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Basal plant immune responses are activated by the recognition of conserved microbe‐associated molecular patterns (MAMPs), or breakdown molecules released from the plants after damage by pathogen penetration, so‐called damage‐associated molecular patterns (DAMPs). While chitin‐oligosaccharide (CHOS), a primary component of fungal cell walls, is most known as MAMP, plant cell wall‐derived oligosaccharides, cello‐oligosaccharides (COS) from cellulose, and xylo‐oligosaccharide (XOS) from hemicellulose are representative DAMPs. In this study, elicitor activities of COS prepared from cotton linters, XOS prepared from corn cobs, and chitin‐oligosaccharide (CHOS) from crustacean shells were comparatively investigated. In Arabidopsis, COS, XOS, or CHOS treatment triggered typical defense responses such as reactive oxygen species (ROS) production, phosphorylation of MAP kinases, callose deposition, and activation of the defense‐related transcription factor WRKY33 promoter. When COS, XOS, and CHOS were used at concentrations with similar activity in inducing ROS production and callose depositions, CHOS was particularly potent in activating the MAPK kinases and WRKY33 promoters. Among the COS and XOS with different degrees of polymerization, cellotriose and xylotetraose showed the highest activity for the activation of WRKY33 promoter. Gene ontology enrichment analysis of RNAseq data revealed that simultaneous treatment of COS, XOS, and CHOS (oligo‐mix) effectively activates plant disease resistance. In practice, treatment with the oligo‐mix enhanced the resistance of tomato to powdery mildew, but plant growth was not inhibited but rather tended to be promoted, providing evidence that treatment with the oligo‐mix has beneficial effects on improving disease resistance in plants, making them a promising class of compounds for practical application. [ABSTRACT FROM AUTHOR]

ABCB1, also known as P‐glycoprotein, is an essential component of many physiological barriers and extrudes a variety of hydrophobic chemicals out of the cell. Structures of ABCB1 provided insights into the structural changes that occur upon ATP binding and the characteristic architecture of the substrate binding site. Yet, the structure–function relationship between substrate binding and transporting still remains largely obscured because there is no robust method for accurately measuring substrate binding constants. The methods currently used cannot identify whether the bound substrates are located in the inner chamber of the molecule in the transmembrane region or not because of the low spatial resolution. Here, we report a system for measuring the affinity of substrate binding to the Cyanidioschyzon merolae ABCB1 (CmABCB1) using site‐specific tryptophan (Trp) fluorescence quenching. We designed a CmABCB1 mutant with an extrinsic Trp residue introduced into the inner chamber. Trp fluorescence was quenched by three substrates and one inhibitor, including rhodamine 6G, in a saturable fashion, allowing for accurate estimation of the dissociation constant (KD) for each molecule. The KD for rhodamine 6G is similar to that determined using a reciprocal fluorescence quenching assay using rhodamine 6G fluorescence, suggesting that Trp fluorescence of the mutant was quenched by the interaction between the extrinsic Trp and substrates bound in the inner chamber. Structural comparison of the ABCB1 structures suggests that the system presented in this study could be ideal method of choice to determine the substrate binding affinities of compounds bound to the chamber of mammalian ABCB1. PDB Code(s): 7VR5; [ABSTRACT FROM AUTHOR]

Introduction: Control of postoperative gastroesophageal reflux (GER) is a critical consideration for patients who undergo proximal gastrectomy. This report describes a new and simple reconstruction method by esophagogastrostomy with placement of the remnant stomach into the lower mediastinum (EG‐PRIME). This approach not only suppresses postoperative GER, but it can also be easily performed by total laparoscopy. Detailed herein are the methods for EG‐PRIME and the results for three patients with cancer of the upper gastric body. Materials and Surgical Technique: At the start of the EG‐PRIME surgical procedure, a pseudo‐fornix and lozenge‐shaped gastric conduit were made by cutting the stomach diagonally. Next, the pseudo‐fornix was inserted into the esophageal hiatus to form a new angle of His. Then overlap anastomosis was performed and the entry site was closed longitudinally. The outcomes assessed were operative time, intraoperative blood loss, postoperative complications and GER according to 24‐hour pH monitoring. Discussion: The operative times were 339, 288 and 236 minutes; in two patients, intraoperative blood loss was 260 and 343 mL, and in the third, blood loss was minimal. No postoperative complications were observed in any of the three patients. The degree of the GER resulting in fraction time pH<4 was 9.0%, 0.3%, and 2.9%, respectively. No esophagitis by upper gastrointestinal endoscopy was observed in any patient. This EG‐PRIME method was technically feasible for reconstruction after proximal gastrectomy by total laparoscopy. This approach may be as simple and useful as esophagogastrostomy for preventing postoperative GER, but more experience with this method is required. [ABSTRACT FROM AUTHOR]

CmABCB1 is a homologue of human P‐glycoprotein, which extrudes various substrates by iterative cycles of conformational changes between the inward‐ and outward‐facing states. Comparison of the inward‐ and outward‐facing structures of CmABCB1 suggested that pivotal joints in the transmembrane domain regulate the tilt of transmembrane helices. Transmembrane helix 1 (TM1) forms a tight helix–helix contact with TM3 at the TM1–3 joint. Mutation of Gly132 to valine at the TM1–3 joint, G132V, caused a 10‐fold increase in ATPase activity, but the mechanism underlying this change remains unclear. Here, we report a crystal structure of the outward‐facing state of the CmABCB1 G132V mutant at a 2.15 Å resolution. We observed structural displacements between the outward‐facing states of G132V and the previous one at the region around the TM1–3 joint, and a significant expansion at the extracellular gate. We hypothesize that steric hindrance caused by the Val substitution shifted the conformational equilibrium toward the outward‐facing state, favoring the dimeric state of the nucleotide‐binding domains and thereby increasing the ATPase activity of the G132V mutant. PDB Code(s): 7DQV; [ABSTRACT FROM AUTHOR]

