Network Structure and Forces Involved in Perilla Globulin Gelation: Comparison with Sesame Globulin

Scanning electron micrographs show that perilla globulin gel had a finer network structure than sesame α-globulin gel. The effects of various reagents on the gel formation and solubility of perilla and sesame gels were compared. The contribution of disulfide bonds to the formation and stability of perilla gel was greater than to sesame gel, despite having the same subunit structure.     

Subunit structure and functional properties of the predominant globulin of perilla (Perilla frutescens var. frutescens) seeds

Approximately 40% of defatted perilla seeds consists of proteins which are primarily composed of globulin (84%). The amino acid profile of perilla proteins demonstrated balanced amounts of all essential amino acids, except for lysine. The molecular mass of the predominant globulin was estimated to be 340 kDa by gel filtration. This globulin was separated into three intermediary subunits (54, 57 and 59 kDa) by SDS-PAGE. It is suggested from these results that the globulin exists as a hexamer. A treatment with 50 mM dithiothreitol enabled the intermediary subunits to be separated into three acidic subunits (31-34 kDa) and four basic subunits (23-25 kDa). It is interesting that this subunit structure is the same as that of sesame α-globulin, despite them coming from different families. Compared to sesame α-globulin, the heat-induced gel of perilla globulin had better water-holding ability, despite it displaying the same degree of gel hardness.     

Gas Chromatographic Analysis of Higher Alcohols and Ethyl Acetate in Table Wines

Approximately 40% of defatted perilla seeds consists of proteins which are primarily composed of globulin (84%). The amino acid profile of perilla proteins demonstrated balanced amounts of all essential amino acids, except for lysine. The molecular mass of the predominant globulin was estimated to be 340 kDa by gel filtration. This globulin was separated into three intermediary subunits (54, 57 and 59 kDa) by SDS–PAGE. It is suggested from these results that the globulin exists as a hexamer. A treatment with 50 mM dithiothreitol enabled the intermediary subunits to be separated into three acidic subunits (31–34 kDa) and four basic subunits (23–25 kDa). It is interesting that this subunit structure is the same as that of sesame α-globulin, despite them coming from different families. Compared to sesame α-globulin, the heat-induced gel of perilla globulin had better water-holding ability, despite it displaying the same degree of gel hardness.     

Oviposition and development response of perilla seed bugs (Nysius sp.) (Heteroptera: Lygaeidae) to five crop seeds

The perilla seed bugs, Nysius plebeius Distant and Nysius hidakai Nakatani, are emerging pests of perilla crop in Korea. Here, we investigated the life-history variables of N. plebeius and N. hidakai on five commonly grown crop seeds: perilla [Perilla frutescens (L.) Britt.], sesame (Sesamum indicum L.), sorghum [Sorghum bicolor (L.)], proso millet (Panicum miliaceum L.) and foxtail millet (Setaria italic L.). Bug oviposition, total developmental period, adult emergence, adult longevity and adult weight of both Nysius species were determined in both choice and no-choice tests. The oviposition and development periods of N. plebeius and N. hidakai varied significantly among seed species. The most preferred seed species for oviposition were perilla seeds, followed by sesame, sorghum and foxtail millet, whereas proso millet seeds were the least preferred. The bugs development period was faster in perilla seeds, with higher adult emergence, greater adult longevity and heavier adult weight. The higher oviposition preference and faster development with longer longevity of both Nysius species on perilla seeds is probably due to the physicochemical and nutritional attributes of the seeds. This information may enable the exploration of resistant genetic materials and chemical traits associated with seeds for breeding programme. Further, the outcomes of this study are discussed in terms of bug's behavioural response to crop seeds and management strategy of Nysius species as alternative to chemical management.     

Intermolecular Forces Involved in the Gelation and Gel Stability of Sesame 13S Globulin

The effect of various reagents on the formation and stability of heat-induced gels of sesame 13S globulins were investigated. Electrostatic interaction, the hydrophobic bond and the disulfide bond were important for forming the network structure of gels, and the hydrogen bond also had an influence on the formation of the gel. Hydrophobic bonds mainly contributed to the stability of the gel. Subunit analyses of the proteins solubilized from the gels showed the presence of a free acidic subunit (AS) and basic subunit (BS), a polymer of AS, a dimer of BS and the dimer of a fragment from AS or BS. From the results, sulfhydryl-disulfide exchange reactions during gelation are suggested.     

