Vasorelaxant effects of macrocyclic bis(bibenzyls) from liverworts

Vasorelaxant effects of a series of bis(bibenzyls) from liverworts such as Marchantia polymorpha and Marchantia paleacea on rat aorta demonstrated that they relaxed phenylephrine (PE)-induced contractions, which may be mediated through the increased release of NO from endothelial cells as well as opening of K(+) channels, and inhibition of Ca(2+) influx through voltage-dependent Ca(2+) channels (VDCs) and/or receptor-operated Ca(2+) channels (ROCs). Structure-activity relationship based on their structures was discussed. The presence of two aromatic rings which can be connected through two atoms bridge spacer may play an important role for vasorelaxant effect.     

Distinct roles of protein disulfide isomerase and P5 sulfhydryl oxidoreductases in multiple pathways for oxidation of structurally diverse storage proteins in rice

In the rice (Oryza sativa) endosperm, storage proteins are synthesized on the rough endoplasmic reticulum (ER), in which prolamins are sorted to protein bodies (PBs) called type-I PB (PB-I). Protein disulfide isomerase (PDI) family oxidoreductase PDIL2;3, an ortholog of human P5, contains a conserved structural disulfide in the redox-inactive thioredoxin-like (TRX) domain and was efficiently targeted to the surface of PB-I in a redox active site-dependent manner, whereas PDIL1;1, an ortholog of human PDI, was localized in the ER lumen. Complementation analyses using PDIL1;1 knockout esp2 mutant indicated that the a and a' TRX domains of PDIL1;1 exhibited similar redox activities and that PDIL2;3 was unable to perform the PDIL1;1 functions. PDIL2;3 knockdown inhibited the accumulation of Cys-rich 10-kD prolamin (crP10) in the core of PB-I. Conversely, crP10 knockdown dispersed PDIL2;3 into the ER lumen. Glutathione S-transferase-PDIL2;3 formed a stable tetramer when it was expressed in Escherichia coli, and the recombinant PDIL2;3 tetramer facilitated α-globulin(C79F) mutant protein to form nonnative intermolecular disulfide bonds in vitro. These results indicate that PDIL2;3 and PDIL1;1 are not functionally redundant in sulfhydryl oxidations of structurally diverse storage proteins and play distinct roles in PB development. We discuss PDIL2;3-dependent and PDIL2;3-independent oxidation pathways that sustain disulfide bonds of crP10 in PB-I.     

Functional and genetic survey of all known single-nucleotide polymorphisms within the human deoxyribonuclease I gene in wide-ranging ethnic groups

The single-nucleotide polymorphisms (SNPs) in the human DNase I gene (DNASE1) might be involved in susceptibility to some common diseases; however, only limited population data are available. Further, the effects of these SNPs on in vivo DNase I activity remain unknown. The genotype and haplotype of all the SNPs in DNASE1 were determined in 3 ethnic groups including 14 populations using newly developed methods. Together with our previous data on the nonsynonymous SNPs, two major haplotypes based on the five exonic SNPs were identified; genetic diversity in the Asian population was low. Among 10 SNPs, other than exonic SNPs in the gene, only 3 were polymorphic among all the populations. Haplotype distribution, based on all the polymorphic SNPs, was clarified to be generally varied in an ethnic-dependent manner. Thus, the genetic aspects of DNASE1 with regard to all the SNPs in wide-ranging ethnic groups could be first demonstrated. Further, there was no correlation of all the polymorphic SNPs other than nonsynonymous ones with serum DNase I activity levels. Polymorphic SNPs other than the exonic SNPs might not be directly related to common diseases through alterations in in vivo levels of the activity.     