ABCB1, also called MDR1 or P‐glycoprotein, exports various hydrophobic compounds and plays an essential role as a protective physiological barrier in several organs, including the brain, testis, and placenta. However, little is known about the structural mechanisms that allow ABCB1 to recognize hydrophobic compounds of diverse structures or the coupling of ATP hydrolysis to uphill substrate export. High‐resolution X‐ray crystal structures of the pre‐ and post‐transport states and FRET analyses in living cells have revealed that an aromatic hydrophobic network at the top of the inner cavity is key for the conformational change in ABCB1 that is triggered by a hydrophobic substrate. ATP binding, but not hydrolysis, induces a progressive network that results in a twisting motion of the whole protein, squeezing out the substrate directly to the extracellular space. This twist‐and‐squeeze mechanism by which ABCB1 exports hydrophobic substrates is distinct from those of other transporters. [ABSTRACT FROM AUTHOR]

Members of the ATP‐binding cassette (ABC) transporter superfamily translocate a broad spectrum of chemically diverse substrates. While their eponymous ATP‐binding cassette in the nucleotide‐binding domains (NBDs) is highly conserved, their transmembrane domains (TMDs) forming the translocation pathway exhibit distinct folds and topologies, suggesting that during evolution the ancient motor domains were combined with different transmembrane mechanical systems to orchestrate a variety of cellular processes. In recent years, it has become increasingly evident that the distinct TMD folds are best suited to categorize the multitude of ABC transporters. We therefore propose a new ABC transporter classification that is based on structural homology in the TMDs. [ABSTRACT FROM AUTHOR]

Aims/Introduction: Low birthweight is reportedly associated with type 2 diabetes mellitus; however, this association has not been confirmed in the Japanese population, and whether high birthweight is associated with type 2 diabetes mellitus is controversial. We aimed to investigate the association between birthweight and type 2 diabetes mellitus among a general Japanese population. Materials and Methods: Overall 1,135 middle‐ to old‐aged Japanese men and women were enrolled in the Toon Health Study. A 75‐g oral glucose tolerance test was used to diagnose type 2 diabetes mellitus, and a questionnaire survey about birthweight was administered. The association between birthweight and the prevalence of type 2 diabetes mellitus in later life of the participants was examined using multivariable logistic regression analysis. Stratified analysis by current body mass index was also carried out. Results: The mean age was 56.5 ± 12.2 years. Type 2 diabetes mellitus was observed in 9.3% of the participants in this study. Compared with the reference group (2,500–3,999 g), the adjusted odds ratio of the low‐birthweight group (<2,500 g) for type 2 diabetes mellitus was 2.46 (95% confidence interval 1.48–4.10). The association between the high‐birthweight group (≥4000 g) and type 2 diabetes mellitus was not significant after including family history of diabetes in the multivariable model. The odds ratio of the low‐birthweight group for type 2 diabetes mellitus was higher in the overweight/obese group than in the non‐overweight group. Conclusions: Low birthweight was associated with an increased risk of type 2 diabetes mellitus in a Japanese population, especially in overweight/obese individuals. [ABSTRACT FROM AUTHOR]

Abstract: Spatial variability of throughfall (TF) isotopic composition, used as tracer input, influences isotope hydrological applications in forested watersheds. Notwithstanding, identification of the dominant canopy factors and processes that affect the patterns of TF isotopic variability remains ambiguous. Here, we examined the spatio‐temporal variability of TF isotopic composition in a Japanese cypress plantation, in which intensive strip thinning was performed and investigated whether canopy structure at a fine resolution of canopy effect analysis is related to TF isotopic composition and how this is affected by meteorological factors. Canopy openness, as an index of canopy structure, was calculated from hemispherical photographs at different zenith angles. TF samples were collected in a 10 × 10 m experimental plot in both pre‐thinning (from July to November 2010) and post‐thinning (from May 2012 to March 2013) periods. Our results show that thinning resulted in a smaller alteration of input δ18O of gross precipitation, whereas the changes in deuterium excess varied in both directions. Despite the temporal stability of spatial patterns in TF amount, the spatial variability of TF isotopic composition was not temporally stable in both pre‐ and post‐thinning periods. Additionally, after thinning, the isotopic composition of TF was best related to canopy openness calculated at the zenith angle of 7°, exhibiting three different relationships, that is, significantly negative, significantly positive, and nonsignificant. Changes in meteorological factors (wind speed, rainfall intensity, and temperature) were found to affect the relationships between TF δ18O and canopy openness. The observed shifts in the relationships reveal different dominant factors (partial evaporation and the selection), and canopy water flowpaths control such differences. This study provides useful insights into the spatial variability of TF isotopic composition and improves our understanding of the physical processes of interception through canopy passage. [ABSTRACT FROM AUTHOR]