Effect of the dietary alpha-linolenate/linoleate balance on leukotriene production and histamine release in rats

Rats were fed semi-purified diets supplemented either with safflower seed oil rich in linoleate (18:2n-6) or with perilla seed oil rich in alpha-linolenate (18:3n-3) through two generations. In the major phospholipids of polymorphonuclear leukocytes (PMNs), the proportions of n-6 polyunsaturated fatty acids (18:2, 20:4, 22:4 and 22:5) were higher but those of n-3 acids (20:5, 22:5 and 22:6) were lower in the safflower group than in the perilla group. When stimulated with a calcium ionophore, the PMNs from the safflower group produced 27% more leukotriene (LT)B4 than those from the perilla group. The formation of LTB5 which has biological activities less than 1/10 those of LTB4, was negligible in the safflower group but was 40 ng/10(7) PMN cells in the perilla group. The amount of the total LTB formed in the perilla group tended to be more than in the safflower group. The formation of SRS-A (slow-reacting substances of anaphylaxis) by PMNs was determined by measuring the spasmogenic activities of LTs on guinea pig ileum. SRS-A activity was 59% higher in the safflower group than in the perilla group. In contrast, histamine release from rat peritoneal mast cells was not significantly different between the two groups. Thus, the increasing the alpha-linolenate/linoleate ratio of diets results in the decreased formation of LTs derived from 20:4n-6 in PMNs. This may be beneficial in lowering the severity of allergic and inflammatory responses caused by LTs, and thereby shifting the pathological symptoms to normal self-defense mechanism.     

Functional complementation of a periila ω3 fatty acid desaturase under the seed-specific SeFAD2 promoter

Functional characterization of the fatty acid desaturase genes and seed-specific promoters is prerequisite for altering the unsaturated fatty acid content of oilseeds by genetic manipulation. The ω-6 fatty acid desaturase (FAD2) and ω-3 fatty acid desaturase (FAD3) catalyze extra-plastidial desaturation of oleic acid to linoleic acid and linoleic acid to linolenic acid, respectively. These are major constituents in seed storage oils. Here, we report the complementation of a perilla linoleic acid desaturase (PrFAD3) cDNA under the seed-specific sesame FAD2 (SeFAD2) promoter in the Arabidopsis fad3 mutant. PrFAD3 is functionally active and the SeFAD2 promoter is applicable for modifying fatty acid composition in developing seeds. Transient expression of the GUS gene under that promoter in the developing seeds and leaves of sesame, soybean, and corn via microprojectile bombardment indicated that the SeFAD2 promoter likely will be useful for altering the seed phenotypes of dicot and monocot crops.     

高粱、紫苏叶脉密度与光合特性的关系

叶脉是植物叶片光合作用水分输送的重要结构。为阐述叶脉与光合特性之间的关系,以C4植物高粱(Sorghum bicolor)、C3植物紫苏(Perilla frutescens)为实验材料研究了叶脉密度和光合特性之间的关系。结果表明,与紫苏相比,高粱叶片叶脉密度大,导水能力强,蒸腾速率高,但气孔密度小。进一步分析表明,高粱叶片近轴侧气孔密度占总气孔的比例明显高于紫苏。叶脉密度大的高粱具有较高的净光合速率;而紫苏叶脉密度小,净光合速率也较低。由此表明,较高的叶脉密度有利于支持较高的光合速率,但研究表明叶脉密度和气孔密度可能不存在严格的协同变异关系。研究结果对理解植物光合作用适应有重要意义。     

Comparative effects of perilla and fish oils on the activity and gene expression of fatty acid oxidation enzymes in rat liver

The activity and mRNA level of hepatic enzymes in fatty acid oxidation and synthesis were compared in rats fed diets containing either 15% saturated fat (palm oil), safflower oil rich in linoleic acid, perilla oil rich in α-linolenic acid or fish oil rich in eicosapentaenoic (EPA) and docosahexaenoic acids (DHA) for 15 days. The mitochondrial fatty acid oxidation rate was 50% higher in rats fed perilla and fish oils than in the other groups. Perilla and fish oils compared to palm and safflower oils approximately doubled and more than tripled, respectively, peroxisomal fatty acid oxidation rate. Compared to palm and safflower oil, both perilla and fish oils caused a 50% increase in carnitine palmitoyltransferase I activity. Dietary fats rich in n-3 fatty acids also increased the activity of other fatty acid oxidation enzymes except for 3-hydroxyacyl-CoA dehydrogenase. The extent of the increase was greater with fish oil than with perilla oil. Interestingly, both perilla and fish oils decreased the activity of 3-hydroxyacyl-CoA dehydrogenase measured using short- and medium-chain substrates. Compared to palm and safflower oils, perilla and fish oils increased the mRNA level of many mitochondrial and peroxisomal enzymes. Increases were generally greater with fish oil than with perilla oil. Fatty acid synthase, glucose-6-phosphate dehydrogenase, and pyruvate kinase activity and mRNA level were higher in rats fed palm oil than in the other groups. Among rats fed polyunsaturated fats, activities and mRNA levels of these enzymes were lower in rats fed fish oil than in the animals fed perilla and safflower oils. The values were comparable between the latter two groups. Safflower and fish oils but not perilla oil, compared to palm oil, also decreased malic enzyme activity and mRNA level. Examination of the fatty acid composition of hepatic phospholipid indicated that dietary α-linolenic acid is effectively desaturated and elongated to form EPA and DHA. Dietary perilla oil and fish oil therefore exert similar physiological activity in modulating hepatic fatty acid oxidation, but these dietary fats considerably differ in affecting fatty acid synthesis.     

