Annual cycle of circulations in Lake Biwa, part 2: mechanisms

We investigate the mechanisms of circulations in Lake Biwa using model results that are driven by the momentum and heat fluxes evaluated from meteorological parameters observed around and on the lake. The kinetic energy budget shows that the wind stress over the lake, which has positive (negative) curl over the northern (southern) region of the lake throughout the year, generates the current system of multiple gyres in the stratified season (May–January), while the thermal forcing makes a minor contribution. This mechanism is quite different from the previous picture, in which the thermal forcing was dominant in forming the gyre system during the heating season. In the cooling season, the overturning of the lake occurs not in a simple one-dimensional (vertical) process, but in a time-dependent three-dimensional one. Cold water that is formed in the shallow eastern region descends the bottom slope to the deep lake in December to March while such a descent hardly occurs in the western region. A large shallow area (small heat capacity) as well as intense cooling in the eastern region effectively produces cold water. Wind again plays an important role by intensifying the cooling through its higher speed and by driving slope current. Further, the fluctuating wind direction causes significantly time-dependent behaviors of the slope current or bottom water formation.     

Variations in amount of TSST-1 produced by clinical methicillin resistant Staphylococcus aureus (MRSA) isolates and allelic variation in accessory gene regulator (agr) locus

Background  

Staphylococcus aureus (S. aureus) is an important pathogen associated with both nosocomial and community-acquired infections and its pathogenicity is attributed to its potential to produce virulence factors. Since the amount of toxin produced is related to virulence, evaluating toxin production should be useful for controlling S. aureus infection. We previously found that some strains produce relatively large amounts of TSST-1; however, no reports have described the amount of TSST-1 produced by clinical isolates.     

Effects of co-exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin and perfluorooctane sulfonate or perfluorooctanoic acid on expression of cytochrome P450 isoforms in chicken (Gallus gallus) embryo hepatocyte cultures

In this study we investigated the effect of a single-compound exposure or two compound co-exposure to tetrachlorodibenzo-p-dioxin (TCDD) plus perfluorooctane sulfonate (PFOS) or perfluorooctanoic acid (PFOA) on the mRNA expression of cytochromes P450 (CYP) 1A4, 4V2 and 3A37, ethoxyresorufin-O-deethylase (EROD) activity and cell viability in chicken (Gallus gallus domesticus) embryo primary hepatocyte cultures. Cell viability after 24 h of incubation was significantly decreased in cells exposed to PFOS at concentrations between 30 µM and 60 µM with or without co-exposure to TCDD (0.3 nM at maximum). PFOA did not decrease cell viability even at maximum concentrations of 60 µM. TCDD induced CYP1A4 mRNA and EROD activity substantially as reported previously. PFOS also increased CYP1A4 mRNA in a concentration-dependent manner. Co-exposure of cells to PFOS plus TCDD did not change CYP1A4 mRNA levels compared to cells treated with TCDD alone. PFOS alone did not induce CYP4V2 mRNA, however 40–50 µM PFOS plus TCDD (0.3 nM) induced CYP4V2 mRNA compared to TCDD alone (P < 0.05). This trend was similar to that observed with co-exposure to TCDD plus PFOA, suggesting that PFOA alone did not induce CYP4V2 mRNA, whereas co-exposure to TCDD plus PFOA induced the expression levels. PFOS alone decreased CYP3A37 mRNA by a maximum of 45%, however after co-exposure to TCDD, recovery of mRNA expression to levels measured in DMSO-treated cells was observed. Our data suggest a complex gene response to mixtures of dioxin-like and perfluorinated compounds.     

