Studies on peroxisomes. VI. Relationship between the peroxisomal core and urate oxidase.

The peroxisomal core from the liver of rats was purified 450-fold as a marker of urate oxidase [EC 1.7.3.3.] activity. This preparation has a high specific activity of urate oxidase but not of other peroxisomal enzymes: D-amino acid oxidase [EC 1.4.3.3.], L-alpha-hydroxy acid oxidase [EC 1.1.3.15], or catalase [EC 1.11.1.6]. No activity of marker enzymes for other subcellular particles; cytochrome c oxidase [EC1.9.3.1] (mitochondria), acid phosphatase [EC 3.1.3.2] (lysosomes), or glucose-6-phosphatase [EC 3.1.3.9] (microsomes), was detected in this preparation. The core obtained showed a single protein band in sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the position of the band was found to correspond to a molecular weight 35,000. When the peroxisomal core was subjected to treatment at various pH's with 0.1 M carbonate buffer, urate oxidase was almost completely solubulized at pH 11.0, although approximately 35% of the core protein still remained in the pellet After solubilization of the core at pH 11.0, the specific activity of urate oxidase in the supernatant increased about 1.6 times; the density of the insoluble protein remaining in the pellet was identical with the that of the original core on sucrose density gradient centrifugation.     

Modulation of chondrocyte migration and aggregation by insulin-like growth factor-1 in cultured cartilage

The effect of insulin-like growth factor-1 (IGF-1) on the behavior of rabbit chondrocytes in cultured collagen (CL) gels initially seeded with 2 × 10⁵ cells/ml was examined. On day 5, the frequency of migrating cells cultured in presence of 100 ng IGF-1/ml was 0.04, which was 54 % of the frequency in IGF-1-free culture. The presence of IGF-1 caused an increase in the frequency of dividing cells from 0.09 to 0.13. These results suggest that IGF-1 suppressed the migration of chondrocytes in the CL gels while stimulating cell division in the initial culture phase. The proteolytic migration of cells was thought to be suppressed by the down-regulation of membrane type 1 matrix metalloproteinase by IGF-1. This contributed to the formation of aggregates with spherical-shaped cells that produced collagen type II.     

C-terminal truncation of Noxa1 greatly enhances its ability to activate Nox2 in a pure reconstitution system

Noxa1 activates Nox2 together with Noxo1 and Rac in a pure reconstitution system, but the resulting activity is considerably lower than that induced by p67^phox and p47^phox. In this study, we found that C-terminal-truncated forms of Noxa1 exhibited higher activities than full-length Noxa1. Of the truncations examined, Noxa1(1-225) showed the highest ability for activation. Kinetic studies revealed that Noxa1(1-225) had a threefold higher V_max value than full-length Noxa1 with a similar EC₅₀ value. The affinities of Noxo1 and RacQ61L were not much altered by the truncation. Conversely, the affinity of FAD for the Nox2 complex was enhanced after the truncation. In the absence of Noxo1, Noxa1(1-225) showed much higher activity with a lower EC₅₀ than full-length Noxa1. Noxa1(1-225) showed comparable activity to that of p67^phox with either Noxo1 or p47^phox, although the stability was lower than that with p67^phox and p47^phox. These findings indicate that the role of the C-terminal half of Noxa1 is autoinhibition. The data suggest a two-step autoinhibition mechanism, comprising self-masking to interrupt the binding to the oxidase, and holding of the activation domain in a suboptimal position to the oxidase. This study reveals that when both types of inhibition are released, Noxa1 achieves high-level superoxide production.     

Production of Acetone and Butanol Using Immobilized Growing-cells of Clostridium acetobutylicum

The biosynthetic route for chloromonilicin, an antifungal substance from cherry rot fungus, was investigated using deuterium-labeled precursors. The incorporation of synthetic deuterium-labeled moniliphenone into chloromonilicin and into its xanthone precursor, 4-chloropinselin, was confirmed by ’H-NMR spectrometry.     

