Soil site response

The response of stiff quaternary soil sites to strong earthquake shaking is an important factor in the development of seismic siting criteria for nuclear power plants. Analytical methods for estimating this site response are in common use, but there is need for empirical verification of these modeling techniques. There are two dense 3-D strong motion arrays installed and operating in California to directly measure the response of stiff quaternary soil to earthquake shaking. These are the Hollister Earthquake Observatory and the Borrego Valley Downhole Arrays. Site response measurements from these arrays, along with detailed geotechnical and geophysical site investigations, will provide important calibration and confirmation of site response modeling techniques used for seismic siting criteria development. This paper presents an overview of the two arrays and initial results of the measurements.     

Uromodulin Regulates Murine Aquaporin−2 Activity via Thick Ascending Limb–Collecting Duct Cross−Talk during Water Deprivation

Uromodulin, a urinary protein synthesized and secreted from the thick ascending limb (TAL) of the loop of Henle, is associated with hypertension through the activation of sodium reabsorption in the TAL. Uromodulin is a potential target for hypertension treatment via natriuresis. However, its biological function in epithelial cells of the distal nephron segment, particularly the collecting duct, remains unknown. Herein, we examined the regulation of uromodulin production during water deprivation in vivo as well as the effect of uromodulin on the activity of the water channel aquaporin−2 (AQP2) in vitro and in vivo using transgenic mice. Water deprivation upregulated uromodulin production; immunofluorescence experiments revealed uromodulin adhesion on the apical surface of the collecting duct. Furthermore, the activation of AQP2 was attenuated in mice lacking uromodulin. Uromodulin enhanced the phosphorylation and apical trafficking of AQP2 in mouse collecting duct cells treated with the vasopressin analog dDAVP. The uromodulin-induced apical trafficking of AQP2 was attenuated via endocytosis inhibitor treatment, suggesting that uromodulin activates AQP2 through the suppression of endocytosis. This study provides novel insights into the cross−talk between TAL and the collecting duct, and indicates that the modulation of uromodulin is a promising approach for diuresis and hypertension treatment.     

Design of an Artificial Peptide Inspired by Transmembrane Mitochondrial Protein for Escorting Exogenous DNA into the Mitochondria to Restore their Functions by Simultaneous Multiple Gene Expression

Mitochondria are vital organelles regulating essential cellular functions. Human mitochondrial DNA (mtDNA) consists of 37 genes, 13 of which encode mitochondrial proteins, and the remaining 24 genes encode two ribosomal RNAs and 22 transfer RNAs needed for the translation of the mtDNA-encoded 13 proteins. However, mtDNA often impairs the expression and function of these genes due to various mutations, ultimately causing mitochondrial dysfunction. To recover from this desperate condition, developing the technology to supply all mitochondrial proteins encoded by mtDNA at once is an urgent task, but there is no established strategy for this purpose. In this study, a simple yet effective mitochondrial gene delivery system is proposed comprising an artificial peptide inspired by a transmembrane mitochondrial membrane protein. The designed mitochondria-targeting peptides presented on the carrier surface effectively guide the encapsulated plasmid to the mitochondria, facilitating mitochondrial uptake and gene expression. The developed system successfully delivers exogenous mtDNA to mtDNA-depleted cells and leads to simultaneous multigene expression, ultimately restoring mitochondrial functions, including the mitochondrial respiration rate. The established multiple gene expression system in each mitochondrion is a game-changing technology that can accelerate the development of mitochondrial engineering technologies as well as clinical applications for mitochondrial diseases.     