Cross-linking of cold-insoluble globulin by fibrin-stabilizing factor.

Cold-insoluble globulin (CI globulin) was purified from human plasma and identified on the basis of its sedimentation coefficient, electrophoretic mobility, and concentration in normal plasma. CI globulin was distinguished from antihemophilic factor (AHF) by amino acid analysis, position of elution from 4% agarose, and electrophoretic migration in polyacrylamide gels in the presence of sodium dodecyl sulfate without prior reduction. CI globulin and AHF could not be distinguished by polyacrylamide gel electrophoresis in sodium dodecyl sulfate after reduction and probably have very similar subunit molecular weights. CI globulin apparently consists of two polypeptide chains, each of molecular weight 2.0 x 10(5), held together by disulfide bonds. CI globulin was a substrate for activated fibrin-stabilizing factor (FSF, blood coagulation factor XIII). FSF catalyzed the incorporation of a fluorescent primary amine, N-(5-aminopentyl)-5-dimethylaminonaphthalene-1-sulfonamide, into CI globulin and also catalyzed the cross-linking of CI globulin into multimers, as judged by polyacrylamide gel electrophoresis in sodium dodecyl sulfate after reduction. In the presence of fibrin, cross-linking of CI globulin by FSF occurred without the formation of CI globulin multimers. Instead, polypeptides with apparent molecular weights of 2.6 x 10(5) and 3.0 x 10(5) were seen. The formation of these polypeptides coincided with the loss of the alpha chain of fibrin and CI globulin. The polypeptides were not seen when fibrin alone was cross-linked. The formation of the polypeptides was greater in fine clots than in coarse clots, and greater in clots incubated at 0 degrees than in clots incubated at 37 degrees. In clots made from purified fibrinogen, CI globulin, and FSF, the concentration of CI globulin in the clot liquor was greater if either FSF or calcium ion was omitted and cross-linking did not take place. These observations suggest that CI globulin is enzymically cross-linked to one of the chains of fibrin, most likely the alpha chain, and is thus covalently incorporated into the fibrin clot. CI globulin is very similar to a protein in the plasma membrane of fibroblasts. The cross-linking of CI globulin to itself and to fibrin may typify reactions also involving the fibroblast membrane protein.     

Effect of dietary alpha-linolenate on platelet-activating factor production in rat peritoneal polymorphonuclear leukocytes.

The effects of dietary alpha-linolenate (18:3, n-3) and linoleate (18:2, n-6) on platelet-activating factor (PAF) production were examined. Rats were fed an alpha-linolenic acid-rich (perilla oil) diet or a linoleic acid-rich (safflower oil) diet for 6 wk, and polymorphonuclear leukocytes (PMN) were elicited by peritoneal injection of casein. The overall phospholipid content and composition as well as the subclass distribution of choline and ethanolamine glycerophospholipids in PMN were not altered by these diets. However, with the perilla oil diet their content of a putative precursor of PAF, 1-alkyl-2-arachidonoyl-sn-glycero-3-phosphocholine was approximately 50% of that with safflower oil diet. On exposure to various concentrations of FMLP, PAF formation by PMN in the perilla oil group was less than 50% of that by PMN in the safflower oil group. A larger difference in PAF productions by PMN in the two dietary groups was observed on their stimulation with calcium ionophore A23187. These results demonstrate that PAF production is modulated in some as yet unknown way by changing the alpha-linolenate/linoleate balance of the diet.     

Formation of thermally induced aggregates of the soya globulin beta-conglycinin.