Regulation of the expression of ribulose-1,5-bisphosphate carboxylase/oxygenase (EC 4.1.1.39) in a cyanobacterium, Synechococcus PCC7942

When cyanobacterium cells are grown under extremely low CO₂ concentration, the number of carboxysomes, structures containing ribulose-bisphosphate carboxylase (Rubisco; EC 4.1.1.39), is known to increase. This suggests that Rubisco helps to regulate photosynthesis in cyanobacteria. However, no studies have been done on the changes of Rubisco content and activity in response to the extracellular CO₂ concentration, and no information is available on its effect on photosynthesis. To elucidate the relationship between the expression responses of Rubisco and extracellular CO₂, wild-type cells (Synechococcus PCC7942) and carboxysome-lacking cells were grown under various CO₂ concentrations, and Rubisco activity was determined. In both strains, Rubisco activity increased when the cells were grown under a CO₂ concentration around, or less than, K _1/2(CO₂) of photosynthesis. In carboxysome-lacking cells, Rubisco activity increased five to six times at most, and a simultaneous increase in the rate of photosynthesis was observed. These results suggest that stimulation of expression of Rubisco occurs to compensate for the decrease in the rate of photosynthesis under CO₂-limited conditions. This revised version was published online in August 2006 with corrections to the Cover Date.     

Identification and characterization of a novel calcineurin-binding protein in scallop testis

Calcineurin has been inferred to function in meiosis and spermiogenesis in testis. Here, we identified a calcineurin-binding protein in scallop testis by Far-Western blot analysis using purified calcineurin as a probe. The molecular mass of the binding protein estimated on the blot was 75 kDa. The isolated cDNA clone encoded a novel 474-residue protein, named CaNBP75. The region between T6 and A210 of CaNBP75 was responsible for the interaction with calcineurin. CaNBP75 was predominantly expressed in testis and ovary of scallop. Thus, CaNBP75 may modulate the physiological function of calcineurin in the testis and ovary of scallop, such as in spermiogenesis or meiosis.     

Crystal structures of Aspergillus oryzae aspartic proteinase and its complex with an inhibitor pepstatin at 1.9A resolution

The X-ray structures of Aspergillus oryzae aspartic proteinase (AOAP) and its complex with inhibitor pepstatin have been determined at 1.9A resolution. AOAP was crystallized in an orthorhombic system with the space group P2(1)2(1)2(1) and cell dimensions of a=49.4A, b=79.4A, and c=93.6A. By the soaking of pepstatin, crystals are transformed into a monoclinic system with the space group C2 and cell dimensions of a=106.8A, b=38.6A, c=78.7A, and beta=120.3 degrees. The structures of AOAP and AOAP/pepstatin complex were refined to an R-factor of 0.177 (R(free)=0.213) and of 0.185 (0.221), respectively. AOAP has a crescent-shaped structure with two lobes (N-lobe and C-lobe) and the deep active site cleft is constructed between them. At the center of the active site cleft, two Asp residues (Asp33 and Asp214) form the active dyad with a hydrogen bonding solvent molecule between them. Pepstatin binds to the active site cleft via hydrogen bonds and hydrophobic interactions with the enzyme. The structures of AOAP and AOAP/pepstatin complex including interactions between the enzyme and pepstatin are very similar to those of other structure-solved aspartic proteinases and their complexes with pepstatin. Generally, aspartic proteinases cleave a peptide bond between hydrophobic amino acid residues, but AOAP can also recognize the Lys/Arg residue as well as hydrophobic amino acid residues, leading to the activation of trypsinogen and chymotrypsinogen. The X-ray structure of AOAP/pepstatin complex and preliminary modeling show two possible sites of recognition for the positively charged groups of Lys/Arg residues around the active site of AOAP.     

PAX4 mutation (R121W) as a prodiabetic variant in Okinawans

We previously reported that a missense mutation at codon 121 (CGG(Arg) to TGG(Trp), R121W) of PAX4 may be associated with the onset of type 2 diabetes in Japanese. In this study, we determined the frequency of the R121W mutation of PAX4 and characterized the prodiabetic phenotype in a population-based study. Healthy 372 residents participated in annual health check-ups in Nishihara (Okinawa, Japan) and unrelated 193 type 2 diabetic patients from the outpatient clinic of Ryukyu University Hospital were enrolled. Diagnosis of diabetes was based on the 1997 American Diabetes Association criteria. The R121W mutation in PAX4 was genotyped by PCR-RFLP analysis. In healthy residents, R121W mutation was detected in 12 of 372 residents (3.1%). The prevalence of newly diagnosed type 3 diabetes (25% vs. 5%, p=0.004) and HbA(1c) (5.6+/-1.9 vs. 5.1+/-0.7, p=0.026) was higher in the variants than in the wild-types. The odds ratio of diabetes in the R121W variants was 5.98 with 95% confidence interval from 1.50 to 23.9. The R121W mutation was observed in 12 of the 193 type 2 diabetic patients (6.2%). Onset-ages of diabetes were earlier (37+/-10 vs. 47+/-13 years, p=0.010) and the rate of insulin user was two times higher (83% vs. 41%, p=0.005) in the variants. The R121W mutation in PAX4 is a predisposing factor for the development of type 2 diabetes in Okinawans.     

