A diterpene quinone from the bark of Cryptomeria japonica

A diterpene, cryptoquinone, was isolated from the bark of Cryptomeria japonica, the structure, 7,11,14-trioxoabieta-8,12-diene, was established by spectral analyses and X-ray crystallography. This diterpene quinone showed moderate antifungal activities against Pyricularia orizae and Alternaria alternata, and cytotoxic activity against mouse lymphoid neoplasm (P388) cells with an IC(50) of 0.26 microg/ml.     

Probing the interaction between N(1),N(4)-dibenzylputrescine and tRNA through (15)N NMR: biological implications

NMR spectroscopy was used to characterize the binding properties of polyamines to Escherichia coli tRNA. The (15)N NMR spectra of three (15)N-enriched N-substituted putrescine derivatives (DMP, DEP and DBP) were recorded in the presence of tRNA, and the spin relaxation times of the nitrogen nuclei were measured. From these data, the activation parameters for the rotational correlation times of the (15)N nuclei were determined. The present data indicate that the nature of the amino substituents does play a relevant role in controlling the polyamine-tRNA interaction. This study also provides a rationale for the in vivo antiproliferative effect of DBP against tumoral cells.     

Substrate recognition by glycoside hydrolase family 74 xyloglucanase from the basidiomycete Phanerochaete chrysosporium

The basidiomycete Phanerochaete chrysosporium produces xyloglucanase Xgh74B, which has the glycoside hydrolase (GH) family 74 catalytic domain and family 1 carbohydrate-binding module, in cellulose-grown culture. The recombinant enzyme, which was heterologously expressed in the yeast Pichia pastoris, had high hydrolytic activity toward xyloglucan from tamarind seed (TXG), whereas other beta-1,4-glucans examined were poor substrates for the enzyme. The existence of the carbohydrate-binding module significantly affects adsorption of the enzyme on crystalline cellulose, but has no effect on the hydrolysis of xyloglucan, indicating that the domain may contribute to the localization of the enzyme. HPLC and MALDI-TOF MS analyses of the hydrolytic products of TXG clearly indicated that Xgh74B hydrolyzes the glycosidic bonds of unbranched glucose residues, like other GH family 74 xyloglucanases. However, viscometric analysis suggested that Xgh74B hydrolyzes TXG in a different manner from other known GH family 74 xyloglucanases. Gel permeation chromatography showed that Xgh74B initially produced oligosaccharides of degree of polymerization (DP) 16-18, and these oligosaccharides were then slowly hydrolyzed to final products of DP 7-9. In addition, the ratio of oligosaccharides of DP 7-9 versus those of DP 16-18 was dependent upon the pH of the reaction mixture, indicating that the affinity of Xgh74B for the oligosaccharides of DP 16-18 is affected by the ionic environment at the active site.     

Three distinct stages of apoptotic nuclear condensation revealed by time-lapse imaging, biochemical and electron microscopy analysis of cell-free apoptosis

During apoptotic execution, chromatin undergoes a phase change from a heterogeneous, genetically active network to an inert highly condensed form that is fragmented and packaged into apoptotic bodies. We have previously used a cell-free system to examine the roles of caspases or other proteases in apoptotic chromatin condensation and nuclear disassembly. But so far, the role of DNase activity or ATP hydrolysis in this system has not yet been elucidated. Here, in order to better define the stages of nuclear disassembly in apoptosis, we have characterized the apoptotic condensation using a cell-free system and time-lapse imaging. We demonstrated that the population of nuclei undergoing apoptosis in vitro appears to follow a reproducible program of nuclear condensation, suggesting the existence of an ordered biochemical pathway. This enabled us to define three stages of apoptotic chromatin condensation: stage 1 ring condensation; stage 2 necklace condensation; and stage 3 nuclear collapse/disassembly. Electron microscopy revealed that neither chromatin nor detectable subnuclear structures were present inside the stage 1 ring-condensed structures. DNase activity was not essential for stage 1 ring condensation, which could occur in apoptotic extracts depleted of all detectable DNase activity. However, DNase(s) were required for stage 2 necklace condensation. Finally, we demonstrated that hydrolyzable ATP is required for stage 3 nuclear collapse/disassembly. This requirement for ATP hydrolysis further distinguished stage 2 from stage 3. Together, these experiments provide the first steps towards a systematic biochemical characterization of chromatin condensation during apoptosis.     

Can biological nitrification inhibition (BNI) genes from perennial Leymus racemosus (Triticeae) combat nitrification in wheat farming?

