Arabidopsis Pol II-Dependent in Vitro Transcription System Reveals Role of Chromatin for Light-Inducible rbcS Gene Transcription.

In vitro transcription is an essential tool to study the molecular mechanisms of transcription. For over a decade, we have developed an in vitro transcription system from tobacco (Nicotiana tabacum)-cultured cells (BY-2), and this system supported the basic activities of the three RNA polymerases (Pol I, Pol II, and Pol III). However, it was not suitable to study photosynthetic genes, because BY-2 cells have lost their photosynthetic activity. Therefore, Arabidopsis (Arabidopsis thaliana) in vitro transcription systems were developed from green and etiolated suspension cells. Sufficient in vitro Pol II activity was detected after the minor modification of the nuclear soluble extracts preparation method; removal of vacuoles from protoplasts and L-ascorbic acid supplementation in the extraction buffer were particularly effective. Surprisingly, all four Arabidopsis Rubisco small subunit (rbcS-1A, rbcS-1B, rbcS-2B, and rbcS-3B) gene members were in vitro transcribed from the naked DNA templates without any light-dependent manner. However, clear light-inducible transcriptions were observed using chromatin template of rbcS-1A gene, which was prepared with a human nucleosome assembly protein 1 (hNAP1) and HeLa histones. This suggested that a key determinant of light-dependency through the rbcS gene transcription was a higher order of DNA structure (i.e. chromatin).

Molecular dynamics simulation of effect of glycerol monostearate on amorphous polyethylene in the presence of water.

Polyethylene (PE) is used widely as an electrical insulating material. However, the deterioration of its insulating ability is accelerated by exposure to a humid environment. To prevent the influence of water molecules, mixing 2,3-dihydroxypropyl octadecanoate-known as glycerol monostearate (GMS)-into PE has been proposed. However, the physical mechanism underlying the effect of GMS remains unclear. In the present study, the behavior of water molecules in amorphous PE with and without GMS molecule(s) was investigated in terms of diffusion and clustering using molecular dynamics (MD) simulations. Analyzing the mean square displacement (MSD), diffusion coefficient and radial distribution function (RDF) of the water molecules revealed that GMS contributed to suppressing the diffusion and dispersion of water molecules. Furthermore, it was demonstrated that, in the case of 4 wt% GMS and less than 2 wt% water, GMS contributes to reducing the diffusion coefficient of water molecules but does not change the glass transition temperature (T g) of the system drastically.

Computational study on acetophenone in amorphous polyethylene.

Polyethylene (PE) is widely used as an electrical insulating material. Acetophenone (AP) is a major residue in PE and is considered one of the causes of insulation deterioration. However, the physicochemical explanation of the influence of AP is still unknown. Therefore, in the present study, the behavior of AP molecules in amorphous PE was investigated using molecular dynamics (MD) simulations and quantum chemical calculation. First, the basic properties of the AP molecule were evaluated from the viewpoint of molecular electrostatic potential (MEP), molecular orbitals, and energy levels. Subsequently, an amorphous PE system containing AP molecules was studied using MD simulations. The results clearly indicate that AP does not greatly change the density and radius of gyration of amorphous PE. Quantum computations were performed using a part of the structure obtained from the MD simulations, suggesting that AP acts as a trap site in amorphous PE. It was also revealed that under the external electric field, the total density of state (DOS) changes with a dependence on the applied direction. Results of these calculations help in explaining previous experimental results.

Treatment with buckwheat bran extract prevents the elevation of serum triglyceride levels and fatty liver in KK-A(y) mice.

Buckwheat powder or protein has been shown to decrease the total serum cholesterol level in non-diabetic mice or rats. However, the lipid-lowering effect of buckwheat bran extract (BBE) in diabetic mice has not been fully elucidated. KK-A(y) mice that received six-week treatment with BBE showed decreased body weight and liver weight compared to those of control (vehicle) mice. However, there was no significant difference in food intake. BBE treatments prevented liver triglyceride accumulation and decreased the serum level of triglycerides. In addition, mRNA expression levels lipogenic enzyme genes, fatty acid synthase, acetyl-coenzyme a oxidase and stearyl-coenzyme a desaturase 1, but not those of ß-oxidized enzyme genes, were decreased in BBE-treated mice. Level of transcription factors ChREBP and SREBP1c, transcripts of lipogenic genes, were also decreased in BBE-treated mice. These results suggest that chronic treatment with BBE derivatives could have beneficial effects on hypertriglycemia in patients with type 2 diabetes mellitus.