This study investigated the effects of plowing on the vertical migration of radioactive Cs, air dose rate, and soil physicochemical properties in temperate pastures after radioactive pollution from the 2011 Fukushima Daiichi Nuclear Power Plant accident. The study area consisted of three plowed pastures (decontamination treatment) and one non-plowed pasture at the Kawatabi Field Science Center of Tohoku University, 150 km north of the nuclear power plant. Air dose rate was measured at the top, middle, and bottom of the slope of each pasture. A profile pit (1 m × 3 m) was excavated at each location. Plants, litter and soil samples (from 0 cm to the parent material layer) were collected from all locations during the summer of 2014 to determine the radioactive Cs concentration. Particle size distribution and bulk density of the soil samples were also measured. Air dose rate was higher on the non-plowed (0.090-0.097 μSv h−1) than that on the plowed (0.047-0.073 μSv h−1) pastures. Radioactive Cs concentration was high in the litter layer (207-475 Bq kg−1 dry matter [DM]) and the topsoil (0-2.5 cm depth; 412-1139 Bq kg−1 DM) in the non-plowed pasture. In the plowed pastures, these variables did not differ within the 0-40 cm layer. In the non-plowed pasture, there were high sand (37.0%) and low silt (61.4%) proportions at 10-20 cm and the topsoil had a high bulk density. In the plowed pastures, these variables were similar within the 0-20 cm layer. The bulk density ( P < 0.001) and organic matter content ( P < 0.05) had significant relationships with the concentration of radioactive Cs. These results indicate that plowing reduced air dose rate. [ABSTRACT FROM AUTHOR]

This study revealed the influence of change in forest canopy structure by forest management practice for throughfall generation and canopy water interception process. Throughfall and stemflow were observed in a mature Japanese cypress ( Chamaecyparis obtusa) plantation before and after thinning. Decreased stem density (from 2400 to 1300 trees ha−1) and basal area (from 107.6 to 60.4 m2 ha−1) resulted in an increase in the throughfall fraction (from 58% to 79%) and decreases in the stemflow fraction (from 14% to 8.6%) and the canopy interception loss fraction (from 27% to 12%). Thinning also decreased spatial variability in throughfall and smoothed temporal persistence of throughfall among sampling points or rainfall events. These results were supported by the change in the throughfall balance between direct throughfall and released throughfall. The proportion of direct throughfall of incident rainfall changed from 0.16 to 0.42 owing to an increase in canopy gap. The distribution ratio of released throughfall from intercepted rainwater by canopy changed from 50% to 64% because of a decrease in foliage; this led to lower water storage in the canopy (from 1.83 to 0.82 mm), and thus less precipitation was needed to saturate the canopy. Consequently, the amount of throughfall increased and less precipitation was needed to stabilize the water flow path during the post-thinning period. This study contributes to knowledge of differential canopy wetting processes and spatiotemporal heterogeneity of throughfall distribution in response to specific silvicultural prescription. Copyright © 2015 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Nutrient concentrations in stream water, rainfall, throughfall, stemflow, surface flow and ground water were compared before, during and after strip thinning (intensive 50%) in plantation forested watersheds in Tochigi, Japan. Influences were evaluated comparing four thinning-applied and two reference basins for 1 year before, 6 months during and 1 year after the thinning. Results show that this strip thinning significantly increased dissolved total phosphorus, total phosphorus and dissolved organic carbon (0.01, 0.04 and 0.53 mg l-1, respectively) during the thinning period and dissolved total nitrogen and total nitrogen (0.34 and 0.46 mg l-1, respectively) after the thinning in stream waters relative to the unthinned basins. The increased phosphorus during thinning indicated ground disturbances by the strip thinning, with a concomitant increase in dissolved organic carbon. Changes in biotic and abiotic conditions resulted in increased nitrogen after the thinning, particularly in the dissolved pool. Changes in hydrological processes due to thinning, for example, a change in flow distributions (less high nutrient stemflow and more low nutrient throughfall) and an increase in water discharge in stream water, possibly weakened the direct influences of thinning on nutrient concentrations. [ABSTRACT FROM AUTHOR]

We evaluated the effects of 50% strip thinning on peak flow responses in 17-ha and 9-ha nested catchments. We observed 117 storm events from April 2010 to December 2013, including 32 events in the pre-thinning period, 15 events during the thinning operation, and 70 events in the post-thinning period. Paired-catchment analysis with multiple pairs of nested gauging stations revealed that changes in peak flow responses varied depending on the location of the nested observation stations. Increases in peak flow during the operation period were significant in gauging stations located at the outlet and the middle of a catchment. Increases in peak flow during the post-treatment period became significant in all nested catchments, but not in the catchment outlet. Rates of increase in peak flow ranged from 17.7% to 44.4%, which is within the range of previous studies. Different responses for changes in peak flow among nested pairs were associated with changes in internal hydrological flow pathways due to removal of timber and hydrological connectivity via overland flow generated by newly installed and reactivated skid trails. Our findings suggest that the effects of timber harvesting on peak flow are scale-dependent with respect to drainage areas, hydrological links, and management practices. Selection of gauging locations and process-based observations within catchments are essential for testing hypotheses related to the effects of timber harvesting with robust hydrological monitoring. [ABSTRACT FROM AUTHOR]

