Comparative proteomic analysis provides new insights into the fiber elongating process in cotton

A comparative proteomic analysis was performed to explore the mechanism of cell elongation in developing cotton fibers. The temporal changes of global proteomes at five representative development stages (5-25 days post-anthesis [dpa]) were examined using 2-D electrophoresis. Among approximately 1800 stained protein spots reproducibly detected on each gel, 235 spots were differentially expressed with significant dynamics in elongating fibers. Of these, 120 spots showed a more than 2-fold change in at least one stage point, and 21 spots appeared to be specific to developmental stages. Furthermore, 106 differentially expressed proteins were identified from mass spectrometry to match 66 unique protein species. These proteins involve different cellular and metabolic processes with obvious functional tendencies toward energy/carbohydrate metabolism, protein turnover, cytoskeleton dynamics, cellular responses and redox homeostasis, indicating a good correlation between development-dependent proteins and fiber biochemical processes, as well as morphogenesis. Newly identified proteins such as phospholipase D alpha, vf14-3-3 protein, small ras-related protein, and GDP dissociation inhibitor will advance our knowledge of the complicated regulatory network. Identification of these proteins, combined with their changes in abundance, provides a global view of the development-dependent protein changes in cotton fibers, and offers a framework for further functional research of target proteins associated with fiber development.     

A proteomic approach to identify developmentally regulated proteins in Leishmania infantum

We used comparative two-dimensional gel electrophoresis (2-DE) and mass spectrometry methodologies to highlight and identify proteins that are differentially expressed in the intracellular stage of the parasite Leishmania donovani infantum, a causative agent of visceral leishmaniasis. During its digenetic life cycle, Leishmania alternates between the alimentary tract of the sandfly vector as an extracellular promastigote and the acidic phagolysosomes of macrophage cells as an intracellular amastigote. Proteins differentially expressed in the intracellular form of the parasite are thought to be important for intracellular survival and pathogenesis. We used narrow pH range strips for isoelectric focusing to resolve soluble proteins of both developmental stages of L. infantum. More than 62 proteins differentially expressed in amastigotes were detected among approximately 2000 protein spots resolved by 2-DE. A quadrupole time-of-flight analysis of few selected protein spots, specifically expressed in the amastigote stage, permitted the identification of two proteins, part of the energetic metabolism pathways, the isocitrate dehydrogenase and the glycolytic enzyme triosephosphate isomerase. The kinetic parameters of these two enzymes were measured in both developmental stages of the parasite and their activity was indeed found to be higher in amastigotes. These findings bring a new insight in our understanding of metabolic and energy requirements of the intracellular form of Leishmania. Comparative analysis of the proteome of both developmental stages of the protozoan parasite Leishmania should permit the identification of protein candidates for the development of vaccines and new drugs.     

Comparative proteomics analysis reveals an intimate protein network provoked by hydrogen peroxide stress in rice seedling leaves

Hydrogen peroxide (H2O2) plays a dual role in plants as the toxic by-product of normal cell metabolism and as a regulatory molecule in stress perception and signal transduction. However, a clear inventory as to how this dual function is regulated in plants is far from complete. In particular, how plants maintain survival under oxidative stress via adjustments of the intercellular metabolic network and antioxidative system is largely unknown. To investigate the responses of rice seedlings to H2O2 stress, changes in protein expression were analyzed using a comparative proteomics approach. Treatments with different concentrations of H2O2 for 6 h on 12-day-old rice seedlings resulted in several stressful phenotypes such as rolling leaves, decreased photosynthetic and photorespiratory rates, and elevated H2O2 accumulation. Analysis of approximately 2000 protein spots on each two-dimensional electrophoresis gel revealed 144 differentially expressed proteins. Of them, 65 protein spots were up-regulated, and 79 were down-regulated under at least one of the H2O2 treatment concentrations. Furthermore 129 differentially expressed protein spots were identified by mass spectrometry to match 89 diverse protein species. These identified proteins are involved in different cellular responses and metabolic processes with obvious functional tendencies toward cell defense, redox homeostasis, signal transduction, protein synthesis and degradation, photosynthesis and photorespiration, and carbohydrate/energy metabolism, indicating a good correlation between oxidative stress-responsive proteins and leaf physiological changes. The abundance changes of these proteins, together with their putative functions and participation in physiological reactions, produce an oxidative stress-responsive network at the protein level in H2O2-treated rice seedling leaves. Such a protein network allows us to further understand the possible management strategy of cellular activities occurring in the H2O2-treated rice seedling leaves and provides new insights into oxidative stress responses in plants.     

Combined effect of gender and caloric restriction on liver proteomic expression profile

We analyzed the combined effect of gender and CR on protein expression profile in liver. We identified 27 differentially expressed proteins involved in several cellular functions such as substrate metabolism, antioxidant systems, stress response, iron homeostasis and cardiovascular protection. This study reveals new cellular pathways liable to be similarly regulated in females and calorie restricted rats and which could be related with the greater longevity in these animals.     

