Akiko Hashiguchi

Proteome analysis of early-stage soybean seedlings under flooding stress.

Proteomic analyses of soybean seedlings responding to flooding were conducted to identify proteins involved in such response. Soybean was germinated for 48 h and then subjected to flooding stress for 6-48 h. Proteomic analysis of hypocotyl and root was used in a time-dependent manner, and altered proteins were identified using soybean protein data file constructed for this research. Under flooding stress, 35 proteins were up-regulated, whereas 16 proteins were down-regulated at a 24-h time point. Changes in energy generation was recognized because several glycolytic enzymes were up-regulated. General stress response was also shown to occur as various reactive oxygen species scavengers were up-regulated. Other identified proteins with diverse functional categories suggest that flooding stress includes not only hypoxic stress, but also other stresses such as weak light, disease, and water stresses. In addition, proteins with unknown functions were shown to be positioned as hubs which activate other proteins in system response networks by protein-protein interaction analysis, suggesting that this type of interaction analysis is useful for screening of important factors in plant response to environmental stresses.

Role of TSC-22 during early embryogenesis in Xenopus laevis.

Transforming growth factor‐β1‐stimulated clone 22 (TSC‐22) encodes a leucine zipper‐containing protein that is highly conserved. During mouse embryogenesis, TSC‐22 is expressed at the site of epithelial–mesenchymal interaction. Here, we isolated Xenopus laevis TSC‐22 (XTSC‐22) and analyzed its function in early development. XTSC‐22 mRNA was first detected in the ectoderm of late blastulae. Translational knockdown using XTSC‐22 antisense morpholino oligonucleotides (XTSC‐22‐MO) caused a severe delay in blastopore closure in gastrulating embryos. This was not due to mesoderm induction or convergent‐extension, as confirmed by whole‐mount in situ hybridization and animal cap assay. Cell lineage tracing revealed that migration of ectoderm cells toward blastopore was disrupted in XTSC‐22‐depleted embryos, and these embryos had a marked increase in the number of dividing cells. In contrast, cell division was suppressed in XTSC‐22 mRNA‐injected embryos. Co‐injection of XTSC‐22‐MO and mRNA encoding p27Xic1, which inhibits cell cycle promotion by binding cyclin/Cdk complexes, reversed aberrant cell division. This was accompanied by rescue of the delay in blastopore closure and cell migration. These results indicate that XTSC‐22 is required for cell movement during gastrulation though cell cycle regulation.

Impact of Post-Translational Modifications of Crop Proteins under Abiotic Stress

The efficiency of stress-induced adaptive responses of plants depends on intricate coordination of multiple signal transduction pathways that act coordinately or, in some cases, antagonistically. Protein post-translational modifications (PTMs) can regulate protein activity and localization as well as protein–protein interactions in numerous cellular processes, thus leading to elaborate regulation of plant responses to various external stimuli. Understanding responses of crop plants under field conditions is crucial to design novel stress-tolerant cultivars that maintain robust homeostasis even under extreme conditions. In this review, proteomic studies of PTMs in crops are summarized. Although the research on the roles of crop PTMs in regulating stress response mechanisms is still in its early stage, several novel insights have been retrieved so far. This review covers techniques for detection of PTMs in plants, representative PTMs in plants under abiotic stress, and how PTMs control functions of representative proteins. In addition, because PTMs under abiotic stresses are well described in soybeans under submergence, recent findings in PTMs of soybean proteins under flooding stress are introduced. This review provides information on advances in PTM study in relation to plant adaptations to abiotic stresses, underlining the importance of PTM study to ensure adequate agricultural production in the future.

TSC-box is essential for the nuclear localization and antiproliferative effect of XTSC-22

Transforming growth factor‐β1‐stimulated clone 22 (TSC‐22) encodes a leucine zipper‐containing protein that is highly conserved among various species. Mammalian TSC‐22 is a potential tumor suppressor gene. It translocates into nuclei and suppresses cell division upon antiproliferative stimuli. In human colon carcinoma cells, TSC‐22 inhibits cell growth by upregulating expression of the p21 gene, a cyclin‐dependent kinase (Cdk) inhibitor. We previously showed that the Xenopus laevis homologue of the TSC‐22 gene (XTSC‐22) is required for cell movement during gastrulation through cell cycle regulation. In this report, we investigated the molecular mechanism of the antiproliferative effect of XTSC‐22. Reverse transcriptase‐polymerase chain reaction (RT‐PCR) analysis suggested that XTSC‐22 did not affect the expression levels of the p21 family of Cdk inhibitors or other cell cycle regulators. Analysis of deletion mutants of XTSC‐22 revealed that nuclear localization of the N‐terminal TSC‐box is necessary for cell cycle inhibition by XTSC‐22. Further experiments suggested that p27Xic1, a key Cdk inhibitor in Xenopus, interacts with XTSC‐22. Because p27Xic1 is a cell cycle inhibitor with a nuclear localization signal, it is possible that XTSC‐22 suppresses cell division by translocating into the nucleus with p27Xic1, where it may potentiate the intranuclear action of p27Xic1.

