Background:Notch receptor has an important role in both development and cancer. We previously reported that inhibition of the Notch3 by γ-secretase inhibitor (GSI) induces apoptosis and suppresses tumour proliferation in non-small-cell lung cancer. Although radiation is reported to induce Notch activation, little is known about the relationship between radiation and Notch pathway.Methods:We examined the effect of combining GSI and radiation at different dosing in three Notch expressing lung cancer cell lines. The cytotoxic effect of GSI and radiation was evaluated using MTT assay and clonogenic assay in vitro and xenograft models. Expressions of Notch pathway, mitogen-activated protein kinase (MAPK) pathway and Bcl-2 family proteins were investigated using western blot analysis.Results:We discovered that the antitumour effect of combining GSI and radiation was dependent on treatment schedule. γ-Secretase inhibitor administration after radiation had the greatest growth inhibition of lung cancer in vitro and in vivo. We showed that the combination induced apoptosis of lung cancer cell lines through the regulation of MAPK and Bcl-2 family proteins. Furthermore, activation of Notch after radiation was ameliorated by GSI administration, suggesting that treatment with GSI prevents Notch-induced radiation resistance.Conclusion:Notch has an important role in lung cancer. Treatment with GSI after radiation can significantly enhance radiation-mediated tumour cytotoxicity.British Journal of Cancer advance online publication, 17 May 2012; doi:10.1038/bjc.2012.178 www.bjcancer.com.

ABSTACT: The fungus Tricholoma matsutake forms an ectomycorrhizal relationship with pine trees. Its sporocarps often develop in a circle, which is commonly known as a fairy ring. The fungus produces a solid, compact, white aggregate of mycelia and mycorrhizae beneath the fairy ring, which in Japanese is called a 'shiro'. In the present study, we used soil dilution plating and molecular techniques to analyze the bacterial communities within, beneath, and outside the T. matsutake fairy ring. Soil dilution plating confirmed previous reports that bacteria and actinomycetes are seldom present in the soil of the active mycorrhizal zone of the T. matsutake shiro. In addition, the results showed that the absence of bacteria was strongly correlated with the presence of T. matsutake mycorrhizae. The results demonstrate that bacteria, especially aerobic and heterotrophic forms, and actinomycetes, are strongly inhibited by T. matsutake. Indeed, neither bacteria nor actinomycetes were detected in 11.3% of 213 soil samples from the entire shiro area by culture-dependent methods. However, molecular techniques demonstrated that some bacteria, such as individual genera of Sphingomonas and Acidobacterium, were present in the active mycorrhizal zone, even though they were not detected in soil assays using the dilution plating technique.

Physicochemical characterization and structural evaluation of a 2:1 naproxen-nicotinamide cocrystal were performed. The 2:1 cocrystal showed rapid naproxen dissolution and less water vapor adsorption, indicating better pharmaceutical properties of naproxen. The unique 2:1 cocrystal formation was evaluated by solid-state nuclear magnetic resonance (NMR). The assignments of all H and (13) C peaks for naproxen and the cocrystal were performed using dipolar-insensitive nuclei enhanced by polarization transfer and (1) H-(13) C cross-polarization (CP)-heteronuclear correlation (HETCOR) NMR measurements. The (13) C chemical shift revealed that two naproxen molecules and one nicotinamide molecule existed in the asymmetric unit of the cocrystal. The (1) H chemical shifts indicated that the carboxylic group of the naproxen in the cocrystal was nonionized, and the CH-π interaction between naproxens was very strong. From the (1) H-(13) C CP-HETCOR NMR spectrum with contact time of 5 ms, two different synthons, carboxylic acid-amide and carboxylic acid-pyridine ring, were found between naproxen and nicotinamide. Single-crystal X-ray analysis, which supported the solid-state NMR results, clarified the geometry and intermolecular interactions in more detail. The structure is unique among pharmaceutical cocrystals because each carboxyl group of the two naproxens formed different intermolecular synthons. © 2012 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci.