We examined the effect of strip thinning on the spatio-temporal variability of throughfall (TF) in a mature Japanese cypress (Chamaecyparis obtusa) stand. Strip thinning, which removed 50% of stems, was performed in a catchment in October 2011. TF was examined by setting up 20 tipping-bucket rain gauges in an experimental plot. Canopy interception parameters were derived from high-temporal-resolution TF data. Observations were obtained during the growing season in the pre-thinning and postthinning periods. The results demonstrated that after thinning, the TF rate increased from 64.3 to 76.2%, and the mean coefficient of variation of the TF rate significantly decreased from 27.3 ± 9.7 to 22.5 ± 6.8%. The spatial TF patterns were largely related to the rainfall class. There was no significant difference in the TF rate for the rainfall amount >25mm between the periods. The temporal persistence of the normalized TF was slightly stable in the post-thinning period and was not significantly related to canopy cover or the distance to the nearest trunk of each rain gauge in either period. Additionally, thinning resulted in an increase in the proportion of direct TF (from 0.14 ± 0.07 to 0.33 ± 0.11) and a decrease in canopy water storage (from 2.4 ± 0.7 to 1.5 ± 0.5 mm). The quantification of the canopy interception parameters can aid in modelling the changes in interception loss and its processes induced by thinning. These findings also help recognize the changes in spatial TF patterns under different forest management practices. [ABSTRACT FROM AUTHOR]

Varicella zoster virus ( VZV) is the etiologic agent of varicella, and it remains common among children in Japan due to low vaccination rates. It can cause a variety of serious and life-threatening complications. Generally, the most frequent complication of varicella in healthy children is bacterial superinfection, but empyema after VZV infection is a rare condition. This case report describes a previously healthy 21-month-old boy who attended nursery school with a recent varicella and group A β-hemolytic streptococcus ( GABHS) pharyngitis outbreak and who presented with a 7 day history of vesicular rash along with progressive fever. Due to continued mild cough and prolonged fever, however, chest radiography was done, which showed a right pleural effusion. Further computed tomography showed a right pulmonary empyema, and purulent material was drained and eventually grew GABHS. This report hereby describes the development of pleural empyema caused by GABHS after VZV infection in a serologically immunocompetent patient. [ABSTRACT FROM AUTHOR]

ABSTRACT We investigated the contribution of three evapotranspiration (ET) components including canopy interception ( Ei), tree transpiration ( Et) and forest floor evaporation ( Ef) and identified the sources depth of evaporated water in a Japanese cypress stand. Monitoring primarily focused on the growing season of July-October 2011. In a 12 × 13 m plot with 28 trees with a DBH of 19.1 ± 3.9 cm, Ei was calculated as the difference between precipitation on an open field and precipitation under a canopy as the sum of throughfall and stemflow, and Et was measured using the Granier method. Ef was measured by weighing lysimeters. Total ET during the monitoring period was 446.3 mm, accounting for 47.5% of the total precipitation of 938.8 mm. Ei was the dominant evaporation flux and accounted for 53.6% of ET or 25.5% of rainfall, followed by Et with 33.7% of ET or 16.0% of rainfall. The average Et was 1.5 ± 0.6 mm d−1. It was well correlated with soil moisture at a depth of 5-15 cm, which reflected the forest properties, i.e. tree roots were exposed on the forest floor. Ef accounted for 12.7% of ET or 6.0% of rainfall with a daily mean of 0.55 ± 0.26 mm d−1, and was best correlated with soil moisture in the upper 5 cm of soil. These results can improve the understanding of water budget in forested watershed and also be used to build and validate hydrologic models for water resource managements. Copyright © 2013 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Peroxisomes require peroxin (Pex) proteins for their biogenesis. The interaction between Pex3p, which resides on the peroxisomal membrane, and Pex19p, which resides in the cytosol, is crucial for peroxisome formation and the post-translational targeting of peroxisomal membrane proteins (PMPs). It is not known how Pex3p promotes the specific interaction with Pex19p for the purpose of PMP translocation. Here, we present the three-dimensional structure of the complex between a cytosolic domain of Pex3p and the binding-region peptide of Pex19p. The overall shape of Pex3p is a prolate spheroid with a novel fold, the 'twisted six-helix bundle.' The Pex19p-binding site is at an apex of the Pex3p spheroid. A 16-residue region of the Pex19p peptide forms an α-helix and makes a contact with Pex3p; this helix is disordered in the unbound state. The Pex19p peptide contains a characteristic motif, consisting of the leucine triad (Leu18, Leu21, Leu22), and Phe29, which are critical for the Pex3p binding and peroxisome biogenesis. [ABSTRACT FROM AUTHOR]

An MsbA deletion mutant ΔC21 that lacks the two C-terminal α-helices was expressed in Escherichia coli strain C41 and purified by metal-affinity and gel-filtration chromatography. Purified ΔC21 retained 26% of the activity of the wild-type ATPase and had a similar binding affinity to fluorescent nucleotide derivatives. Although crystals of wild-type MsbA complexed with adenosine 5'- (β,γ-imido)triphosphate could not be obtained, crystals of ΔC21 that diffracted to 4.5 A resolution were obtained. The preliminary ΔC21 structure had the outward-facing conformation, in contrast to the previously reported E. coli MsbA structure. This result suggests that deletion of the C-terminal α-helices may play a role in facilitating the outward-facing nucleotide-bound crystal structure of EcMsbA. [ABSTRACT FROM AUTHOR]