Comparative proteome analysis of alkaliphilic <Emphasis Type="Italic">Bacillus</Emphasis> sp. N16-5 grown on different carbon sources

To determine the impact of carbohydrates on the metabolic pathway in alkaliphiles, proteomes were obtained from cultures containing different carbohydrates and were resolved on two-dimensional gel electrophoresis (2-DE). The proteomes were compared to determine differentially expressed proteins. A novel alkaliphilic bacterium (alkaliphilic Bacillus sp. N16-5 isolated from Wudunur Soda Lake, China) was isolated in media with five different carbon sources (glucose, mannose, galactose, arabinose, and xylose). Comparative proteome analysis identified 61 differentially expressed proteins, which were mainly involved in carbohydrate metabolism, amino acid transport, and metabolism, as well as energy production and conversion. The comparison was based on the draft genome sequence of strain N16-5. The abundance of enzymes involved in central metabolism was significantly changed when exposed to various carbohydrates. Notably, catabolite control protein A (CcpA) was up-regulated under all carbon sources compared with glucose. In addition, pentose exhibited a stronger effect than hexose in CcpA-mediated carbon catabolite repression. These results provided a fundamental understanding of carbohydrate metabolism in alkaliphiles.     

Quantitative Proteomic Analysis Reveals That Anti-Cancer Effects of Selenium-Binding Protein 1 In Vivo Are Associated with Metabolic Pathways

Previous studies have shown the tumor-suppressive role of selenium-binding protein 1 (SBP1), but the underlying mechanisms are unclear. In this study, we found that induction of SBP1 showed significant inhibition of colorectal cancer cell growth and metastasis in mice. We further employed isobaric tags for relative and absolute quantitation (iTRAQ) to identify proteins that were involved in SBP1-mediated anti-cancer effects in tumor tissues. We identified 132 differentially expressed proteins, among them, 53 proteins were upregulated and 79 proteins were downregulated. Importantly, many of the differentially altered proteins were associated with lipid/glucose metabolism, which were also linked to Glycolysis, MAPK, Wnt, NF-kB, NOTCH and epithelial-mesenchymal transition (EMT) signaling pathways. These results have revealed a novel mechanism that SBP1-mediated cancer inhibition is through altering lipid/glucose metabolic signaling pathways.     

A high‐confidence reference dataset of differentially expressed proteins in elongating cotton fiber cells

In our previous study, we used a comparative proteomic approach based on 2DE to profile dynamic proteomes of cotton fibers and found 235 protein spots differentially expressed during the elongation process ranging from 5 to 25 days post‐anthesis. Of them, only 106 differentially expressed proteins (DEPs) were identified by MS due to database limitations at the time. In the present work, we successfully identified the remaining 129 DEPs from the same experimental system using high‐resolution MS with an updated database. Bioinformatic analysis revealed that proteins involved in carbohydrate and protein metabolism, transport, and redox homeostasis are the most abundant, and glycolysis was found to be the most significantly regulated process during fiber elongation. Our high‐confidence reference dataset, composed of 235 DEPs, provides a valuable resource for future studies on the molecular mechanism of cotton fiber elongation.     

Protein expression changes during cotton fiber elongation in response to drought stress and recovery

An investigation to better understand the molecular mechanism of cotton (Gossypium hirsutum L.) fiber elongation in response to drought stress and recovery was conducted using a comparative proteomics analysis. Cotton plants (cv. NuCOTN 33B) were subjected to water deprivation for 10 days followed by a recovery period (with watering) of 5 days. The temporal changes in total proteins in cotton fibers were examined using 2DE. The results revealed that 163 proteins are significantly drought responsive. MS analysis led to the identification of 132 differentially expressed proteins that include some known as well as some novel drought‐responsive proteins. These drought responsive fiber proteins in NuCOTN 33B are associated with a variety of cellular functions, i.e. signal transduction, protein processing, redox homeostasis, cell wall modification, metabolisms of carbon, energy, lipid, lignin, and flavonoid. The results suggest that the enhancement of the perception of drought stress, a new balance of the metabolism of the biosynthesis of cell wall components and cytoskeleton homeostasis plays an important role in the response of cotton fibers to drought stress. Overall, the current study provides an overview of the molecular mechanism of drought response in cotton fiber cells.     

Nano-LC-MS/MS based proteomics of hepatocellular carcinoma cells compared to Chang liver cells and tanshinone IIA induction

Human hepatocellular carcinoma (HCC) is one of the most malignant tumors, being particularly induced by unregulated growth and metastasis, and is a leading cause of death and major health problems in many countries. We report here the identification of 167 differentially expressed proteins between HCC (MHCC97-H) cells and Chang liver cells using enhanced nano-liquid chromatography/mass spectrometry (LC/MS). The most relevant pathways of differentially expressed proteins are involved in cytoskeleton organization, stress defense, and energy homeostasis etc. Moreover, of the identified proteins, there are 59 known or putative membrane-associated proteins with multitransmembrane domains confirmed by bioinformatic analysis. These proteins may be associated with cancer, reflecting tumorigenesis of HCC, and would be useful for the development of diagnostic and subsequently pharmaceutical targets of HCC. In addition, we identify a total of 41 proteins that are found to be up- or down-regulated following tanshinone IIA treatment for MHCC97H cells in a time-depended manner. Also, several proteins that are involved in actin cytoskeleton and stress resistance are mainly down-regulated, whereas proteins associated with cell redox homeostasis, mitochondrial, and microtubule-based movement are identified as mostly up-regulated after the treatment. Determination of functional roles of those differentially expressed proteins will enable further understanding of the mechanism of HCC tumorigenesis and exploration of new drugs for therapeutic intervention.     

