Molecular evidence for the taxonomic status of three species of the <Emphasis Type="Italic">Sphyraena obtusata</Emphasis> group (Perciformes: Sphyraenidae) from East Asia

Genetic differences among the three species of Sphyraena with two gill rakers from East Asia (S. iburiensis, S. obtusata, and S. pinguis, defined recently as the S. obtusata group), were investigated using 799-bp sequences of the mitochondrial cytochrome b gene. Pairwise sequence differences within each of the three species were 0.0–0.4% (S. iburiensis), 0.0–0.4% (S. obtusata), and 0.0–0.6% (S. pinguis), respectively, pairwise sequence differences among the three species being 18.3–18.6% (S. iburiensis vs. S. obtusata), 14.9–15.4% (S. iburiensis vs. S. pinguis), and 17.6–18.3% (S. obtusata vs. S. pinguis), respectively. The extent of the latter were comparable to those among four other congeneric species, S. africana, S. forsteri, S. helleri, and S. japonica (16.0–24.5%). Phylogenetic analysis using the maximum-likelihood method indicated reciprocal monophyly of the three species, each clade being supported by a bootstrap value of 100%. These findings fully supported the taxonomic status of these species, recently elucidated by detailed morphological comparisons. In addition, the S. obtusata group formed a strongly supported clade against the four other congeneric species.     

Genetic diversity of <Emphasis Type="Italic">Salanx ariakensis</Emphasis> (Salangidae) from Korea and Japan inferred from AFLP

The genetic diversity and population structures within and between Korean and Japanese populations of Salanx ariakensis were investigated using AFLP (amplified fragment length polymorphism). Using seven primer pair sets, 411 fragments were amplified from 31 specimens of S. ariakensis, 393 fragments (96%) being polymorphic. The expected heterozygosities were 0.211 and 0.245 for the Japanese and Korean populations, respectively, levels similar to those of other freshwater and diadromous fishes, but lower than those of marine fishes. The difference in genetic diversity between the two populations may result from their different habitat sizes. Principal coordinate analysis using 393 polymorphic fragments resulted in specimens from each population being plotted separated, suggesting an absence (or little if present) gene flow between them since Tsushima Strait has been open. Therefore, each population of S. ariakensis is relevant from the conservation perspective and should at least be treated as a “management unit” for conservation purposes.     

Synaptosomal-associated protein 25 gene expression is hormonally regulated during ovulation and is involved in cytokine/chemokine exocytosis from granulosa cells

During ovulation, granulosa cells and cumulus cells synthesize and secrete a wide variety of factors including members of the IL cytokine family via the process of exocytosis. Exocytosis is controlled by the soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptor complex consisting of proteins residing in the vesicle membrane and the plasma membrane. One of the soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptor proteins, synaptosomal-associated protein (SNAP)25, is expressed abundantly in neuronal cells and is also induced transiently in the rat ovary in response to LH. Therefore, we sought to determine the molecular mechanisms controlling ovarian expression of the Snap25 gene, and the role of SNAP25 in exocytosis of secreted factors, such as ILs from cumulus cells and granulosa cells. In preovulatory follicles of equine (e) chorionic gonadotropin (CG)-primed mice, expression of Snap25 mRNA was negligible but was induced markedly 8 h after human (h) CG stimulation. In Pgr null mice Snap25 mRNA and protein levels were significantly lower at 8 h after hCG compared with wild-type mice. To analyze the molecular mechanisms by which progesterone receptor regulates this gene, a 1517-bp murine Snap25 promoter-luciferase reporter construct was generated and transfected into granulosa cell cultures. Three specificity protein (SP)-1/SP-3 sites, but not consensus activator protein 1 or cAMP response element sites, were essential for basal and forskolin/phorbol 12-myristate 13-acetate-induced promoter activity in granulosa cells. The induction was significantly suppressed by PGR antagonist, RU486. Treatment of cumulus oocyte complexes or granulosa cells with FSH/amphiregulin, LH, or forskolin/phorbol 12-myristate 13-acetate-induced elevated expression of Snap25 mRNA and increased the secretion of eight cytokine and chemokine factors. Transfection of granulosa cells with Snap25 small interfering RNA significantly reduced the levels of both SNAP25 protein and the secretion of cytokines. From these results, we conclude that progesterone-progesterone receptor-mediated SNAP25 expression in cumulus oocyte complexes and granulosa cells regulates cytokine and chemokine secretion via an exocytosis system.     

