Identification of a novel cytotoxic protein, Cry45Aa, from Bacillus thuringiensis A1470 and its selective cytotoxic activity against various mammalian cell lines

Parasporal inclusion proteins produced by Bacillus thuringiensis strain A1470 exhibit strong cytotoxicity against human leukemic T cells when activated by protease treatment. One of the cytotoxic proteins was separated by anion exchange chromatography and gel filtration chromatography and designated Cry45Aa. Its gene was then expressed in recombinant Escherichia coli, in which the Cry45Aa precursor was accumulated in an inclusion body. It was solubilized in sodium carbonate buffer and processed with proteinase K, and cytotoxic activities of the protein against various mammalian cell lines were evaluated using the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H tetrazolium bromide assay. The protein exhibited high cytotoxic activity against CACO-2, Sawano, MOLT-4, TCS, and HL60 cells and moderate activity against U-937 DE-4, PC12, and HepG2 cells. On the other hand, the EC50 values against Jurkat, K562, HeLa, A549, Vero, COS-7, NIH3T3, CHO, and four normal tissue cells (human primary hepatocyte cells, UtSMC, MRC-5, and normal T cells) were >2 microg/mL.     

Applicability of Aegilops tauschii drought tolerance traits to breeding of hexaploid wheat

Few genes are available to develop drought-tolerant bread wheat (Triticum aestivum L.) cultivars. One way to enhance bread wheat’s genetic diversity would be to take advantage of the diversity of wild species by creating synthetic hexaploid wheat (SW) with the genomic constitution of bread wheat. In this study, we compared the expression of traits encoded at different ploidy levels and evaluated the applicability of Aegilops tauschii drought-related traits using 33 Ae. tauschii accessions along with their corresponding SW lines under well-watered and drought conditions. We found wide variation in Ae. tauschii, and even wider variation in the SW lines. Some SW lines were more drought-tolerant than the standard cultivar Cham 6. Aegilops tauschii from some regions gave better performing SW lines. The traits of Ae. tauschii were not significantly correlated with their corresponding SW lines, indicating that the traits expressed in wild diploid relatives of wheat may not predict the traits that will be expressed in SW lines derived from them. We suggest that, regardless of the adaptability and performance of the Ae. tauschii under drought, production of SW could probably result in genotypes with enhanced trait expression due to gene interactions, and that the traits of the synthetic should be evaluated in hexaploid level.     

Role for piRNAs and noncoding RNA in de novo DNA methylation of the imprinted mouse Rasgrf1 locus

Genomic imprinting causes parental origin-specific monoallelic gene expression through differential DNA methylation established in the parental germ line. However, the mechanisms underlying how specific sequences are selectively methylated are not fully understood. We have found that the components of the PIWI-interacting RNA (piRNA) pathway are required for de novo methylation of the differentially methylated region (DMR) of the imprinted mouse Rasgrf1 locus, but not other paternally imprinted loci. A retrotransposon sequence within a noncoding RNA spanning the DMR was targeted by piRNAs generated from a different locus. A direct repeat in the DMR, which is required for the methylation and imprinting of Rasgrf1, served as a promoter for this RNA. We propose a model in which piRNAs and a target RNA direct the sequence-specific methylation of Rasgrf1.     

Asynchronous climate changes in the North Atlantic and Japan during the last termination

Pollen records from the annually laminated sediment sequence in Lake Suigetsu, Japan, suggest a sequence of climate changes during the Last Termination that resembles that of the North Atlantic region but with noticeable differences in timing. An interstadial interval commenced a few centuries earlier [approximately 15,000 years before the present (yr B.P.)] than the North Atlantic GI-1 (B?lling) event. Conversely, the onset of a Younger Dryas (YD)-like cold reversal (12,300 to 11,250 yr B.P.) postdated the North Atlantic GS-1 (YD) event by a few centuries. Climate in the Far East during the Last Termination reflected solar insolation changes as much as Atlantic influences.     

