Development of a 16S rRNA gene primer and PCR-restriction fragment length polymorphism method for rapid detection of members of the genus Megasphaera and species-level identification

The genus Megasphaera is relevant to the environment, human health and food, and renewable energy for the future. In this study, a primer set was designed for PCR-restriction fragment length polymorphism (RFLP) analyses to detect and identify the members of Megasphaera. Direct detection and identification were achieved for environmental samples and isolates.     

17,20-Lyase inhibitors. Part 3: Design, synthesis, and structure-activity relationships of biphenylylmethylimidazole derivatives as novel 17,20-lyase inhibitors

A novel series of biphenylylmethylimidazole derivatives and related compounds were synthesized as inhibitors of 17,20-lyase, a key enzyme in the production of steroid hormones, and their biological activities were evaluated. In an attempt to identify potent and selective inhibitors of 17,20-lyase over the related CYP3A4 enzyme, a homology model for human 17,20-lyase was developed using the X-ray crystallographic structure of the mammalian CYP2C5 enzyme. With the aid of molecular modeling, optimization of the biphenyl moiety was performed to give an acetamide derivative, which was resolved by HPLC to give the active (-)-enantiomer. The obtained active enantiomer showed not only potent inhibition of both rat and human 17,20-lyase,with IC(50) values of 14 and 26 nM, respectively, but also excellent selectivity (>300-fold) for inhibition of 17,20-lyase over CYP3A4. Moreover, the active enantiomer significantly reduced both serum testosterone and DHEA concentrations in a monkey model after single oral administration. Asymmetric synthesis of the active enantiomer was also developed via a chiral intermediate using a diastereoselective Grignard reaction.     

Novel 12-membered non-antibiotic macrolides from erythromycin A; EM900 series as novel leads for anti-inflammatory and/or immunomodulatory agents

Herein, we report the design and synthesis of the novel 12-membered non-antibiotic macrolide (8R,9S)-8,9-dihydro-6,9-epoxy-8,9-anhydropseudoerythromycin A (EM900), which was found to be a potent anti-inflammatory and/or immunomodulatory agent, capable of promoting monocyte to macrophage differentiation. This molecule shows improved acid stability, does not exhibit any anti-bacterial activity and has relatively low cytotoxicity against THP-1 cells. In addition, one of its analogues, (8R,9S)-4″,13-O-diacetyl-8,9-dihydro-6,9-epoxy-8,9-anhydropseudoerythromycin A (EM911), was found to be twice as effective as EM900.     

An Arabidopsis RNase III-like protein,AtRTL2, cleaves double-stranded RNA in vitro

Class 1 ribonuclease III (RNase III), found in bacteria and yeast, is involved in processing functional RNA molecules such as ribosomal RNAs (rRNAs). However, in Arabidopsis thaliana, the lack of an obvious phenotype or quantitative change in mature rRNAs in class 1 RNase III (AtRTL2) mutants and overexpressing plants suggests that AtRTL2 is not involved in rRNA maturation. We characterized the in vitro activity of AtRTL2 to consider its in vivo function. AtRTL2 cleaved double-stranded RNA (dsRNA) specifically in vitro, yielding products of approximately 25 nt or longer in length, in contrast to 10–20 nt long products in bacteria and yeasts. Although dsRNA-binding activity was not detected, the dsRNA-binding domains in AtRTL2 were essential for its dsRNA-cleaving activity. Accumulation of small RNAs derived from transgene dsRNAs was increased when AtRTL2 was transiently expressed in Nicotiana benthamiana leaves by agroinfiltration. These results raise the possibility that AtRTL2 has functions distinct from those of other class 1 RNase IIIs in vivo.     

The relationship between chromosomal positioning within the nucleus and the SSD1 gene in Saccharomyces cerevisiae

Eukaryotic cells are characterized by very large chromosomal DNAs efficiently packed within the nucleus. To identify the mechanism of chromosomal packaging based on the uniqueness of the centromere region in Saccharomyces cerevisiae, we isolated the HCH6 mutant, which shows 2.5-fold higher efficiency of site-specific recombination between the CEN5 and HIS3 loci than the wild-type CH53 strain. This mutant also displayed defects in cell integrity at high temperature. The SSD1 gene was perhaps responsible for this defect. The efficiency of site-specific recombination was decreased by the introduction of SSD1 in HCH6 cells and increased by disruption of SSD1 in the wild-type cells. Furthermore, the distances between the CEN5 and HIS3 loci and between the CEN5 locus and the spindle pole body (SPB) indicated that disrupting SSD1 caused a loss of the anchoring of the CEN5 locus near SPB. These results suggest Ssd1p-dependent cross-talk between chromosomal positioning within the nucleus and the positioning of cellular components within the cell.     

Comparison of antioxidative effects between radon and thoron inhalation in mouse organs

Radon therapy has been traditionally performed globally for oxidative stress-related diseases. Many researchers have studied the beneficial effects of radon exposure in living organisms. However, the effects of thoron, a radioisotope of radon, have not been fully examined. In this study, we aimed to compare the biological effects of radon and thoron inhalation on mouse organs with a focus on oxidative stress. Male BALB/c mice were randomly divided into 15 groups: sham inhalation, radon inhalation at a dose of 500 Bq/m³ or 2000 Bq/m³, and thoron inhalation at a dose of 500 Bq/m³ or 2000 Bq/m³ were carried out. Immediately after inhalation, mouse tissues were excised for biochemical assays. The results showed a significant increase in superoxide dismutase and total glutathione, and a significant decrease in lipid peroxide following thoron inhalation under several conditions. Additionally, similar effects were observed for different doses and inhalation times between radon and thoron. Our results suggest that thoron inhalation also exerts antioxidative effects against oxidative stress in organs. However, the inhalation conditions should be carefully analyzed because of the differences in physical characteristics between radon and thoron.