Selective photo-cross-linking of 2'-O-psoralen-conjugated oligonucleotide with RNAs having point mutations

It has been reported that point mutations in genes are responsible for various cancers and the selective regulation of the gene expression is an important issue to develop a new type of anticancer drugs. In this report, we present a new type of antisense molecule that photo-cross-links to an oligoribonucleotide having a point mutation site in a sequence specific manner. 2'-O-psoralen-conjugated adenosine was synthesized in four steps from adenosine and introduced in the middle of an oligodeoxyribonucleotide (2'-Ps-oligo). Compared with 5'-O-psoralen-conjugated oligodeoxyribonucleotide (5'-Ps-oligo), which has a psoralen at the 5'-end, 2'-Ps-oligo more selectively photo-cross-linked to a pyrimidine base of the site of alteration from purine to pyrimidine in the oligoribonucleotide.     

Comparisons of soil‐water content between a Moso bamboo (Phyllostachys pubescens) forest and an evergreen broadleaved forest in western Japan

In Japan, forests of Moso bamboo (Phyllostachys pubescens, an exotic invasive giant bamboo) have naturalized and expanded rapidly, replacing surrounding broadleaved and coniferous forests. To evaluate impacts caused by these forest‐type replacements on the hydrological cycle, soil‐water content and its spatial variability in a Moso bamboo forest were compared with those in an adjacent evergreen broadleaved forest, in a case study of a stand in western Japan (northern Kyushu). The volumetric soil‐water content averaged over depths between 0 and 60 cm was consistently higher in the bamboo stand than that in the broadleaved stand. These results contrast with previous studies comparing the soil‐water content in Moso bamboo forests with that in other forest types. The sum of canopy transpiration and soil evaporation (E) in the bamboo stand tended to be larger than that in the broadleaved stand. Small canopy interception loss was reported in the bamboo forest. Therefore, the large amount of E would counterbalance the small canopy interception loss in the bamboo forest. Differences in soil characteristics between the two stands may be the main factor causing differences in soil‐water content. Spatial variation in soil‐water content in the bamboo stand was larger than that in the broadleaved stand, confirming findings in a previous series of our study. This could happen because the well‐developed root‐system in the bamboo forest enhances preferential flow in the soil. To evaluate the effects of aggressive invasion of alien giant bamboo on the ecosystem functions, we recommend further studies measuring various hydrological components in various Moso bamboo forests.     

Developmental changes in crossover frequency in Arabidopsis

Parental genomes are generally rearranged by two processes during meiosis: one is the segregation of homologous chromosomes and the other is crossing over between such chromosomes. Although the mechanisms underlying chromosome segregation and crossing over are well understood because of numerous genetic and molecular investigations, their contributions to the rearrangement of genetic information have not yet been analysed at a genome-wide level in Arabidopsis thaliana. We established 343 CAPS or SSLP markers to identify polymorphisms between two different Arabidopsis ecotypes, Col and Ler, which are distributed at an average distance of approximately 400kb between pairs of markers throughout the entire genome. Using these markers, crossover frequencies and chromosome segregation were quantified with respect to sex and age. Our large-scale analysis demonstrated that: (i) crossover frequencies during pollen formation were 1.79 and 1.37 times higher than those during megaspore formation in early and late flowers, respectively (P<0.001); (ii) the crossover frequencies during pollen formation were not significantly different between early and late flowers of main shoots (P>0.05), whereas the frequencies increased 1.30 times with shoot age during megaspore formation (P<0.001); (iii) the effect of aging depended on the developmental age of the individual shoot rather than on the age of the whole plant; and (iv) five chromosomes were randomly selected and mixed during meiosis.     

Yeast thioredoxin-enriched extracts for mitigating the allergenicity of foods

Thioredoxin (TRX) catalyzes the reduction of disulfide bonds in proteins via the NADPH-dependent thioredoxin reductase system. Reducing the disulfide bonds of allergenic proteins in food by TRX lowers the allergenicity. We established in this study a method to prepare TRX-enriched extracts from the edible yeast, Saccharomyces cerevisiae, on a large and practical scale, with the objective of developing TRX-containing functional foods to mitigate food allergy. Treating with the yeast TRX-enriched extracts together with NADPH and yeast thioredoxin reductase enhanced the pepsin cleavage of β-lactoglobulin and ovomucoid (OM). We also examined whether yeast TRX can mitigate the allergenicity of OM by conducting immediate allergy tests on guinea pigs. The treatment with TRX reduced the anaphylactic symptoms induced by OM in these tests. These results indicate that yeast TRX was beneficial against food allergy, raising the possibility that yeast TRX-enriched extracts can be applied to food materials for mitigating food allergy.     

Structure-activity relationship studies of sphingosine-1-phosphate receptor agonists with N-cinnamyl-β-alanine moiety

Structure-activity relationship of sphingosine-1-phosphate receptor agonist was examined. In terms of reducing the flexibility of molecule, hit compound 1 was modified to improve S1P₁ agonistic activity as well as selectivity over S1P₃ agonistic activity. Novel S1P agonists with cinnamyl scaffold or 1,2,5,6-tetrahydropyridine scaffold were identified.