. Ten monogalactosyldiacylglycerols (MGDGs), twelve digalactosyldiacylglycerols (DGDGs), five sulfoquinovosyldiacylglycerols (SQDGs), five phosphatidylglycerols (PGs), fifteen phosphatidylcholins (PCs), three phosphatidic acids (PAs), and three phosphatidylethanolamines (PEs) were identified in P. haitanensis. We found the SQDG was the most abundant species, followed by MGDG, DGDG, PG, PC, PE, and PA of the total glycerolipids. The predominant lipid species contained C20 fatty acids at sn-1/sn-2 positions, which was different from higher plants. Changes in membrane lipid species occurred when P. haitanensis were treated with agaro-oligosaccharides. At first, agaro-oligosaccharides induced an increase in total glycerolipids including the galactolipids such as MGDG (20:5/20:5) and phospholipids such as PC (18:3/20:5), suggesting that agaro-oligosaccharides caused changes of lipids in chloroplasts and plasma membrane. With increased treatment time, a large decline in major plasma membrane lipids (PCs and PEs) was observed, but not galactolipids (MGDGs and DGDGs), suggesting that the lipid changes occurred mainly at the plasma membrane after prolonged treatment.     

Proteomic analysis of the oxidative stress response in Candida albicans

An efficient oxidative stress response (OSR) is important for the facultative pathogenic yeast Candida albicans to survive within the human host. We used a large scale 2-D protein gel electrophoresis approach to analyze the stress response mechanisms of C. albicans after treatment with hydrogen peroxide and the thiol oxidizing agent, diamide. Quantitation of in vivo protein synthesis after pulse labeling of the proteins with radioactive L-[35S]-methionine resulted in characteristic proteome signatures for hydrogen peroxide and diamide with significant overlap of 21 up-regulated proteins for both stressors. Among the induced proteins were enzymes with known antioxidant functions like catalase or thioredoxin reductase and a set of oxidoreductases. 2-D gel analysis of mutants in the CAP1 gene revealed that the synthesis of 12 proteins is controlled by the oxidative stress regulator Cap1p. Stressing its importance for the C. albicans OSR, all 12 proteins were also induced after oxidative challenge by hydrogen peroxide or diamide.     

Proteomic analysis of response to long-term continuous stress in roots of germinating soybean seeds

Germination is a complex process, highly dependent on various environmental factors, including temperature and water availability. Germinating soybean seeds are especially vulnerable to unfavorable environmental conditions and exposure to long-term abiotic stresses may result in diminishing much of the yield and most importantly – restrained germination. In the present study, a proteomic approach was employed to analyze influence of cold and osmotic stress on roots of germinated soybean (Glycine max, L.) seeds. Seeds were germinating under continuous conditions of cold stress (+10 °C/H₂O), osmotic stress (+25 °C/−0.2 MPa) as well as cold and osmotic stress combined (+10 °C/−0.2 MPa). Proteome maps established for control samples and stress-treated samples displayed 1272 CBB-stained spots. A total of 59 proteins, present in both control and stress-treated samples and showing significant differences in volume, were identified with LC/nanoESI-MS. Identified proteins divided into functional categories, revealed 9 proteins involved in plant defense, 8 proteins responsible for plant destination and storage and 10 proteins involved in various tracks of carbohydrate metabolism. Furthermore, a number of proteins were assigned to electron transport, range of metabolic pathways, secondary metabolism, protein synthesis, embryogenesis and development, signal transduction, cellular transport, translocation and storage. By analyzing differences in expression patterns, it was possible to trace the soybean response to long-term abiotic stress as well as to distinguish similarities and differences between response to cold and osmotic stress.     

Proteomic analysis of the response of Arabidopsis chloroplast proteins to high light stress

Light is an essential environmental factor in the progression of plant growth and development but prolonged exposure to high levels of light stress can cause cellular damage and ultimately result in the death of the plant. Plants can respond defensively to this stress for a limited period and this involves changes to their gene expression profiles. Proteomic approaches were therefore applied to the study of the response to high light stress in the Arabidopsis thaliana plant species. Wild-type Arabidopsis was grown under normal light (100 micromol photons.m(-2).s(-1)) conditions and then subjected to high light (1000 micromol photons.m(-2).s(-1)) stress. Chloroplasts were then isolated from these plants and both soluble and insoluble proteins were extracted and subjected to two-dimensional (2-D) gel electrophoresis. The resolved proteins were subsequently identified by matrix assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS) and comparative database analysis. 64 protein spots, which were identified as candidate factors that responded to high light stress, were then selected for analysis and 52 of these were successfully identified using MALDI-TOF-MS analysis. 35 of the 52 identified proteins were found to decrease their expression levels during high light stress and a further 14 of the candidate proteins had upregulated expression levels under these conditions. Most of the proteins that were downregulated during high light stress are involved in photosynthesis pathways. However, many of the 14 upregulated proteins were identified as previously well-known high light stress-related proteins, such as heat shock proteins (HSPs), dehydroascorbate reductase (DHAR), and superoxide dismutase (SOD). Three novel proteins that were more highly expressed during periods of high light stress but had no clear functional relationship to these conditions, were also identified in this study.     