Correlation between the activities and the oligomeric forms of pig gastric H/K-ATPase

Membrane-bound H/K-ATPase was solubilized by octaethylene glycol dodecyl ether (C(12)E(8)) or n-octyl glucoside (nOG). H/K-ATPase activity and the distribution of protomeric and oligomeric components were evaluated by high-performance gel chromatography (HPGC) and by single-molecule detection using total internal reflection fluorescence microscopy (TIRFM). As evidenced by HPGC of the C(12)E(8)-solubilized enzyme, the distribution of oligomers was 12% higher oligomeric, 44% diprotomeric, and 44% protomeric species, although solubilization by C(12)E(8) reduced the H/K-ATPase activity to 1.8% of that of the membrane-bound enzyme. The electron microscopic images of the C(12)E(8)-solubilized enzyme showed the presence of protomers and a combination of two and more protomers. While the nOG-solubilized H/K-ATPase retained the same turnover number and 71% of the specific activity as that of the membrane-bound enzyme, 56% higher oligomeric, 34% diprotomeric, and 10% protomeric species were detected. TIRFM analysis of solubilized fluorescein 5'-isothiocyanate (FITC)-modified H/K-ATPase at Lys-518 of the alpha-chain showed a quantized photobleaching of the FITC fluorescence intensity. For the C(12)E(8)-solubilized FITC-enzyme, the fraction of each of the initial relative fluorescence intensity units of 4, 2, and 1 was, respectively, 5%, 44% and 51%. In the case of the nOG-solubilized FITC-enzyme, each fraction of 4 and 2 units was, respectively, 54% and 46% with no detectable 1 unit fraction. This represents the first direct observation of H/K-ATPase in aqueous solution. The correlation between the enzymatic activities and distribution of oligomeric forms of H/K-ATPase by HPGC and the observation of a single molecule of H/K-ATPase and others suggests that the tetraprotomeric form of H/K-ATPase molecules represents the functional species in the membrane.     

Microbial ecology of Bacillus thuringiensis: fecal populations recovered from wildlife in Korea

A total of 34 fecal samples, collected from 14 species of wild mammals in Korea, were examined for the occurrence of Bacillus thuringiensis. The organism was detected in 18 (53%) samples. Among the three food-habit groups, herbivorous animals yielded the highest frequency (69%) of samples positive for B. thuringiensis, followed by omnivorous animals (50%). Of the six fecal samples from carnivorous animals, only one sample contained B. thurin giensis. Among 527 isolates belonging to the Bacillus cereus - B. thuringiensis group, 43 (8%) were assigned to B. thurin giensis on the basis of the formation of parasporal inclusions. Of the 43 isolates, 13 were serologically allocated to the nine H-antigenic serotypes: H3ad (serovar sumiyoshiensis), H15 (dakota), H17/27 (tohokuensis/ mexicanensis), H19 (tochigiensis), H21 (colmeri), H29 (amagiensis), H31/49 (toguchini/muju), H42 (jinghongiensis), and H44 (higo). Other isolates were untestable or untypable by the 55 reference H antisera available. Insecticidal activity was associated with 23% of the fecal populations: three isolates killed larvae of the silkworm, Bombyx mori (Lepidoptera), and seven exhibited larvicidal activity against the mosquito, Aedes aegypti (Diptera). There was no larvicidal activity against the three lepidopterous insects: Plutella xylostella, Spodoptera exigua, and Spodoptera litura. The overall results suggest that wild animals in Korea are in contact with naturally occurring B. thuringiensis at high frequencies through the daily food intake of plants.