The effect of ionic strength (I) on the formation of thermally induced aggregates by the 7S globular storage protein of soya, beta-conglycinin, has been studied using atomic force microscopy. Aggregates were only apparent when I> or =0.1, and had a fibrous appearance, with a height (diameter) of 8-11 nm. At high ionic strength (I=1.0) the aggregates appeared to associate into clumps. When aggregate formation was studied at I=0.2, it was clear that aggregation only began at temperatures above the main thermal transition for the protein at 75 degrees C, as determined by differential scanning calorimetry. This coincided with a small change in secondary structure, as indicated by circular dichroism spectroscopy, suggesting that a degree of unfolding was necessary for aggregation to proceed. Despite prolonged heating the size of the aggregates did not increase indefinitely, suggesting that certain beta-conglycinin isoforms were able to act as chain terminators. At higher protein concentrations (1% w/v) the linear aggregates appeared to form large macroaggregates, which may be the precursors of protein gel formation. The ability of beta-conglycinin to form such distinctive aggregates is discussed in relation to the presence of acidic inserts in certain of the beta-conglycinin subunits, which may play an important role in limiting aggregate length.     

Tolerant and Susceptible Sesame Genotypes Reveal Waterlogging Stress Response Patterns

Waterlogging is a common adverse environmental condition that limits plant growth. Sesame (Sesamum indicum) is considered a drought-tolerant oil crop but is typically susceptible to harmful effects from waterlogging. The present study used comparative analysis to explore the waterlogging stress response associated with two sesame genotypes. The RNA-seq dataset generated during a time course of 0, 3, 9 and 15 h of waterlogging as well as 20 h post-drainage indicated that stress gradually suppressed the expression of sesame genes, with 9 h as the critical time point for the response of sesame to waterlogging stress. Of the 19,316 genes expressed during waterlogging, 72.1% were affected significantly. Sesame of both tolerant and susceptible genotypes showed decreased numbers of upregulated differentially expressed genes (DEGs) but increased numbers of downregulated DEGs at the onset of waterlogging. However, the tolerant-genotype sesame exhibited 25.5% more upregulated DEGs and 29.7% fewer downregulated DEGs than those of the susceptible-genotype strain between 3 and 15 h. The results indicated that the tolerant sesame displayed a more positive gene response to waterlogging. A total of 1,379 genes were significantly induced and commonly expressed in sesame under waterlogging conditions from 3 to 15 h regardless of tolerance level; of these genes, 98 are known homologous stress responsive genes, while the remaining 1,281 are newly reported here. This gene set may represent the core genes that function in response to waterlogging, including those related mainly to energy metabolism and phenylpropanoid biosynthesis. Furthermore, a set of 3,016 genes functioning in energy supply and cell repair or formation was activated in sesame recovery from waterlogging stress. A comparative analysis between sesame of the tolerant and susceptible genotypes revealed 66 genes that may be candidates for improving sesame tolerance to waterlogging. This study provided a comprehensive picture of the sesame gene expression pattern in response to waterlogging stress. These results will help dissect the mechanism of the sesame response to waterlogging and identify candidate genes to improve its tolerance.     

Competition between two similar plant varieties,green shrunk perilla and red shrunk perilla,in mixed cultures

Influences of plant density and time after seeding on the growth of two horticultural forms of perilla (Perilla frutescens var.crispa), green shrunk perilla (f.viridi-crispa) and red shrunk perilla (f.crispa), were examined in a mixed culture experiment. Relationships between mean individual plant weight and plant density in mixed populations were approximated by Ogawa's non-interaction type (NI-type) reciprocal equation. The density conversion factors in the equation for green and red perillas were always, respectively, smaller and larger than unity, suggesting that effects of a green perilla on the other individuals were always stronger than those of a red one in a mixed population. All coefficients in the NI-type reciprocal equation were expressed as functions of time after seeding. As a result, time trends of mean individual plant weights for both species in mixed populations could be reasonably estimated for different plant densities and mixed proportions. The results were also applied to Lotka-Volterra's equation. Time trends of Lotka-Volterra's competition coefficients for both plants could be calculated and were compared with those of density conversion factors.     