Analyses of the Proteomes of the Leaf, Hypocotyl, and Root of Young Soybean Seedlings

The functions of organs in young soybean seedling were determined by means of proteomic analysis. Extracts from leaves, hypocotyls, and roots were separated by two-dimensional polyacrylamide gel electrophoresis, and the proteins were identified by mass spectrometry and protein sequencing. The identified proteins were categorized into various groups according to their function. The leaf was abundant in proteins associated with energy production (50.0%), the hypocotyl was rich in defense proteins (31.8%), and the root contained defense-related proteins (16.7%) and destination and storage proteins (26.7%). Stem 31-kDa glycoprotein, 20 kDa chaperonin, 50S ribosomal protein, and trypsin inhibitor were common to all three tissues. The sequence information obtained from the soybean proteome should be helpful in predicting the functions of unknown proteins.

Proteomics and metabolomics-driven pathway reconstruction of mung bean for nutraceutical evaluation

Mung bean is a legume crop which has a various health-promoting effects. Although rich flavonoids are reported to be responsible for its biological activities, little is known about other nutrients that may potentiate the activities. To obtain information on mung bean nutritional properties, gel-free/label-free proteomic analysis and metabolomic profiling were combined. Pathway reconstruction detected that amino acid metabolism is more active in flesh. Coat contained wider variety of lipids and phenolic acids/flavonoids than flesh. Among the compounds detected in coat, sphingolipids, arachidonic acid, and prostaglandin E2 are compounds which are related to immune response induction. Furthermore, identification of prostaglandin F2α added scientific support to empirical validity of mung bean usage. The abundance of bioactive compounds such as naringenin, which can be metabolized into vitexin, varied among cultivars. These results suggest that lipids together with flavonoids might be potential responsible compounds for biological activity of mung bean coat and flesh.

Proteomic analysis of soybean seedling leaf under waterlogging stress in a time-dependent manner

Leaf is sensitive to environmental changes and exhibits specific responses to abiotic stress. To identify the response mechanism in soybean leaf under waterlogging stress, a gel-free/label-free proteomic technique combined with polyethylene glycol fractionation was used. Attenuated photosynthesis by waterlogging stress in the leaf of soybean seedlings was indicated from proteomic results. Defensive mechanisms such as reactive oxygen species (ROS) scavenging was also recognized. Cluster analysis revealed that proteins that exhibit characteristic dynamics in response to waterlogging were mainly related to photosynthesis. Among the identified photorespiration-related proteins, the protein abundance and enzyme activity of hydroxypyruvate reductase were transiently increased in control plants, but were clearly decreased in response to waterlogging stress. These results suggest that waterlogging directly impairs photosynthesis and photorespiration. Furthermore, hydroxypyruvate reductase may be a critical enzyme controlling the rate of photorespiration.

Subcellular Proteomics: Application to Elucidation of Flooding-Response Mechanisms in Soybean

Soybean, which is rich in protein and oil, is cultivated in several climatic zones; however, its growth is markedly decreased by flooding. Proteomics is a useful tool for understanding the flooding-response mechanism in soybean. Subcellular proteomics has the potential to elucidate localized cellular responses and investigate communications among subcellular components during plant growth and during stress. Under flooding, proteins related to signaling, stress and the antioxidative system are increased in the plasma membrane; scavenging enzymes for reactive-oxygen species are suppressed in the cell wall; protein translation is suppressed through inhibition of proteins related to preribosome biogenesis and mRNA processing in the nucleus; levels of proteins involved in the electron transport chain are reduced in the mitochondrion; and levels of proteins related to protein folding are decreased in the endoplasmic reticulum. This review discusses the advantages of a gel-free/label-free proteomic technique and methods of plant subcellular purification. It also summarizes cellular events in soybean under flooding and discusses future prospects for generation of flooding-tolerant soybean.

Differences in fennel seed responses to drought stress at the seed formation stage in sensitive and tolerant genotypes

To understand the effects of drought on fennel seed production and determine the underlying molecular processes, various fennel genotypes were exposed to drought stress. The yield and quality, including aromatic oil content, of fennel seeds were reduced by drought during seed development. To explore drought-induced biological processes in fennel, a label-free/gel-free proteomic analysis was performed. In Gaziantep and Tatmaj cultivars, which are sensitive and tolerant fennel genotypes, respectively, 106 and 92 drought-responsive proteins were identified. Comparison of protein-functional profiles indicated that proteins classified in stress, cell, and protein synthesis/degradation categories consisted important responsive mechanisms against drought stress. Pathway analysis visualized that the tricarboxylic acid cycle is important for both cultivars. In Tatmaj, moderate activation of proteins related to oxidative pentose phosphate pathway was detected along with an increase in photosynthesis-related proteins. Furthermore, cluster analysis of drought-responsive proteins using protein abundance at milky, dough, and mature stages identified protein homeostasis as a mechanism of drought tolerance in fennel. These results suggest that coordinated energy consumption and supply might be a drought-tolerance mechanism in fennel plants.

Proteomics of Soybean Plants

Abstract Soybean ( Glycine max L. Merr.) is one of the most important protein sources throughout the world. However, ever-changing environmental conditions and limited availability of arable land warrant the development of soybean cultivars with increased stress tolerance. Proteomic approaches have been used to identify target proteins responsible for important agronomic traits of soybean. The major challenge using this approach is the inherent complexity of interactions that occur between plants and the environment during different stages of plant development and in response to various environmental stresses. Proteomic analyses have provided significant insight into the molecular mechanisms underlying soybean responses to stress. In this chapter, the findings and applications of proteomics in soybean research are reviewed. In addition, proteomic descriptions of soybean development and growth are summarized, and plant–environment interactions, including symbiosis and soybean responses to abiotic stresses, particularly flooding stress, are described. Finally, the application of proteomic techniques for the evaluation of allergenic proteins and functional peptides in soybean is discussed.