Magmatic dike resonances inferred from very-long-period seismic signals

Very-long-period (VLP) signals showing simple decaying harmonic oscillations with periods near 10 seconds and lasting for about 300 seconds were observed in association with an earthquake swarm that occurred beneath Hachijo Island, Japan. Results from the source-mechanism analysis and waveform simulation based on a fluid-filled crack model consistently point to the resonance of a dike filled with a basaltic magma as the source of the VLP signals. Thus, VLP signals can be used to probe the state of the fluid and dynamic processes within a volcanic system.     

A planar rhombic charge-separated tetrasilacyclobutadiene

The cyclobutadiene (CBD) molecule C(4)H(4) deviates from a high-symmetry square geometry to compensate for its antiaromatic electronic structure. Here, we report a CBD silicon analog, Si(4)(EMind)(4) (1), stabilized by the bulky 1,1,7,7-tetraethyl-3,3,5,5-tetramethyl-s-hydrindacen-4-yl (EMind) groups, obtained as air- and moisture-sensitive orange crystals by the reduction of (EMind)SiBr(3) with three equivalents of lithium naphthalenide. X-ray crystallography reveals a planar and rhombic structure of the Si(4) four-membered ring, with alternating pyramidal and planar configurations at the silicon atoms. The large (29)Si chemical shift differences (Δδ > 350 parts per million) in the solid-state nuclear magnetic resonance spectra suggest a contribution of an alternately charge-separated structure. The rhombic-shaped charge-separated singlet state of compound 1 thus stabilizes its cyclic 4π-electron antiaromaticity in a manner that contrasts sharply with the bond-length alternation, characterizing the rectangular distortion of carbon-based CBD.     

Essential role of Fkbp6 in male fertility and homologous chromosome pairing in meiosis

Meiosis is a critical stage of gametogenesis in which alignment and synapsis of chromosomal pairs occur, allowing for the recombination of maternal and paternal genomes. Here we show that FK506 binding protein (Fkbp6) localizes to meiotic chromosome cores and regions of homologous chromosome synapsis. Targeted inactivation of Fkbp6 in mice results in aspermic males and the absence of normal pachytene spermatocytes. Moreover, we identified the deletion of Fkbp6 exon 8 as the causative mutation in spontaneously male sterile as/as mutant rats. Loss of Fkbp6 results in abnormal pairing and misalignments between homologous chromosomes, nonhomologous partner switches, and autosynapsis of X chromosome cores in meiotic spermatocytes. Fertility and meiosis are normal in Fkbp6 mutant females. Thus, Fkbp6 is a component of the synaptonemal complex essential for sex-specific fertility and for the fidelity of homologous chromosome pairing in meiosis.     

Regulation of muscular glutamate metabolism by high‐protein diet in broiler chicks

The major taste active component, glutamate (Glu), improves the taste of meat. In this study, we investigated the effect of a short‐term high‐protein (HCP) diet on the intramuscular free Glu content to improve the taste of meat. Furthermore, we elucidated how the muscle free Glu content was controlled by the HCP diet. Chicks (14 days old) were fed the control diet or HCP diet for 10 days. Plasma and muscle free amino acid concentrations, and activity and messenger RNA (mRNA) expression of muscle enzymes related to Glu metabolism were determined. Muscle free Glu content was increased (P < 0.01) by 51%. Activity and mRNA expression of glutaminase (GA), which is one of the major Glu‐related enzymes, were significantly decreased (P < 0.05) in the HCP group because of feedback inhibition. The mRNA expression of lysine α‐ketoglutarate reductase (LKR), which is the enzyme involved in lysine (Lys) degradation and Glu production, was significantly increased (P < 0.001) in the HCP group. These results suggest that short‐term dietary HCP feeding is an effective treatment for improving the taste of meat. Furthermore, our results suggest that the free Glu content in muscle is regulated by GA and LKR.     

Plectosporium blight of monkshood caused by <Emphasis Type="Italic">Plectosporium tabacinum</Emphasis>

Severe blight of potted seedlings of monkshood caused by Plectosporium tabacinum was found in glasshouses in Kagawa Prefecture in southwest Japan in May 2001. Root rot and browning of stem bases were followed by wilting and yellowing of leaves, then blighting of leaves, flower buds and stems. A fungus was isolated from diseased plants and confirmed to cause the disease. The new disease was named “Plectosporium blight of monkshood”.