A comparative proteomic analysis of Gluconacetobacter diazotrophicus PAL5 at exponential and stationary phases of cultures in the presence of high and low levels of inorganic nitrogen compound

A proteomic view of G. diazotrophicus PAL5 at the exponential (E) and stationary phases (S) of cultures in the presence of low (L) and high levels (H) of combined nitrogen is presented. The proteomes analyzed on 2D-gels showed 131 proteins (42E+32S+29H+28L) differentially expressed by G. diazotrophicus, from which 46 were identified by combining mass spectrometry and bioinformatics tools. Proteins related to cofactor, energy and DNA metabolisms and cytoplasmic pH homeostasis were differentially expressed in E growth phase, under L and H conditions, in line with the high metabolic rate of the cells and the low pH of the media. Proteins most abundant in S-phase cells were stress associated and transporters plus transferases in agreement with the general phenomenon that binding protein-dependent systems are induced under nutrient limitation as part of hunger response. Cells grown in L condition produced nitrogen-fixation accessory proteins with roles in biosynthesis and stabilization of the nitrogenase complex plus proteins for protection of the nitrogenases from O(2)-induced inactivation. Proteins of the cell wall biogenesis apparatus were also expressed under nitrogen limitation and might function in the reshaping of the nitrogen-fixing G. diazotrophicus cells previously described. Genes whose protein products were detected in our analysis were mapped onto the chromosome and, based on the tendency of functionally related bacterial genes to cluster, we identified genes of particular pathways that could be organized in operons and are co-regulated. These results showed the great potential of proteomics to describe events in G. diazotrophicus cells by looking at proteins expressed under distinct growth conditions.     

Proteomics Analysis of Soybean Seedlings under Short-Term Water Deficit

Soybeans are one of the most important grain crops worldwide. Water deficit, which seriously affects the yield and quality of soybeans, is the main abiotic stress factor in soybean production. As a follow-up study, the droughttolerant soybean variant Heinong 44 was analyzed via proteome analysis. Soybean was exposed to water deficit for 0, 8, and 24 h, and protein samples were extracted for detection of differentially expressed proteins. Protein sequencing of leaf tissues under water stress yielded a total of 549 differentially expressed proteins: 75 and 320 upregulated proteins as well as 70 and 84 downregulated proteins were obtained after 8 and 24 h of water deficit, respectively. Gene Ontology analysis revealed that most of the differentially expressed proteins (DEPs) were involved in catalytic activity, molecular function, and metabolic processes, whereas some of them were involved in photosynthesis, carbon metabolism, and energy metabolism. We also identified some differentially expressed proteins that may be involved in the regulation of water deficit response. Our study provides a theoretical basis for the breeding of drought-resistant soybean varieties.     

Genetic, proteomic and metabolic analysis of the regulation of energy storage in rice seedlings in response to drought

We used proteomic analysis to determine the response of rice plant seedlings to drought-induced stress. The expression of 71 protein spots was significantly altered, and 60 spots were successfully identified. The greatest down-regulated protein functional category was translation. Up-regulated proteins were mainly related to protein folding and assembly. Additionally, many proteins involved in metabolism (e.g. carbohydrate metabolism) also showed differences in expression. cDNA microarray and GC-MS analysis showed 4756 differentially expressed mRNAs and 37 differentially expressed metabolites. Once these data were integrated with the proteomic analysis, we were able to elucidate the metabolic pathways affected by drought-induced stress. These results suggest that increased energy consumption from storage substances occurred during drought. In addition, increased expression of the enzymes involved in anabolic pathways corresponded with an increase in the content of six amino acids. We speculated that energy conversion from carbohydrates and/or fatty acids to amino acids was increased. Analysis of basic metabolism networks allowed us to understand how rice plants adjust to drought conditions.     

双孢蘑菇子实体发育后期差异表达蛋白质分析

为探讨双孢蘑菇子实体发育后期的蛋白质表达变化,对双孢蘑菇As2796子实体采收期、成熟期和开伞期的蛋白质组进行了双向电泳(2-DE)分析,发现了16个表达差异明显的蛋白质。通过质谱分析(MALDI-TOF/TOF MS)和数据库检索,有14个差异蛋白质获得鉴定。其中磷酸烯醇式丙酮酸水合酶与能量代谢相关,T-蛋白复合体1、蛋白酶体、5-甲基四氢三谷氨酸-同型半胱氨酸甲基转移酶、1-吡咯琳-5-羧酸脱氢酶、精氨酸酶与氨基酸或蛋白质代谢直接相关,而GTP结合蛋白则参与细胞的多种生命活动,在细胞的生长发育过程中起着重要的作用。另外7个为功能未知的蛋白质。