Characteristic seasonal variation in dissolved uranium concentration induced by the change of lake water pH in Lake Biwa,Japan

Epilimnion-dominated profiles of dissolved uranium (U) have been observed during summer in an oxygenated Japanese lake, Lake Biwa, contrary to the commonly accepted view that U shows conservative behavior in oxygenated seas and lakes. Monthly observations were conducted to reveal the mechanism for such characteristic distribution and geochemical behavior of dissolved U in the lake. In the surface water, dissolved U concentration started to increase in spring, peaked in summer, and decreased from autumn to winter. In contrast, the concentration remained almost constant in the middle layer (40 m depth) and decreased slightly in the bottom layer (70 m depth) throughout the stagnation period. Mass balance calculations of U suggest that the major mechanism for seasonal variations in the surface layer is the supply of U, not via water inflow from the watershed, but by internal chemical reactions within the lake. A laboratory experiment using the lake water and sediment demonstrated that U was desorbed from and adsorbed onto sediment in response to variations in lake water pH. From these results, it is inferred that the seasonal variation in the concentration of dissolved U in the epilimnion results mainly from the desorptive/adsorptive processes of U between sediment/water interface in response to variation in water pH, which is affected by biological activity in the lake.     

p53DINP1, a p53-inducible gene, regulates p53-dependent apoptosis

Using the differential display method combined with a cell line that carries a well-controlled expression system for wild-type p53, we isolated a p53-inducible gene, termed p53DINP1 (p53-dependent damage-inducible nuclear protein 1). Cell death induced by DNA double-strand breaks (DSBs), as well as Ser46 phosphorylation of p53 and induction of p53AIP1, were blocked when we inhibited expression of p53DINP1 by means of an antisense oligonucleotide. Overexpression of p53DINP1 and DNA damage by DSBs synergistically enhanced Ser46 phosphorylation of p53, induction of p53AIP1 expression, and apoptotic cell death. Furthermore, the protein complex interacting with p53DINP1 was shown to phosphorylate Ser46 of p53. Our results suggest that p53DINP1 may regulate p53-dependent apoptosis through phosphorylation of p53 at Ser46, serving as a cofactor for the putative p53-Ser46 kinase.     

Total and high molecular weight adiponectin and hepatocellular carcinoma with HCV infection

Background

Adiponectin is shown to be inversely associated with development and progression of various cancers. We evaluated whether adiponectin level was associated with the prevalence and histological grade of hepatocellular carcinoma (HCC), and liver fibrosis in patients with hepatitis C virus (HCV) infection.

Methods

A case-control study was conducted on 97 HCC patients (cases) and 97 patients (controls) matched for sex, Child-Pugh grade and platelet count in patients with HCV infection. The serum total and high molecular weight (HMW) adiponectin levels were measured by enzyme-linked immunosorbent assays and examined in their association with the prevalence of HCC. In addition, the relationship between these adiponectin levels and body mass index (BMI), progression of liver fibrosis, and histological grade of HCC was also evaluated. Liver fibrosis was assessed using the aspartate aminotransferase to platelet ratio index (APRI).

Results

There were no significant differences in the serum total and HMW adiponectin levels between cases and controls. Moreover, there were no inverse associations between serum total and HMW adiponectin levels and BMI in both cases and controls. On the other hand, serum total and HMW adiponectin levels are positively correlated with APRI in both cases (r = 0.491, P<0.001 and r = 0.485, P<0.001, respectively) and controls (r = 0.482, P<0.001 and r = 0.476, P<0.001, respectively). Interestingly, lower serum total (OR 11.76, 95% CI: 2.97–46.66 [P<0.001]) and HMW (OR 10.24, CI: 2.80–37.40 [P<0.001] adiponectin levels were independent risk factors of worse histological grade of HCC.

Conclusions

Our results suggested that serum total and HMW adiponectin levels were predictors of liver fibrosis, but not prevalence of HCC in patients with HCV infection. Moreover, low these adiponectin levels were significantly associated with worse histological grades.     

Copulatory courtship behavior and sine song as a mate recognition cue in Drosophila lini and its sibling species

Most Drosophila species sing species-specific pulse songs during their "precopulatory courtship." Three sibling species of the Drosophila montium species subgroup performed "copulatory courtship": males generated courtship songs by vibrating either wing only after mounting and during copulation. In these three species, strong sexual isolation was detected between D. ohnishii and D. lini and between D. ohnishii and D. ogumai, but not between D. lini and D. ogumai. Female showed strong repelling behavior when they were mounted by a heterospecific male in the species combinations including D. ohnishii, resulting in failure of the copulation attempt of the male. Acoustic analyses of courtship songs revealed that the pulse song was irregular, without any species-specific parameters, but that the frequency of the sine song was different among the three species in accordance with the modes of sexual isolation between them; it was significantly lower in D. ohnishii (mean ± SE = 193.0 ± 1.7 Hz) but higher in D. lini (253.4 ± 2.7 Hz) and D. ogumai (246.7 ± 5.3 Hz). We suggest that this difference in the sine song frequency is a sexual signal in the Specific Mate Recognition System (SMRS) among these three Drosophila species.