Sudachitin, a polymethoxylated flavone, improves glucose and lipid metabolism by increasing mitochondrial biogenesis in skeletal muscle.

BACKGROUND: Obesity is a major risk factor for insulin resistance, type 2 diabetes, and stroke. Flavonoids are effective antioxidants that protect against these chronic diseases. In this study, we evaluated the effects of sudachitin, a polymethoxylated flavonoid found in the skin of the Citrus sudachi fruit, on glucose, lipid, and energy metabolism in mice with high-fat diet-induced obesity and db/db diabetic mice. In our current study, we show that sudachitin improves metabolism and stimulates mitochondrial biogenesis, thereby increasing energy expenditure and reducing weight gain. METHODS: C57BL/6 J mice fed a high-fat diet (40% fat) and db/db mice fed a normal diet were treated orally with 5 mg/kg sudachitin or vehicle for 12 weeks. Following treatment, oxygen expenditure was assessed using indirect calorimetry, while glucose tolerance, insulin sensitivity, and indices of dyslipidemia were assessed by serum biochemistry. Quantitative polymerase chain reaction was used to determine the effect of sudachitin on the transcription of key metabolism-regulating genes in the skeletal muscle, liver, and white and brown adipose tissues. Primary myocytes were also prepared to examine the signaling mechanisms targeted by sudachitin in vitro. RESULTS: Sudachitin improved dyslipidemia, as evidenced by reduction in triglyceride and free fatty acid levels, and improved glucose tolerance and insulin resistance. It also enhanced energy expenditure and fatty acid ß-oxidation by increasing mitochondrial biogenesis and function. The in vitro assay results suggest that sudachitin increased Sirt1 and PGC-1α expression in the skeletal muscle. CONCLUSIONS: Sudachitin may improve dyslipidemia and metabolic syndrome by improving energy metabolism. Furthermore, it also induces mitochondrial biogenesis to protect against metabolic disorders.

Extracts of common buckwheat bran prevent sucrose digestion.

Buckwheat has been shown to have various health benefits such as reduction of hypertension and improvement of hypercholesterolemia; however, its effect on diabetes has not been fully elucidated. In this study, buckwheat bran extracts (BBE) inhibited sucrase activity in vitro more effectively than buckwheat. Balb/c mice pretreated with BBE showed dose-dependent reductions of blood glucose, greater than those observed with control mice, within 60 min following oral sucrose administration. Blood glucose levels in mice pretreated with buckwheat extracts were also significantly lower compared to those in control mice within 30 min following oral administration of sucrose. However, rutin, one of the abundant polyphenols of BBE, did not lower blood glucose level. Our data indicate that components of BBE other than rutin have inhibitory activity against sucrase in vivo. These results suggest that BBE could have beneficial effects on diabetes.

Subjective feeling of psychological fatigue is related to decreased reactivity in ventrolateral prefrontal cortex.

The purpose of this study is to examine the relationship between subjective fatigue and brain function. Twenty-three healthy young volunteers participated in this study. Relationships were investigated between subjective fatigue assessed using visual-analogue scale (VAS) score and sleep duration, and cerebral cortex reactivity during a verbal fluency task by 52-channel near-infrared spectroscopy (NIRS). The VAS score negatively correlated with oxygenated hemoglobin concentration ([oxy-Hb]) increases in the bilateral channels over the regions from the ventrolateral part of the frontal lobe to the upper part of the temporal lobe during the verbal fluency task. Sleep duration in the previous night positively correlated with [oxy-Hb] increases in the bilateral channels over the dorsolateral prefrontal lobe also during the verbal fluency task. No significant correlations between the VAS score and sleep duration in the previous night with [oxy-Hb] increases were found during a control task, the left-finger-tapping task. The subjective feeling of psychological fatigue is related to decreased reactivities in the lateral frontal and superior temporal cortices and is unrelated to sleep duration in the previous night, which is reflected in the reactivity in the dorsolateral prefrontal cortices. These results suggest that transient hypofunction and persistent dysfunction in the lateral prefrontal and temporal lobes are among the brain substrates of fatigue. These also demonstrate the advantage of NIRS for investigating brain function during subjective phenomena such as fatigue because it enables examination in a natural setting.