Evaluation of Sycp3, Plzf and Cyclin B3 expression and suitability as spermatogonia and spermatocyte markers in zebrafish

Recent studies in mammals have revealed the heterogeneity of spermatogonial populations which contain differentiated and undifferentiated cells that further divide into actual stem cells and potential stem cells. In fish however, there are no functional definitions, and very few molecular markers, for germ cells. In our present study, specific antibodies were raised against Sycp3, Plzf and Cyclin B3 in zebrafish and then used to determine the localization of these proteins in the testis. We wished to confirm whether these molecules were potential markers for spermatocytes and spermatogonia. Immunohistochemical observations revealed that Sycp3 is specifically localized in spermatocytes in typical nuclear patterns at each meiotic stage. Plzf was found to be localized in the nucleus of both type A and type B spermatogonia until the 8-cell clone, similar to the pattern in Plzf-positive A(single)-A(aligned) undifferentiated spermatogonia in rodents. In addition to Plzf, the localization of Cyclin B3 was predominantly detected in the nuclei of type A and early type B spermatogonia until the 16-cell clone. Additionally, Cyclin B3 protein signals were detected in germ cells in large cysts, possibly corresponding to spermatocytes at the preleptotene stage. Our present data thus show that these molecules have properties that will enable their use as markers of spermatocytes and early spermatogonia in zebrafish.     

Effect of Coccolith Polysaccharides Isolated from the Coccolithophorid, <Emphasis Type="Italic">Emiliania huxleyi</Emphasis>, on Calcite Crystal Formation in In Vitro CaCO<Subscript>3</Subscript> Crystallization

Marine coccolithophorids (Haptophyceae) produce calcified scales “coccoliths” which are composed of CaCO₃ and coccolith polysaccharides (CP) in the coccolith vesicles. CP was previously reported to be composed of uronic acids and sulfated residues, etc. attached to the polymannose main chain. Although anionic polymers are generally known to play key roles in biomineralization process, there is no experimental data how CP contributes to calcite crystal formation in the coccolithophorids. CP used was isolated from the most abundant coccolithophorid, Emiliania huxleyi. CaCO₃ crystallization experiment was performed on agar template layered onto a plastic plate that was dipped in the CaCO₃ crystallization solution. The typical rhombohedral calcite crystals were formed in the absence of CP. CaCO₃ crystals formed on the naked plastic plate were obviously changed to stick-like shapes when CP was present in the solution. EBSD analysis proved that the crystal is calcite of which c-axis was elongated. CP in the solution stimulated the formation of tabular crystals with flat edge in the agarose gel. SEM and FIB-TEM observations showed that the calcite crystals were formed in the gel. The formation of crystals without flat edge was stimulated when CP was preliminarily added in the gel. These observations suggest that CP has two functions: namely, one is to elongate the calcite crystal along c-axis and another is to induce tabular calcite crystal formation in the agarose gel. Thus, CP may function for the formation of highly elaborate species-specific structures of coccoliths in coccolithophorids.     

Beyond molting--roles of the steroid molting hormone ecdysone in regulation of memory and sleep in adult Drosophila

The molting hormone 20-hydroxyecdysone (20E) is an active metabolite of ecdysone and plays vital roles during ontogeny of the fruit fly Drosophila, coordinating critical developmental transitions such as molting and metamorphosis. Although 20E is known to exist throughout life in both male and female flies, its functions in adult physiology and behavior remain largely elusive. Notably, findings from previous studies suggest that this hormone may be involved in adult stress responses. Consistent with this possibility, we have found that ecdysone signaling in adult flies is activated by "stressful" social interactions and plays a role in the formation of long-term courtship memory [Ishimoto et al. (2009). Ecdysone signaling regulates the formation of long-term courtship memory in adult Drosophila melanogaster. PNAS 106, 6381-6386]. In addition, we recently reported that ecdysone signaling contributes to the regulation of sleep, affecting transitions between sleep and wakefulness [Ishimoto and Kitamoto. (2010). The steroid molting hormone ecdysone regulates sleep in adult Drosophila melanogaster. Genetics 185, 269-281]. Here in Extra Views, we first summarize our findings on the unconventional roles of 20E in regulating memory and sleep in adult flies. We then discuss speculative ideas concerning the stress hormone-like features of 20E, as well as the possibility that ecdysone signaling contributes to remodeling of the adult nervous system, at both the functional and structural levels, through epigenetic mechanisms.