Environmental metabolomics is an emerging field that is promoting new understanding in how organisms respond to and interact with the environment and each other at the biochemical level(1). Nuclear magnetic resonance (NMR) spectroscopy is one of several technologies, including gas chromatography-mass spectrometry (GC-MS), with considerable promise for such studies. Advantages of NMR are that it is suitable for untargeted analyses, provides structural information and spectra can be queried in quantitative and statistical manners against recently available databases of individual metabolite spectra(2,3). In addition, NMR spectral data can be combined with data from other omics levels (e.g. transcriptomics, genomics) to provide a more comprehensive understanding of the physiological responses of taxa to each other and the environment(4,5,6). However, NMR is less sensitive than other metabolomic techniques, making it difficult to apply to natural microbial systems where sample populations can be low-density and metabolite concentrations low compared to metabolites from well-defined and readily extractable sources such as whole tissues, biofluids or cell-cultures. Consequently, the few direct environmental metabolomic studies of microbes performed to date have been limited to culture-based or easily defined high-density ecosystems such as host-symbiont systems, constructed co-cultures or manipulations of the gut environment where stable isotope labeling can be additionally used to enhance NMR signals(7,8,9,10,11,12). Methods that facilitate the concentration and collection of environmental metabolites at concentrations suitable for NMR are lacking. Since recent attention has been given to the environmental metabolomics of organisms within the aquatic environment, where much of the energy and material flow is mediated by the planktonic community(13,14), we have developed a method for the concentration and extraction of whole-community metabolites from planktonic microbial systems by filtration. Commercially available hydrophilic poly-1,1-difluoroethene (PVDF) filters are specially treated to completely remove extractables, which can otherwise appear as contaminants in subsequent analyses. These treated filters are then used to filter environmental or experimental samples of interest. Filters containing the wet sample material are lyophilized and aqueous-soluble metabolites are extracted directly for conventional NMR spectroscopy using a standardized potassium phosphate extraction buffer(2). Data derived from these methods can be analyzed statistically to identify meaningful patterns, or integrated with other omics levels for comprehensive understanding of community and ecosystem function.

A statistical approach was used to characterize the heterogeneous structures of bacterial cellulose samples pretreated with four kinds of ionic liquids (ILs). The structural heterogeneity of these samples was measured by Fourier transform infrared spectroscopy, as well as solid-state NMR methods such as cross-polarization magic angle spinning and dipolar-assisted rotational resonance. The obtained data matrices were then evaluated by principal components analysis. The measured one-dimensional data clearly revealed the modification of crystalline cellulose; in addition, the statistical approach revealed subtle structural changes that occurred upon pretreatment with different kinds of ILs. To investigate whether such regenerated structural changes occurred because of solubilization, the intermolecular nuclear Overhauser effect (NOE) between cellulose and an IL was examined. Our results clarify how the nucleophilic imidazole is attacked, and suggest that the cation of the IL is associated with the collapse of hydrogen bonds in cellulose.

Ecosystems can be conceptually thought of as interconnected environmental and metabolic systems, in which small molecules to macro-molecules interact through diverse networks. State-of-the-art technologies in post-genomic science offer ways to inspect and analyze this biomolecular web using omics-based approaches. Exploring useful genes and enzymes, as well as biomass resources responsible for anabolism and catabolism within ecosystems will contribute to a better understanding of environmental functions and their application to biotechnology. Here we present ECOMICS, a suite of web-based tools for ECosystem trans-OMICS investigation that target metagenomic, metatranscriptomic, and meta-metabolomic systems, including biomacromolecular mixtures derived from biomass. ECOMICS is made of four integrated webtools. E-class allows for the sequence-based taxonomic classification of eukaryotic and prokaryotic ribosomal data and the functional classification of selected enzymes. FT2B allows for the digital processing of NMR spectra for downstream metabolic or chemical phenotyping. Bm-Char allows for statistical assignment of specific compounds found in lignocellulose-based biomass, and HetMap is a data matrix generator and correlation calculator that can be applied to trans-omics datasets as analyzed by these and other web tools. This web suite is unique in that it allows for the monitoring of biomass metabolism in a particular environment, i.e., from macromolecular complexes (FT2DB and Bm-Char) to microbial composition and degradation (E-class), and makes possible the understanding of relationships between molecular and microbial elements (HetMap). This website is available to the public domain at: https://database.riken.jp/ecomics/.