Effect of dietary alpha-linolenate/linoleate balance on mean survival time, incidence of stroke and blood pressure of spontaneously hypertensive rats

Following the suckling period, stroke-prone spontaneously hypertensive rats (SHR-SP) were fed semi-purified diets supplemented either with safflower seed oil (rich in linoleic acid) or with perilla seed oil (rich in alpha-linolenic acid). The mean survival time of male SHR-SP fed the perilla diet was longer than that fed the safflower diet by 17% (p less than 0.001) while the difference was 15% in female SHR-SP (p less than 0.05). The mean survival times of female SHR-SP were more than 40% longer than those of male SHR-SP in both dietary groups. Post-mortem examinations of brains revealed apoplexy-related symptoms as the major cause of the death in both dietary groups. The systolic blood pressure was lower by ca. 10% (21 mmHg) in the perilla group than in both the safflower group and conventional diet group. The eicosapentaenoate (20:5 n-3)/arachidonate (20:4 n-6) ratio of platelet phospholipids in spontaneously hypertensive rat (SHR), a measure of platelet aggregability, was much higher in the perilla group than in the safflower group. Thus, increasing the dietary alpha-linolenate/linoleate ratio resulted in an increased mean survival time of SHR-SP rats, possibly by lowering blood pressure and platelet aggregability.     

Differential gene expression profiles of red and green forms of Perilla frutescens leading to comprehensive identification of anthocyanin biosynthetic genes

Differential screening by PCR-select subtraction was carried out for cDNAs from leaves of red and green perilla, two chemovarietal forms of Perilla frutescens regarding anthocyanin accumulation. One hundred and twenty cDNA fragments were selected as the clones preferentially expressed in anthocyanin-accumulating red perilla over the nonaccumulating green perilla. About half of them were the cDNAs encoding the proteins related presumably to phenylpropanoid-derived metabolism. The cDNAs encoding glutathione S-transferase (GST), PfGST1, and chalcone isomerase (CHI), PfCHI1, were further characterized. The expression of PfGST1 in an Arabidopsis thaliana tt19 mutant lacking the GST-like gene involved in vacuole transport of anthocyanin rescued the lesion of anthocyanin accumulation in tt19, indicating a function of PfGST1 in vacuole sequestration of anthocyanin in perilla. The recombinant PfCHI1 could stereospecifically convert naringenin chalcone to (2S)-naringenin. PfGST1 and PfCHI1 were preferentially expressed in the leaves of red perilla, agreeing with the accumulation of anthocyanin and expression of other previously identified genes for anthocyanin biosynthesis. These results suggest that the genes of the whole anthocyanin biosynthetic pathway are regulated in a coordinated manner in perilla.     

Interactions and compatibility of 11 S globulin from Vicia faba seeds and sodium salt of carboxymethylcellulose in an aqueous medium

This work studies specific interactions and compatibility between a legumin and a linear carboxylated polysaccharide using gel permeation chromatography, sedimentation analysis, SDS gel electrophoresis, viscometry and phase analysis measurements. It uses the system water/11 S globulin/CMC as a model. Carboxymethylcellulose (CMC) molecules are able to cause a partial dissociation of the protein, subsequent formation of soluble interbiopolymeric complexes and partial aggregation of the free non combined protein at room temperature and pH 6.0-6.5. The maximal binding of biopolymers is observed at their equimolar ratio. The decrease in temperature of the mixture from 293 to 277 K leads to formation of the complex coacervate. The increase in pH from 6.0 to 7.6 results in suppression of complex formation and manifestation of the phenomenon of thermodynamic incompatibility if the total concentration of biopolymers in the system exceeds the critical concentration of segregative phase separation.     

Analysis of globulin maturation in developing sunflower seeds

The synthesis of major storage globulin polypeptides has been examined in developing seeds of sunflower(Helianthus annuus L.). Analyses of total proteins and purified globulins, also called helianthinin, by gel electrophoresis and immunoelectrophoresis have shown that a burst of protein synthesis and accumulation occurs around 10 d after flowering. There is no mature globulin before that time and only small amounts of precursor forms can be detected. Thus, 10–12 d after flowering appears to be a transition period during which genetic information for the globulin becomes actively expressed. Immunoelectrophoresis has confirmed that globulin is the main storage protein, at seed maturation, accounting up to 70 % of total proteins per kernel. Pulse chase experiments have shown that synthesis initially involves the formation of high molecular mass precursors and that storage proteins are post-translationally processed. Intermediary products, with molecular mass higher than early translational products, can be detected, together with mature globulin polypeptides.     

Anti-inflammatory and anti-allergic actions by oral administration of a perilla leaf extract in mice

The anti-inflammatory and anti-allergic activity of perilla leaf extract was investigated. The oral administration of perilla leaf extract to mice inhibited two types of acute inflammatory models, arachidonic acid-induced ear edema and 12-o-tetradecanoylphorbol-13-acetate-induced ear edema. Oral administration of perilla leaf extract also inhibited the contact dermatitis model, oxazolone-induced ear edema, by affecting sensitization.