Human P-glycoprotein (P-gp), which conveys multidrug resistance, is an ATP-dependent drug efflux pump that transports a wide variety of structurally unrelated compounds out of cells. P-gp possesses a ‘linker region’ of ∼ 75 amino acids that connects two homologous halves, each of which contain a transmembrane domain followed by a nucleotide-binding domain. To investigate the role of the linker region, purified human P-gp was cleaved by proteases at the linker region and then compared with native P-gp. Based on a verapamil-stimulated ATP hydrolase assay, size-exclusion chromatography analysis and a thermo-stability assay, cleavage of the P-gp linker did not directly affect the preservation of the overall structure or the catalytic process in ATP hydrolysis. However, linker cleavage increased the kcat values both with substrate ( ksub) and without substrate ( kbasal), but decreased the ksub/ kbasal values of all 10 tested substrates. The former result indicates that cleaving the linker activates P-gp, while the latter result suggests that the linker region maintains the tightness of coupling between the ATP hydrolase reaction and substrate recognition. Inspection of structures of the P-gp homolog, MsbA, suggests that linker-cleaved P-gp has increased ATP hydrolase activity because the linker interferes with a conformational change that accompanies the ATP hydrolase reaction. Moreover, linker cleavage affected the specificity constants [ ksub/ Km(D)] for some substrates (i.e. linker cleavage probably shifts the substrate specificity profile of P-gp). Thus, this result also suggests that the linker region regulates the inherent substrate specificity of P-gp. [ABSTRACT FROM AUTHOR]

The occurrence of a sound event in a sound stream can be perceived without difficulty even when it is difficult to define the acoustic boundary. This study was designed to investigate which acoustical feature functions as an effective cue to “mark” the occurrence of a new event. When two steady sounds are connected by a short frequency glide, at which point do we perceive the occurrence of the second sound: the starting point of the frequency glide or its ending point? To answer this question, experimental stimuli were designed to establish an isochronous structure that was conveyed exclusively by the starting or by the ending point of the glide. The frequency glide was made by modulating the carrier frequency of a sinusoid or the resonant frequency of a complex tone. In two experiments, the deviation from the isochronous structure was detected more easily under the starting-point condition than under the ending-point condition. The results suggest that the starting point functions as a more effective cue for marking a new event than does the ending point. [ABSTRACT FROM AUTHOR]

Δ1-Pyrroline-5-carboxylate dehydrogenase (P5CDh) is known to preferentially use NAD+ as a coenzyme. The kcat value of Thermus thermophilus P5CDh ( TtP5CDh) is four times lower for NADP+ than for NAD+. The crystal structure of NADP+-bound TtP5CDh was solved in order to study the structure–activity relationships for the coenzymes. The binding mode of NADP+ is essentially identical to that in the previously solved NAD+-bound form, except for the regions around the additional 2′-phosphate group of NADP+. The coenzyme-binding site can only accommodate this group by the rotation of a glutamate residue and subtle shifts in the main chain. The 2′-phosphate of NADP+ increases the number of hydrogen bonds between TtP5CDh and NADP+ compared with that between TtP5CDh and NAD+. Furthermore, the phosphate of the bound NADP+ would restrict the `bending' of the coenzyme because of steric hindrance. Such bending is important for dissociation of the coenzymes. These results provide a plausible explanation of the lower turnover rate of NADP+ compared with NAD+. [ABSTRACT FROM AUTHOR]

PsbP is a membrane-extrinsic subunit of the water-oxidizing complex photosystem II (PS II). The evolutionary origin of PsbP has long been a mystery because it specifically exists in higher plants and green algae but not in cyanobacteria. We report here the crystal structure of PsbP from Nicotiana tabacum at a resolution of 1.6 Å. Its structure is mainly composed of ß-sheet, and is not similar to any structures in cyanobacterial PS II. However, the electrostatic surface potential of PsbP is similar to that of cyanobacterial PsbV (cyt c550), which has a function similar to PsbP. A structural homology search with the DALI algorithm indicated that the folding of PsbP is very similar to that of Mog1p, a regulatory protein for the nuclear transport of Ran GTPase. The structure of PsbP provides insight into its novel function in GTP-regulated metabolism in PS II. [ABSTRACT FROM AUTHOR]

Abstract: To investigate mechanisms for perceiving the duration of an auditory event, an effect of perceptual grouping upon perceived duration was studied psychophysically. In the first experiment, the perceived duration of a spoken word was measured under three conditions of acoustic continuity (i.e., (a) intact, (b) noise-replaced, and (c) gap-replaced) as a function of the duration of the target stimulus. Under the noise-replaced condition, a portion of the target stimulus was physically replaced with a noise burst. Under the gap-replaced condition, the replacement was made with a gap. The gap-replacement resulted in a prominent shrinkage of the perceived duration. In the case of noise-replacement, the amount of shrinkage was moderate but highly significant, although the word employed was perceived to be phonetically intact. Independent of this effect of replacement, the amount of shrinkage was also affected by the physical duration of the target stimulus. The second experiment tested an effect of noise replacement on the perceived duration of a tone burst. In this case, the noise replacement also shrunk the perceived duration of the non-speech stimulus. This noise-induced shrinkage could be regarded as being general for the auditory duration. The phenomenon is discussed in relation to a revised model for perceived duration. [ABSTRACT FROM AUTHOR]