Growth densities regulate the response to elevated CO2 in a farmed seaweed Pyropia haitanensis (Bangiales,Rhodophyta)

Journal of Applied Phycology - Atmospheric CO2 concentration is predicted to nearly double by the end of this century. There are a large number of reports on the effects of elevated atmospheric CO2...     

Response of Pyropia haitanensis to agaro-oligosaccharides evidenced mainly by the activation of the eicosanoid pathway

The marine red alga Pyropia haitanensis (Protoflorideophyceae, Bangiaceae) has a nonvascular and multicellular structure and emerged earlier in evolution than other cultivatable red algae. It has been reported that lipid mediators from both the eicosanoid and octadecanoid pathways are involved in the innate immunity of other marine algae. But the defense strategies of P. haitanensis are not clearly understood. Here, we investigated the lipid defense of P. haitanensis elicited by agaro-oligosaccharides. The results indicate that the resistance of P. haitanensis was elicited and hydrogen peroxide was released by agaro-oligosaccharides. In P. haitanensis, C20 fatty acids are the essential fatty acids. Phospholipase A₂ was activated, and the free fatty acids decreased 3 h after treatment with agaro-oligosaccharides. Gas chromatography–mass spectrometry analyses revealed that the contents of volatile organic compounds increased after treatment for 3 h, which indicated that these free fatty acids were metabolized to volatile organic compounds. In conclusion, the lipid metabolic defense pathway of P. haitanensis was mainly via the C20 metabolism pathway. The C20 fatty acid was rapidly metabolized to volatile organic compounds, but not oxidized to oxylipins in response to agaro-oligosaccharides.     

Proteomic changes in response to low-light stress during cotton fiber elongation

Numerous studies have illustrated that low light is one of the major abiotic stresses limiting cotton (Gossypium hirsutum L.) fiber length, but studies addressing molecular mechanisms contributing to reduced fiber growth under low light are lacking. To investigate the molecular mechanisms of cotton fiber elongation in response to low light, an experiment of low light caused by shading was conducted with cotton cultivar NuCOTN 33B. The results showed that low light resulted in shorter fiber length. Proteomic analysis of four developmental stages (5, 10, 15 and 20 days post-anthesis) showed that 49 proteins were significantly responsive to low light. 39 differentially expressed proteins that included some known as well as some novel low-light stress-responsive proteins were identified. These differentially expressed proteins were involved in signal transduction, carbohydrate/energy metabolism, cell wall component synthesis, protein metabolism, cytoskeleton, nitrogen metabolism and stress responses. The results also showed that the decrease in fiber length might be because the levels of signal-related protein (phospholipase D), cytoskeletal proteins (two annexins isoforms), cell wall component-related proteins (sucrose synthase, UDP-d-glucuronic acid 4-epimerase and rhamnose synthase), carbohydrate metabolism-proteins (phosphofructokinase, dihydrolipoamide dehydrogenase, vacuolar H⁺-ATPase catalytic subunit, malate dehydrogenase and isocitrate dehydrogenase), and stress-related proteins (peroxisomal catalase, short chain alcohol dehydrogenase) were decreased under low light.     

Proteomic analyses of the response of cyanobacteria to different stress conditions

Cyanobacteria are significant contributors to global photosynthetic productivity, thus making it relevant to study how the different environmental stresses can alter their physiological activities. Here, we review the current research work on the response of cyanobacteria to different kinds of stress, mainly focusing on their response to metal stress as studied by using the modern proteomic tools. We also report a proteomic analysis of plastocyanin and cytochrome c₆ deletion mutants of the cyanobacterium Synechocystis sp. PCC 6803 grown under copper or iron deprivation, as compared to wild-type cells, so as to get a further understanding of the metal homeostasis in cyanobacteria and their response to changing environmental conditions.     

Proteomic analysis of cellular response to osmotic stress in thick ascending limb of Henle's loop (TALH) cells

Epithelial cells of the thick ascending limb of Henle's loop (TALH cells) play a major role in the urinary concentrating mechanism. They are normally exposed to variable and often very high osmotic stress, which is particularly due to high sodium and chloride reabsorption and very low water permeability of the luminal membrane. It is already established that elevation of the activity of aldose reductase and hence an increase in intracellular sorbitol are indispensable for the osmotic adaptation and stability of the TALH cells. To identify new molecular factors potentially associated with the osmotic stress-resistant phenotype in kidney cells, TALH cells exhibiting low or high levels of resistance to osmotic stress were characterized using proteomic tools. Two-dimensional gel analysis showed a total number of 40 proteins that were differentially expressed in TALH cells under osmotic stress. Twenty-five proteins were overexpressed, whereas 15 proteins showed a down-regulation. Besides the sorbitol pathway enzyme aldose reductase, whose expression was 15 times increased, many other metabolic enzymes like glutathione S-transferase, malate dehydrogenase, lactate dehydrogenase, alpha enolase, glyceraldehyde-3-phosphate dehydrogenase, and triose-phosphate isomerase were up-regulated. Among the cytoskeleton proteins and cytoskeleton-associated proteins vimentin, cytokeratin, tropomyosin 4, and annexins I, II, and V were up-regulated, whereas tubulin and tropomyosins 1, 2, and 3 were down-regulated. The heat shock proteins alpha-crystallin chain B, HSP70, and HSP90 were found to be overexpressed. In contrast to the results in oxidative stress the endoplasmic reticulum stress proteins like glucose-regulated proteins (GRP78, GRP94, and GRP96), calreticulin, and protein-disulfide isomerase were down-regulated under hypertonic stress.     