A supramolecular system that can activate an enzyme through photo-isomerization was constructed by using a liposomal membrane scaffold. The design of the system was inspired by natural signal transduction systems, in which enzymes amplify external signals to control signal transduction pathways. The liposomal membrane, which provided a scaffold for the system, was prepared by self-assembly of a photoresponsive receptor and a cationic synthetic lipid. NADH-dependent L-lactate dehydrogenase, the signal amplifier, was immobilized on the liposomal surface by electrostatic interactions. Recognition of photonic signals by the membrane-bound receptor induced photo-isomerization, which significantly altered the receptor's metal-binding affinity. The response to the photonic signal was transmitted to the enzyme by Cu(2+) ions. The enzyme amplified the chemical information through a catalytic reaction to generate the intended output signal.

A comprehensive and large scale metabolome quantitative trait loci (mQTL) analysis was performed to investigate the genetic backgrounds associated with metabolic phenotypes in rice grains. The metabolome dataset consisted of 759 metabolite signals was obtained from the grains of 85 lines of rice (Oryza sativa, Sasanishiki × Habataki back-crossed inbred lines). Metabolome analysis was performed using 4 different mass spectrometry pipelines to enhance detection of different classes of metabolites. mQTL analysis of a wide range of metabolites highlighted an uneven distribution of 802 mQTLs around the rice genome, as well as different modes of metabolomic trait (m-trait) control among various types of metabolites. The levels of most metabolites within rice grains were highly sensitive to environmental factors, but only weakly associated with mQTLs. Coordinated control was observed for several groups of metabolites, such as amino acids that linked to the mQTL hotspot on chromosome 3. For flavonoids, m-trait variation among the experimental lines was tightly governed by genetic factors that alter the glycosylation of flavones. Many loci affecting m-trait levels were detected by QTL analysis and plausible gene candidates were evaluated by in silico analysis. There were several mQTLs that profoundly influenced metabolite levels, providing insight into the control of rice metabolism. The genomic region and genes potentially responsible for the biosynthesis of apigenin-6,8-di-C-α-l-arabinoside are presented as an example of a critical mQTL identified by the analysis. © 2012 The Authors. The Plant Journal© 2012 Blackwell Publishing Ltd.

Lipid-membrane-incorporating C(60) and C(70) (LMIC(60) and LMIC(70)) were prepared by the fullerene-exchange reaction from the γ-cyclodextrin cavity to vesicles (we call this method the "exchange method"). An advantage of this method is that the ratios of [C(60)]/[lipids] and [C(70)]/[lipids] can be arbitrarily controlled by adjusting the ratios of the fullerenes and liposome. The maximum ratio (30 mol %) obtained was approximately 14 and 100 times higher than those achieved for LMIC(60) and LMIC(70), respectively, that were prepared by the classical method, which we call the "premixing method"(dissolving lipids and C(60) or C(70) in chloroform, followed by concentration and extraction with water). Furthermore, the stabilities and photodynamic activities of the LMIC(60) and LMIC(70) solutions prepared by the exchange method were shown to be much higher than those prepared by the premixing method. That is, the exchange method was found to be superior to the premixing method as a preparative method of LMIC(60) and LMIC(70) for applications in photomedical and photomaterials chemistry.

Relatively little is known about the regulatory mechanisms of the Drosha/DGCR8 complex, which processes miRNAs at the initial step of biogenesis. We found that histone deacetylase 1 (HDAC1) increases the expression levels of mature miRNAs despite repressing the transcription of host genes. HDAC1 is an integral component of the Drosha/DGCR8 complex and enhances miRNA processing by increasing the affinity of DGCR8 to primary miRNA transcripts via deacetylation of critical lysine residues in the RNA-binding domains of DGCR8. This finding suggests that HDACs have two arms for gene silencing: transcriptional repression by promoter histone deacetylation and post-transcriptional inhibition by increasing miRNA abundance.