Using a recombinant luminescent Nitrosomonas europaea assay to quantify biological nitrification inhibition (BNI), we found that a wild relative of wheat (Leymus racemosus (Lam.) Tzvelev) had a high BNI capacity and releases about 20 times more BNI compounds (about 30 ATU g⁻¹ root dry weight 24 h⁻¹) than Triticum aestivum L. (cultivated wheat). The root exudate from cultivated wheat has no inhibitory effect on nitrification when applied to soil; however, the root exudate from L. racemous suppressed formation and kept more than 90% of the soil’s inorganic-N in the -form for 60 days. The high-BNI capacity of L. racemosus is mostly associated with chromosome Lr#n. Two other chromosomes Lr#J, and Lr#I also have an influence on BNI production. Tolerance of L. racemosus to is controlled by chromosome 7Lr#1-1. Sustained release of BNI compounds occurred only in the presence of in the root environment. Given the level of BNI production expressed in DALr#n and assuming normal plant growth, we estimated that nearly 87,500,000 ATU of BNI activity ha⁻¹ day⁻¹ could be released in a field of vigorously growing wheat; this amounts to the equivalent of the inhibitory effect from the application of 52.5 g of the synthetic nitrification inhibitor nitrapyrin (one AT unit of BNI activity is equivalent to 0.6 μg of nitrapyrin). At this rate of BNI production it would take only 19 days for a BNI-enabled wheat crop to produce the inhibitory power of a standard commercial application of nitrapyrin, 1 kg ha⁻¹. The synthetic nitrification inhibitor, dicyandiamide, blocked specifically the AMO (ammonia monooxygenase) pathway, while the BNI from L. racemosus blocked the HAO (hydroxylamine oxidoreductase) pathway in Nitrosomonas. Here we report the first finding of high production of BNI in a wild relative of any cereal and its successful introduction and expression in cultivated wheat. These results demonstrate the potential for empowering the new generation of wheat cultivars with high-BNI capacity to control nitrification in wheat-production systems. Responsible Editor: Hans Lambers.     

The Inner Centromere Protein (INCENP) Coil Is a Single α-Helix (SAH) Domain That Binds Directly to Microtubules and Is Important for Chromosome Passenger Complex (CPC) Localization and Function in Mitosis

The chromosome passenger complex (CPC) is a master regulator of mitosis. Inner centromere protein (INCENP) acts as a scaffold regulating CPC localization and activity. During early mitosis, the N-terminal region of INCENP forms a three-helix bundle with Survivin and Borealin, directing the CPC to the inner centromere where it plays essential roles in chromosome alignment and the spindle assembly checkpoint. The C-terminal IN box region of INCENP is responsible for binding and activating Aurora B kinase. The central region of INCENP has been proposed to comprise a coiled coil domain acting as a spacer between the N- and C-terminal domains that is involved in microtubule binding and regulation of the spindle checkpoint. Here we show that the central region (213 residues) of chicken INCENP is not a coiled coil but a ∼32-nm-long single α-helix (SAH) domain. The N-terminal half of this domain directly binds to microtubules in vitro. By analogy with previous studies of myosin 10, our data suggest that the INCENP SAH might stretch up to ∼80 nm under physiological forces. Thus, the INCENP SAH could act as a flexible “dog leash,” allowing Aurora B to phosphorylate dynamic substrates localized in the outer kinetochore while at the same time being stably anchored to the heterochromatin of the inner centromere. Furthermore, by achieving this flexibility via an SAH domain, the CPC avoids a need for dimerization (required for coiled coil formation), which would greatly complicate regulation of the proximity-induced trans-phosphorylation that is critical for Aurora B activation.     

Efficient reinforcement learning: computational theories, neuroscience and robotics

Reinforcement learning algorithms have provided some of the most influential computational theories for behavioral learning that depends on reward and penalty. After briefly reviewing supporting experimental data, this paper tackles three difficult theoretical issues that remain to be explored. First, plain reinforcement learning is much too slow to be considered a plausible brain model. Second, although the temporal-difference error has an important role both in theory and in experiments, how to compute it remains an enigma. Third, function of all brain areas, including the cerebral cortex, cerebellum, brainstem and basal ganglia, seems to necessitate a new computational framework. Computational studies that emphasize meta-parameters, hierarchy, modularity and supervised learning to resolve these issues are reviewed here, together with the related experimental data.     

Lactoferrin as a suppressor of cell migration of gastrointestinal cell lines

The effects of lactoferrin (Lf), an iron-binding glycoprotein, on cell migration were investigated. Lf inhibited the cell migration of three gastrointestinal cell lines (Caco-2 cells, AGS cells, and IEC-18 cells) in vitro. Both iron-saturated (holo) and iron-depleted (apo) Lf showed this inhibitory effect. Chelation of iron in the culture medium by desferrioxamine did not affect the activity of either form of Lf. A pepsin hydrolysate of Lf exhibited effectiveness similar to that of intact Lf. These results demonstrate a novel activity of Lf and suggest a potential role for this molecule in gastrointestinal wound healing, which is independent of its iron-binding capacity. J. Cell. Physiol. 170:101–105, 1997. © 1997 Wiley-Liss, Inc.     

Endo-beta-1,3-galactanase from winter mushroom Flammulina velutipes

Arabinogalactan proteins are proteoglycans found on the cell surface and in the cell walls of higher plants. The carbohydrate moieties of most arabinogalactan proteins are composed of β-1,3-galactan main chains and β-1,6-galactan side chains, to which other auxiliary sugars are attached. For the present study, an endo-β-1,3-galactanase, designated FvEn3GAL, was first purified and cloned from winter mushroom Flammulina velutipes. The enzyme specifically hydrolyzed β-1,3-galactan, but did not act on β-1,3-glucan, β-1,3:1,4-glucan, xyloglucan, and agarose. It released various β-1,3-galactooligosaccharides together with Gal from β-1,3-galactohexaose in the early phase of the reaction, demonstrating that it acts on β-1,3-galactan in an endo-fashion. Phylogenetic analysis revealed that FvEn3GAL is member of a novel subgroup distinct from known glycoside hydrolases such as endo-β-1,3-glucanase and endo-β-1,3:1,4-glucanase in glycoside hydrolase family 16. Point mutations replacing the putative catalytic Glu residues conserved for enzymes in this family with Asp abolished activity. These results indicate that FvEn3GAL is a highly specific glycoside hydrolase 16 endo-β-1,3-galactanase.