Mass spectrometry-based analysis of proteomic changes in the root tips of flooded soybean seedlings

Flooding injury is a major problem in soybean cultivation. A proteomics approach was used to clarify the occurrence of changes in protein expression level and phosphorylation in soybeans under flooding stress. Two-day-old seedlings were flooded for 1 day, proteins were extracted from root tips of the seedlings and digested with trypsin, and their expression levels and phosphorylation states were compared to those of untreated controls using mass spectrometry-based proteomics techniques. Phosphoproteins were enriched using a phosphoprotein purification column prior to digestion and mass spectrometry. The expression of proteins involved in energy production increased as a result of flooding, while expression of proteins involved in protein folding and cell structure maintenance decreased. Flooding induced changes of phosphorylation status of proteins involved in energy generation, protein synthesis and cell structure maintenance. The response to flooding stress may be regulated by both modulation of protein expression and phosphorylation state. Energy-demanding and production-related metabolic pathways may be particularly subject to regulation by changes in protein phosphorylation during flooding.     

Search for immunobiological parameters predictive of clinical effects of OK-432 in patients with malignant ascites

Although OK-432, a potent BRM, has been known to induce the remarkable improvement of clinical conditions in cancer patients through its strong effects on their immune capabilities, no specific immune parameters have been identified to best predict the clinical outcome after the OK-432 treatment. In an attempt to identify early parameters indicative of the clinical effects, we have administered 0.1 mg of OK-432 intraperitoneally to a total of 12 patients with malignant ascites and examined peritoneal fluid and peripheral blood obtained on 4 days before, 1, 3, and 7 days after the OK-432 injection using various immunobiological assays. Four weeks later, clinical improvements were evaluated by the disappearance of malignant cells from and/or substantial decrease in ascites. Four patients (responders) showed the improvements while 8 patients (nonresponders) showed no clinical evidence for improvement. In a few parameters among the many examined, significantly different patterns of changes were noted between responders and nonresponders. Thus, in nonresponder patients MØ and T cell population returned to an initial low level after early increases (on days 1 and/or 3), while they remained increased day 1 through 7 in responders. In responder patients, the cytotoxicity of peritoneal mononuclear cells against K562 and Daudi cells were augmented on day 7, but not in nonresponder patients. Thein vitro stimulation of the mononuclear cells with OK-432 enhanced the cytotoxic activity and induced the interferon (IFN) production in the responders but not in nonresponders. These parameters will be useful for the early prediction of the expected clinical effects of OK-432.     

Proteomic characterization of tissue expansion of rice scutellum stimulated by abscisic acid

We found that appropriate treatment with a highly potent and long-lasting abscisic acid analog enhanced the tissue expansion of scutellum during early seedling development of rice, accompanied by increases of protein and starch accumulation in the tissue. A comparative display of the protein expression patterns in the abscisic acid analog-treated and non-treated tissues on two dimensional gel electrophoretogram indicated that approximately 30% of the scutellar proteins were induced by abscisic acid. The abscisic acid-induced proteins included sucrose metabolizing, glycolytic, and ATP-producing enzymes. Most of these enzyme proteins also increased during the seedling growth. In addition, the expression of some isoforms of UDP-glucose pyrophosphorylase, 3-phosphoglycerate kinase, and mitochondrial ATP synthase beta chain was stimulated in the scutellum, with suppressed expression of alpha-amylase. We concluded that abscisic acid directly and indirectly stimulates the expression of numerous proteins, including carbohydrate metabolic enzymes, in scutellar tissues.     

Symmetry and entropy of biological patterns: discrete Walsh functions for 2D image analysis

To quantify symmetry and entropy inherent in the discrete patterns such as spatial self-organization in cell sorting and mussel bed ecosystems, we introduce the discrete Walsh analysis. This analysis enables us to estimate the degree of the complicated symmetry, and to extract the symmetry from the pattern that seems to be asymmetric. The results obtained in this paper are summarized as follows. (I) The geometrical patterns of the cell sorting become systematic with the predominance of the particular symmetry. This implies that not only the entropy but also the particular symmetry can decrease in the biological process. (II) The magnitude of the symmetry is related to the absolute value of the adhesion, and the type of the symmetry is related to the sign of the adhesion. That is, centro-symmetry dominates in the cell sorting pattern caused by large negative adhesion, and double symmetry dominates in the pattern caused by large positive adhesion. (III) Spatial self-organization in mussel bed is accompanied by the decreasing of the centro-symmetry. This implies that the positive "adhesion" between mussel individuals increases with time. (IV) In the biological process, the Curie symmetry breaking occurs at intervals.     