Abstract Background: The purpose of this study was to investigate the potential of combining cytosine deaminase/5-fluorocytosine (CD/5-FC) gene therapy and radiation therapy (either external beam radiation or radioimmunotherapy [RIT]), for the treatment of prostate cancer. Methods: Tumor xenografts of CD-transduced LNCaP cells grown in the testes of severe combined immunodeficiency (SCID) mice were used to evaluate antitumor effect. The mice were injected intraperitoneally with 500 mg/kg of 5-FC, or with 5, 15 or 30 mg/kg of 5-fluorouracil (5-FU), for 9 days. The tumors were treated with fractionated radiation at a dose of 1 or 3 Gy/day for 3 days, or I-131 labelled anti-prostate specific antigen (anti-PSA) monoclonal antibody (mAb) administration at a subtherapeutic dose of 20 or 80 µCi. Intratumoral and serum concentrations of 5-FU were measured using high performance liquid chromatography. Results: Mice treated with CD/5-FC gene therapy presented a significant tumor growth inhibition comparable to that obtained with 15 mg/kg, 5-FU systemic administration without marked weight loss. Treatment with CD/5-FC gene therapy resulted in higher tumor but lower serum concentrations of 5-FU than treatment with systemic 5-FU chemotherapy. An additive antitumor effect was obtained when CD/5-FC therapy was combined with 1 Gy irradiation, which by itself did not produce a significant antitumor effect. However, the efficacy of CD/5-FC therapy was not enhanced when combined with RIT, probably due to poor accumulation of the mAb as the tumor/blood ratio never exceeded 1. Conclusion: These findings indicate that CD/5-FC gene therapy for prostate cancer may function with enhanced antitumor effect when combined with external beam radiation. However, combining CD/5-FC gene therapy and RIT using an anti-PSA mAb may not be effective because of insufficient accumulation of the mAb at the target tumors. [ABSTRACT FROM AUTHOR]

Abstract Background: The 150-kDa oxygen-regulated protein ORP150, a new member of the heat shock protein family that functions as a molecular chaperone in the endoplasmic reticulum, was found to increase in infiltrating cancer cells. Since enhancement of ORP150 expression and the presence of vascular endothelial growth factor (VEGF) in human prostate cancer glands were immunohistochemically demonstrated, we examined whether transduced antisense ORP150 cDNA can reduce angiogenicity and tumorigenicity through suppression of VEGF secretion. Methods: Human prostate cancer specimens were immunohistochemically stained with fluorescein isothiocyanate (FITC) for ORP150 or vascular endothelial growth factor (VEGF). An adenovirus vector (Ad) carrying antisense ORP150 cDNA (AdCA-Antisense ORP150) was constructed and infected to prostate cancer DU145 cells. Expression of ORP150 in the cells was analyzed with western blotting and secretion of VEGF into the supernatant with an enzyme-linked immunoabsorbent assay (ELISA). Angiogenicity was evaluated by chorioallantoic membrane (CAM) assay. A nude mouse xenograft model was used to examine tumorigenicity. Results: Immunohistochemical study proved that the expression of ORP150 and VEGF was enhanced in the cytoplasm of prostate cancer cells. The Ad showed 100% transduction efficiency and minimum cytotoxicity when the cells were infected at a multiplicity of infection (MOI) of 20 for 24 h. Expression of ORP150 was substantially reduced by the antisense treatment. Secretion of VEGF into the culture supernatant was reduced to 30%. Consequently, the CAM assay showed relatively low angiogenicity, while marked suppression of tumor formation was observed in the xenograft model. Conclusion: Adenoviral-mediated antisense ORP150 cDNA transfer is well worth considering as an option for prostate cancer gene therapy. [ABSTRACT FROM AUTHOR]

Abstract Background : The aim of this study was to investigate the efficacy and safety of high-dose chemotherapy (HDCT) for the treatment of patients with advanced testicular cancer. Methods : Fourteen patients were treated with high-dose carboplatin, etoposide and cyclophosphamide (with or without THP-adriamycin) followed by peripheral blood stem cell transplantation. The treatment was used for two refractory cases, a second relapse, and for consolidation after the first relapse in one case each. It was also used for nine cases as part of the first-line treatment following primary conventional-dose chemotherapy, and for one case as the first salvage for a late recurrent tumor of teratoma with malignant transformation. Results : The first two patients who received intensive pretreatment with cisplatin-based chemotherapy did not respond to HDCT. The two patients who were treated with HDCT as the first or second salvage therapy achieved successful outcomes. The results for the subsequent nine patients (consisting of two with stage IIIC, five with IIIB2, one with IIB, and one extragonadal seminoma) were two progressive disease, three no change and four partial remission. Only three are alive with NED following salvage surgery. Finally, a case of teratoma with malignant transformation did not respond well to two cycles of HDCT. There were no marked adverse reactions except one episode of severe neutropenic colitis. Conclusions : The results demonstrated the limited efficacy of HDCT even in cases with a good to intermediate risk rating according to classification by the International Germ Cell Cancer Collaborative Group. Because treatment for relapse after HDCT is extremely difficult, new HDCT regimens consisting of drugs that are not used in induction chemotherapy need to be established. [ABSTRACT FROM AUTHOR]

