Substructure of the Cytoplasmic Membrane of Bacillus megaterium

The components of fractions obtained by dialyzing and differential centrifuging the “Ghost” of Bacillus megaterium were analyzed in detail. The compositions of amino acids in the main fractions (Fraction 2 and 3) of the “Ghosts”, were estimated. Fraction 2 was rich in non-polar amino acids, while Fraction 3 was scanty of them. Most of the fatty acids in Fraction 2 were 12-methyl tetradecanoic acid, while in Fraction 3 many kinds of fatty acid were detected.

As for the localization of enzymes, the three enzymes, glucose oxidase, succinic dehydrogenase and reduced nicotinamide-adenine dinucleotide oxidase, which were present in the original “Ghosts”, were mostly observed in Fraction 2, and a very little amount of them was found in the other fractions. Further, Fraction 2 could be dissolved in formic acid and dialysis of the solution brought about reaggregation to form membrane-like structure in the presence of Ca or Mg ion.     

Vinyldiaminotriazine-acridine conjugate as G-quadruplex alkylating agent

Higher-order structures of nucleic acids have become widely noted for their biological consequences and the discovery of an alkylating small molecule for these structures has been of interest due to its therapeutic potential. We previously developed the vinyldiaminotriazine (VDAT)-acridine conjugate as a T-T mismatch alkylating agent. In this report, we focused on the finding of the alkylation to the G-quadruplex (G4) DNA with VDAT-acridine conjugates. The VDAT-acridine conjugates exhibited a considerable alkylation ability to G4 under mild conditions. Moreover, the investigation of properties with the alkylated G4 revealed that alkylation by this conjugate significantly increased the stability of the G4 structure. This study provides a starting point in the further development of selective G4 alkylating small molecules.     

Involvement of Proteasome-Ubiquitin System in Wound-Signaling in Tobacco Plants

Treatment of wounded tobacco leaves with a proteasome-specificinhibitor lactacystin resulted in; (1) enhanced expression ofproteasome subunit genes, (2) suppressed expression of proteasomeactivity, (3) induced accumulation of Ub-conjugated proteins,and (4) suppressed expression of wound-inducible genes, indicatingthat the Ub-proteasome system is involved in wound-signalingin plants. ³ Present address: Department of Plant Breeding, National AgricultureResearch Center, 3-1-1 Kannondai, Tsukuba, Ibaraki, 305-8666Japan.     

Transgenic-cloned pigs systemically expressing red fluorescent protein, Kusabira-Orange

Genetically engineered pigs with cell markers such as fluorescent proteins are highly useful in lines of research that include the tracking of transplanted cells or tissues. In this study, we produced transgenic-cloned pigs carrying a gene for the newly developed red fluorescent protein, humanized Kusabira-Orange (huKO), which was cloned from the coral stone Fungia concinna. The nuclear transfer embryos, reconstructed with fetal fibroblast cells that had been transduced with huKO cDNA using retroviral vector DDeltaNsap, developed efficiently in vitro into blastocysts (28.0%, 37/132). Nearly all (94.6%, 35/37) of the cloned blastocysts derived from the transduced cells exhibited clear huKO gene expression. A total of 429 nuclear transfer embryos were transferred to four recipients, all of which became pregnant and gave birth to 18 transgenic-cloned offspring in total. All of the pigs highly expressed huKO fluorescence in all of the 23 organs and tissues analyzed, including the brain, eyes, intestinal and reproductive organs, skeletal muscle, bone, skin, and hoof. Furthermore, such expression was also confirmed by histological analyses of various tissues such as pancreatic islets, renal corpuscles, neuronal and glial cells, the retina, chondrocytes, and hematopoietic cells. These data demonstrate that transgenic-cloned pigs exhibiting systemic red fluorescence expression can be efficiently produced by nuclear transfer of somatic cells retrovirally transduced with huKO gene.     

Correlation between reduction potentials and inhibitions of Epstein-Barr virus activation by anthraquinone derivatives

As a continuation of studies using natural and synthetic products as cancer chemopreventive agents, we used cyclic voltammetry to examine the reduction–oxidation potentials of methylated emodin derivatives prepared from emodin in phosphate buffer at pH 7.2. A good correlation was found between the inhibitory effects on Epstein–Barr virus early antigen (EBV-EA) activation and the reduction potential of methylated emodin derivatives. Furthermore, there was significant correlation between EBV-EA activation and the reduction potential of 35 anthraquinone derivatives including methylated emodin derivatives. It was further shown that the correlation could be enhanced by including LUMO energy and the number of hydroxy groups as additional parameters.     

Development of novel 2-[4-(aminoalkoxy)phenyl]-4(3H)-quinazolinone derivatives as potent and selective histamine H3 receptor inverse agonists

Novel 2-[4-(aminoalkoxy)phenyl]-4(3H)-quinazolinone derivatives were identified as potent human H(3) receptor inverse agonists. After systematic modification of lead 5a, the potent and selective analog 5r was identified. Elimination of hERG K(+) channel and human alpha(1A)-adrenoceptor activities is the main focus of the present study.     

Heterotypy in the N-terminal region of growth/differentiation factor 5 (GDF5) mature protein during teleost evolution

Heterotypy is now recognized as a generative force in the formationof new proteins through modification of existing proteins. Wereport that heterotypy in the N-terminal region of the maturegrowth/differentiation factor 5 (GDF5) protein occurred duringevolution of teleosts. N-terminal length variation of GDF5 wasfound among teleost interfamilies and interorders but not withinteleost families or among tetrapods. We further show that increaseof proline and glutamine to the N-terminal region of matureGDF5 occurred in Eurypterygii, the higher lineage of teleosts.Because the basic amino acids, believed to control diffusion,are conserved in this region across all species examined, wesuggest that the N-terminal elongation of the mature GDF5 proteinduring evolution has altered the protein diffusion in Eurypterygii,leading to high concentrations of the protein in the joint ofthe pharyngeal skeleton, the location of cartilage formationduring development.     

Genetic Evidence That the Non-Homologous End-Joining Repair Pathway Is Involved in LINE Retrotransposition

Long interspersed elements (LINEs) are transposable elements that proliferate within eukaryotic genomes, having a large impact on eukaryotic genome evolution. LINEs mobilize via a process called retrotransposition. Although the role of the LINE-encoded protein(s) in retrotransposition has been extensively investigated, the participation of host-encoded factors in retrotransposition remains unclear. To address this issue, we examined retrotransposition frequencies of two structurally different LINEs—zebrafish ZfL2-2 and human L1—in knockout chicken DT40 cell lines deficient in genes involved in the non-homologous end-joining (NHEJ) repair of DNA and in human HeLa cells treated with a drug that inhibits NHEJ. Deficiencies of NHEJ proteins decreased retrotransposition frequencies of both LINEs in these cells, suggesting that NHEJ is involved in LINE retrotransposition. More precise characterization of ZfL2-2 insertions in DT40 cells permitted us to consider the possibility of dual roles for NHEJ in LINE retrotransposition, namely to ensure efficient integration of LINEs and to restrict their full-length formation.