Random PCR‐based genotyping by sequencing technology GRAS‐Di (genotyping by random amplicon sequencing,direct) reveals genetic structure of mangrove fishes

While various technologies for high‐throughput genotyping have been developed for ecological studies, simple methods tolerant to low‐quality DNA samples are still limited. In this study, we tested the availability of a random PCR‐based genotyping‐by‐sequencing technology, genotyping by random amplicon sequencing, direct (GRAS‐Di). We focused on population genetic analysis of estuarine mangrove fishes, including two resident species, the Amboina cardinalfish (Fibramia amboinensis, Bleeker, 1853) and the Duncker's river garfish (Zenarchopterus dunckeri, Mohr, 1926), and a marine migrant, the blacktail snapper (Lutjanus fulvus, Forster, 1801). Collections were from the Ryukyu Islands, southern Japan. PCR amplicons derived from ~130 individuals were pooled and sequenced in a single lane on a HiSeq2500 platform, and an average of three million reads was obtained per individual. Consensus contigs were assembled for each species and used for genotyping of single nucleotide polymorphisms by mapping trimmed reads onto the contigs. After quality filtering steps, 4,000–9,000 putative single nucleotide polymorphisms were detected for each species. Although DNA fragmentation can diminish genotyping performance when analysed on next‐generation sequencing technology, the effect was small. Genetic differentiation and a clear pattern of isolation‐by‐distance was observed in F. amboinensis and Z. dunckeri by means of principal component analysis, F_ST and the admixture analysis. By contrast, L. fulvus comprised a genetically homogeneous population with directional recent gene flow. These genetic differentiation patterns reflect patterns of estuary use through life history. These results showed the power of GRAS‐Di for fine‐grained genetic analysis using field samples, including mangrove fishes.     

Ozone stress‐induced proteomic changes in leaf total soluble and chloroplast proteins of soybean reveal that carbon allocation is involved in adaptation in the early developmental stage

Considerable soybean yield losses caused by ozone (O₃) stress have been demonstrated by large‐scale meta‐analyses of free‐gas concentration enrichment systems. In this study, comparative proteomic approach was employed to explore the differential changes of proteins in O₃ target structures such as leaf and chloroplasts of soybean seedlings. Acute O₃ exposure (120 parts‐per‐billion) for 3 days did not cause any visible symptoms in developing leaves. However, higher amounts of ROS and lipid peroxidation indicated that severe oxidative burst occurred. Immunoblot analysis of O₃‐induced known proteins revealed that proteins were modulated before symptoms became visible. Proteomic analysis identified a total of 20 and 32 differentially expressed proteins from O₃‐treated leaf and chloroplast, respectively. Proteins associated with photosynthesis, including photosystem I/II and carbon assimilation decreased following exposure to O₃. In contrast, proteins involved in antioxidant defense and carbon metabolism increased. The activity of enzymes involved in carbohydrate metabolism increased following exposure to O₃, which is consistent with the decrease in starch and increase in sucrose concentrations. Taken together, these results suggest that carbon allocation is tightly programmed, and starch degradation probably feeds the tricarboxylic acid cycle while the photosynthesis pathway is severely affected during O₃ stress.     

Properdin factor B frequencies in four Asian populations

The distribution of Properdin factor B (Bf) phenotypes and their gene frequencies were investigated in four Asian populations (Chinese, Filipino, Thai and Japanese). The frequency of the BfS phenotype in Filipinos (0.717) was significantly lower than that in Chinese (0.900) and Thai (0.889) (p less than 0.01), but not different from the Japanese (0.840). One variant, BfF 0.65 S, was identified in a Japanese subject. Thus, in the Asian populations studied, Bfs frequencies were high and the frequency of variants other than F and S were low.     

Ovarian development in Meishan pigs

Ovaries collected from a total of 35 Meishan gilts of various ages were morphologically and histologically examined. Vesicular follicles were first observed in ovarian sections at 45 d of age. Simultaneously, protruding follicles appeared on the surface of the ovaries, and then ovarian weight increased rapidly with an increasing number of protruding follicles. About half of the gilts between 75 and 90 d of age had ovaries with corpora lutea, indicating that some of them had started estrous cycles before reaching 75 d of age. In Meishan gilts treated with exogenous gonadotropins, ovaries were stimulated as early as 45 d of age. These results suggest that the precocity of Meishan gilts may include the development of vesicular follicles in the ovaries at an early age.