Different Responses to Heat Shock Stress Revealed Heteromorphic Adaptation Strategy of Pyropia haitanensis (Bangiales,Rhodophyta)

Pyropia has a unique heteromorphic life cycle with alternation stages between thallus and conchocelis, which lives at different water temperatures in different seasons. To better understand the different adaptation strategies for temperature stress, we tried to observe comparative biochemical changes of Pyropia haitanensis based on a short term heat shock model. The results showed that: (1) At normal temperature, free-living conchocelis contains significantly higher levels of H₂O₂, fatty acid-derived volatiles, the copy number of Phrboh and Phhsp70 genes,the activities of NADPH oxidase and floridoside than those in thallus. The released H₂O₂ and NADPH oxidase activity of conchocelis were more than 7 times higher than those of thallus. The copy number of Phrboh in conchocelis was 32 times that in thallus. (2) After experiencing heat shock at 35°C for 30 min, the H₂O₂ contents, the mRNA levels of Phrboh and Phhsp70, NADPH oxidase activity and the floridoside content in thallus were all significantly increased. The mRNA levels of Phrboh increased 5.78 times in 5 min, NADPH oxidase activity increased 8.45 times in 20 min. (3) Whereas, in conchocelis, the changes in fatty acids and their down-stream volatiles predominated, significantly increasing levels of saturated fatty acids and decreasing levels of polyunsaturated fatty acids occurred, and the 8-carbon volatiles were accumulated. However, the changes in H₂O₂ content and expression of oxidant-related genes and enzymatic activity were not obvious. Overall, these results indicate that conchocelis maintains a high level of active protective apparatus to endure its survival at high temperature, while thallus exhibit typical stress responses to heat shock. It is concluded that Pyropia haitanensis has evolved a delicate strategy for temperature adaptation for its heteromorphic life cycle.     

Proteomic analysis of rice endosperm cells in response to expression of hGM-CSF

The accumulation of significant levels of transgenic products in plant cells is required not only for crop improvement, but also for molecular pharming. However, knowledge about the fate of transgenic products and endogenous proteins in grain cells is lacking. Here, we utilized a quantitative mass spectrometry-based proteomic approach for comparative analysis of expression profiles of transgenic rice endosperm cells in response to expression of a recombinant pharmaceutical protein, human granulocyte-macrophage colony stimulation factor (hGM-CSF). This study provided the first available evidence concerning the fate of exogenous and endogenous proteins in grain cells. Among 1883 identified proteins with a false positive rate of 5%, 103 displayed significant changes (p-value < 0.05) between the transgenic and the wild-type endosperm cells. Notably, endogenous storage proteins and most carbohydrate metabolism-related proteins were down-regulated, while 26S proteasome-related proteins and chaperones were up-regulated in the transgenic rice endosperm. Furthermore, it was observed that expression of hGM-CSF induced endoplasmic reticulum stress and activated the ubiquitin/26S-proteasome pathway, which led to ubiquitination of this foreign gene product in the transgenic rice endosperm.     

Proteomic analysis of cellular response to microcystin in human amnion FL cells

Microcystins (MC), the potent inhibitor of protein phosphatase 1 and 2A, are hepatotoxins of increasing importance due to its high acute toxicity and potent tumor promoting activity. So far, the exact mechanisms of MC-induced hepatotoxicity and tumor promoting activity have not been fully elucidated. To better understand the mechanisms underlying microcystin-RR (MC-RR) induced toxicity as well as provide the possibility for the establishment of biomarkers for MC-RR exposure, differential proteome analysis on human amnion FL cells treated by MC-RR was carried out using two-dimensional gel electrophoresis (2-DE) followed by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry. Image analysis of silver-stained 2-dimensional gels revealed that 89 proteins showed significant differential expression in MC-RR treated cells compared with control, and 8 proteins were unique to MC-RR treated cells and 8 proteins were only detected in control cells. Sixty-six proteins were further identified with high confidence by peptide mass fingerprinting. Some of the identified differentially expressed proteins have clearly relationship with the process of apoptosis, signal transduction, and cytoskeleton alteration which are consistent with the literature. The functional implications of alterations in the levels of these proteins were discussed. However, most of which have not been reported previously to be involved in cellular processes responded to MC-RR. Therefore, this work will provide new insight into the mechanism of MC-RR toxicity.