We report a rare case of ectopic ureter accompanied by a giant ureteral stone and pyonephrosis. A 19-year-old Japanese woman consulted our hospital due to low-grade fever and pain in the left flank. The patient had seeping urinary incontinence since childhood. Ultrasonography demonstrated left dysplastic lumbar kidney with hydronephrosis and plain kidney-ureter-bladder film showed a left giant ureteral stone. The left kidney showed no excretion of contrast medium on drip infusion pyelography. After further examination, a diagnosis of left simple ectopic ureter emptying into the vagina was made. Percutaneous nephrostomy penetrating the major psoas muscle was carried out for intractable pyonephrosis, followed by left nephroureterectomy and lithectomy. These procedures resulted in improvement in the patient's symptoms. [ABSTRACT FROM AUTHOR]

Pz peptidase A is an intracellular M3 metallopeptidase found in the thermophile Geobacillus collagenovorans MO-1 that recognizes collagen-specific tripeptide units (Gly-Pro-Xaa). Pz peptidase A shares common reactions with mammalian thimet oligopeptidase (TOP) and neurolysin, but has extremely low primary sequence identity to these enzymes. In this work, Pz peptidase A was cocrystallized with a phosphine peptide inhibitor (PPI) that selectively inhibits TOP and neurolysin. The crystals belong to space group P21, with unit-cell parameters a = 56.38, b = 194.15, c = 59.93 Å, β = 106.22°. This is the first crystallographic study of an M3 family peptidase–PPI complex. [ABSTRACT FROM AUTHOR]

To elucidate the structure and molecular mechanism of a characteristic proline-specific aminopeptidase produced by the thermophile Aneurinibacillus sp. strain AM-1, its gene was cloned and the recombinant protein was overexpressed in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method. X-ray diffraction data were collected to 1.8 Å resolution from the recombinant aminopeptidase crystal. The crystals belong to the orthorhombic space group P21212, with unit-cell parameters a = 93.62, b = 68.20, c = 76.84 Å. A complete data set was also obtained from crystals of SeMet-substituted aminopeptidase. Data in the resolution range 20–2.1 Å from the MAD data set from the SeMet-substituted crystal were used for phase determination. [ABSTRACT FROM AUTHOR]

Bacterial fructosyl amino acid oxidase [fructosyl α-L-amino acid:oxygen oxidoreductase (defructosylating); EC 1.5.3] has been crystallized by the hanging-drop vapour-diffusion technique using sodium citrate as the precipitant. Two types of crystals were grown: one type are rhombic prismatic yellow crystals that belong to space group C with unit-cell parameters a = 101.08, b = 63.36, c = 83.07 Å, β = 108.80° and diffract to at least 1.8 Å resolution, while the second type are rod-like crystals that belong to space group P4122 or P4322 with unit-cell parameters a = b = 119.09, c = 164.66 Å and diffract to 2.7 Å resolution. [ABSTRACT FROM AUTHOR]

Escherichia coli B γ-glutamylcysteine synthetase (γOCS) catalyzes the ATP-dependent coupling of L-Glu and L-Cys to form the glutathione precursor γ-L-Glu-Cys and is a target for development of potential therapeutic agents. By introducing four point mutations of surface-exposed cysteine residues to serine, the γOCS was purified to homogeneity; single crystals have been obtained using the hanging-drop vapour-diffusion method with sodium formate. The γOCS crystal diffracted to 2.8 Å and belongs to space group R3, with unit-cell parameters a = b = 326.7, c = 103.9 Å. [ABSTRACT FROM AUTHOR]

Ribosome recycling factor (RRF) plays a central role during the recycling of ribosomes in the final step of protein biosynthesis in prokaryotes and is therefore a favourable target for the development of new antibiotics. The crystal structure of Escherichia coli RRF has been reported to have an open L-shaped conformation, while other RRFs from thermophilic bacteria have a strict L-shaped conformation [Yun et al. (2000), Acta Cryst. D56, 84-85]. Wild-type E. coli RRF has so far not been crystallized free from bound detergent. Here, a mutant of RRF, Arg132Gly, has been crystallized without any detergent. A complete data set from a crystal of this mutant obtained by the hanging-drop vapour-diffusion method has been collected at 2.2 Å resolution using synchrotron radiation at 100 K. The crystal belongs to the monoclinic space group P21, with unit-cell parameters a = 46.02, b = 49.27, c = 49.37 Å, β = 110.1°. The currently refined structure indicates that RRF has a tRNA-like L-shaped conformation. [ABSTRACT FROM AUTHOR]

Crystals of endopolygalacturonase I from Stereum purpureum have been obtained by the vapour-diffusion method. Prior to crystallization work, endopolygalacturonase I was deglycosylated with endo-β-N-acetylglucosaminidase H. The crystal diffracts to ultrahigh (0.96 Å) resolution using synchrotron radiation and belongs to space group P1, with unit-cell parameters a = 37.26, b = 46.34, c = 52.05 Å, α = 67.17, β = 72.44, γ = 68.90°. [ABSTRACT FROM AUTHOR]

The case is reported of incarcerated intestinal hernia in a hernia sac of reversed ileal conduit wall protruding through the stoma. An 82-year-old woman presented with suspected parastomal intestinal hernia. The patient had undergone total cystectomy with ileal conduit construction 3 years previously, followed by stomal reconstruction surgery for stomal stenosis 2 years later. She had been taking various kinds of purgatives for severe constipation. Initially, this case was diagnosed as parastomal hernia, but emergency surgery demonstrated that incarcerated intestinal hernia in a hernia sac of reversed ileal conduit wall was protruding through the stoma. This case was apparently caused by high abdominal pressure and thinning of the ileal conduit wall. [ABSTRACT FROM AUTHOR]

Pyruvate phosphate dikinase (PPDK) from maize catalyzes the reversible conversion of ATP, orthophosphate and pyruvate to AMP, pyrophosphate and PEP. In higher plants, this enzyme is believed to be involved in the C&sub4; dicarboxylic acid pathway. PPDK was crystallized by the vapour-diffusion method using polyethylene glycol as a precipitant. The crystals belong to the orthorhombic space group C2, with unit-cell parameters a = 108.2, b = 100.2, c = 108.4 Å, β = 96.5 &degrees;, and diffract to 2.3 Å using Spring-8 synchrotron radiation. [ABSTRACT FROM AUTHOR]

In order to establish a new method for simultaneous determination of sulfated bile acids without deconjugation, separation of 3-sulfates of unconjugated, glyco- and tauroconjugated bile acids by high-performance liquid chromatography has been undertaken. The preliminary experiment indicated that reversed phase chromatography on an ODS SC-02 column using ammonium carbonate aqueous solution/acetonitrile as a mobile phase would be promising. The ratio of peak width to retention time was varied depending upon the salt concentration. The content of water in mobile phase exerted an influence on the capacity ratio to a certain extent, but not on the resolution. The use of 0.5% ammonium carbonate/acetonitrile (26∶8 and 20∶8, v/v) was found to be suitable for complete resolution of sulfated cholate, chenodeoxycholate, deoxycholate, lithocholate and their glyco- and tauro-conjugates. [ABSTRACT FROM AUTHOR]

Δ1-Pyrroline-5-carboxylate dehydrogenase (P5CDh) is known to preferentially use NAD+ as a coenzyme. The kcat value of Thermus thermophilus P5CDh ( TtP5CDh) is four times lower for NADP+ than for NAD+. The crystal structure of NADP+-bound TtP5CDh was solved in order to study the structure–activity relationships for the coenzymes. The binding mode of NADP+ is essentially identical to that in the previously solved NAD+-bound form, except for the regions around the additional 2′-phosphate group of NADP+. The coenzyme-binding site can only accommodate this group by the rotation of a glutamate residue and subtle shifts in the main chain. The 2′-phosphate of NADP+ increases the number of hydrogen bonds between TtP5CDh and NADP+ compared with that between TtP5CDh and NAD+. Furthermore, the phosphate of the bound NADP+ would restrict the `bending' of the coenzyme because of steric hindrance. Such bending is important for dissociation of the coenzymes. These results provide a plausible explanation of the lower turnover rate of NADP+ compared with NAD+. [ABSTRACT FROM AUTHOR]

Fructosyl peptide oxidase (FPOX) catalyses the oxidation of α-glycated dipeptides such as N(α)-(1-deoxy-D-fructos-1-yl)-L-valyl-L-histidine (Fru-ValHis) and is used in the diagnosis of diabetes mellitus. Here, two thermostable mutants of FPOX, CFP-T7 and EFP-T5M, were crystallized by the sitting-drop vapour-diffusion method. The crystal of CFP-T7 belonged to the tetragonal space group P4(1)2(1)2, with unit-cell parameters a = b = 110.09, c = 220.48 Å, and that of EFP-T5M belonged to the monoclinic space group P2(1), with unit-cell parameters a = 43.00, b = 230.05, c = 47.27 Å, β = 116.99°. The crystals of CFP-T7 and EFP-T5M diffracted to 1.8 and 1.6 Å resolution, respectively.

An MsbA deletion mutant DeltaC21 that lacks the two C-terminal alpha-helices was expressed in Escherichia coli strain C41 and purified by metal-affinity and gel-filtration chromatography. Purified DeltaC21 retained 26% of the activity of the wild-type ATPase and had a similar binding affinity to fluorescent nucleotide derivatives. Although crystals of wild-type MsbA complexed with adenosine 5'-(beta,gamma-imido)triphosphate could not be obtained, crystals of DeltaC21 that diffracted to 4.5 A resolution were obtained. The preliminary DeltaC21 structure had the outward-facing conformation, in contrast to the previously reported E. coli MsbA structure. This result suggests that deletion of the C-terminal alpha-helices may play a role in facilitating the outward-facing nucleotide-bound crystal structure of EcMsbA.

Delta(1)-Pyrroline-5-carboxylate dehydrogenase (P5CDh) is known to preferentially use NAD(+) as a coenzyme. The k(cat) value of Thermus thermophilus P5CDh (TtP5CDh) is four times lower for NADP(+) than for NAD(+). The crystal structure of NADP(+)-bound TtP5CDh was solved in order to study the structure-activity relationships for the coenzymes. The binding mode of NADP(+) is essentially identical to that in the previously solved NAD(+)-bound form, except for the regions around the additional 2'-phosphate group of NADP(+). The coenzyme-binding site can only accommodate this group by the rotation of a glutamate residue and subtle shifts in the main chain. The 2'-phosphate of NADP(+) increases the number of hydrogen bonds between TtP5CDh and NADP(+) compared with that between TtP5CDh and NAD(+). Furthermore, the phosphate of the bound NADP(+) would restrict the ;bending' of the coenzyme because of steric hindrance. Such bending is important for dissociation of the coenzymes. These results provide a plausible explanation of the lower turnover rate of NADP(+) compared with NAD(+).