Dynamic simulation of flow-induced fiber fracture

A dynamic simulation method has been developed of the fracture process of a fiber in a flow field using the particle simulation method proposed i a previous paper. The fiber is modeled with bonded spheres as a fiber model. The flexibility of the fiber model is altered by changing three parameters of the stretching, bending, and twisting constants between adjacent spheres. The stress induced in each bond of the fiber model as a result of deformation is formulated using displacement of the bodn distanc, bond angle, and torsion angle fr each pair of spheres. After deformation, the fiber model fractures at the bond at which the stress surpasses the strength of the fiber. The motion of the fiber model in a flow field is determined by solving the translational and rotational motion equations for individual spheres under the hydrodynamic force and torque exerted on them. The correctness of the method and formulation was verified by comparing the simulated deflection curve of a cantilever beam (with a concentrated load at the end) with the theoretical curve. Good agreement was found in both the deflection and slope of the beam. The fracture process of a fiber after bending deformation in a two-dimensional siimple shear flow was simulated under assumptions of an infinitely dilute system, no hydrodynamic interaction, and a low Reynolds number of a particle. The calculated critical conditions of the flow field for fiber fracture were compared with Forgacs and Mason's theoretical ones. Simulated values of the fracture condition of the fluid shear stress related to the Young's modulus of a fiber agree with theoretical ones over an aspect ratio of 15.     

Investigating the Influence of GABA Neurons on Dopamine Neurons in the Ventral Tegmental Area Using Optogenetic Techniques

Dopamine (DA) is the key regulator of reward behavior. The DA neurons in the ventral tegmental area (VTA) and their projection areas, which include the prefrontal cortex (PFC), nucleus accumbens (NAc), and amygdala, play a primary role in the process of reward-driven behavior induced by the drugs of addiction, including nicotine and alcohol. In our previous study, we developed a novel platform consisting of micro-LED array devices to stimulate a large area of the brain of rats and monkeys with photo-stimulation and a microdialysis probe to estimate the DA release in the PFC. Our results suggested that the platform was able to detect the increased level of dopamine in the PFC in response to the photo-stimulation of both the PFC and VTA. In this study, we used this platform to photo-stimulate the VTA neurons in both ChrimsonR-expressing (non-specific) wild and dopamine transporter (DAT)-Cre (dopamine specific) mice, and measured the dopamine release in the nucleus accumbens shell (NAcShell). We measured the DA release in the NAcShell in response to optogenetic stimulation of the VTA neurons and investigated the effect of GABAergic neurons on dopaminergic neurons by histochemical studies. Comparing the photo-stimulation frequency of 2 Hz with that of 20 Hz, the change in DA concentration at the NAcShell was greater at 20 Hz in both cases. When ChrimsonR was expressed specifically for DA, the release of DA at the NAcShell increased in response to photo-stimulation of the VTA. In contrast, when ChrimsonR was expressed non-specifically, the amount of DA released was almost unchanged upon photo-stimulation. However, for nonspecifically expressed ChrimsonR, intraperitoneal injection of bicuculline, a competitive antagonist at the GABA-binding site of the GABA_A receptor, also significantly increased the release of DA at the NAcShell in response to photo-stimulation of the VTA. The results of immunochemical staining confirm that GABAergic neurons in the VTA suppress DA activation, and also indicate that alterations in GABAergic neurons may have serious downstream effects on DA activity, NAcShell release, and neural adaptation of the VTA. This study also confirms that optogenetics technology is crucial to study the relationship between the mesolimbic dopaminergic and GABAergic neurons in a neural-specific manner.     

Synthesis of Metal Oxide Particles Using Reaction Route from Rare-Earth Metal-EDTA Complexes

Highly dense, spherical yttria (Y₂O₃) and erbia (Er₂O₃) particles were synthesized from their corresponding metal-ethylenediaminetetraacetic (EDTA) complexes. The EDTA·Y·H and EDTA·Er·H complexes were prepared in powdered form. These complexes were used as the staring materials for synthesis of the Y₂O₃ and Er₂O₃ particles. The particles were synthesized using an H₂-O₂ flame produced with a commercial flame spray apparatus. Crystalline structure, surface and cross-sectional morphologies, and elemental distribution of the synthesized particles were investigated. It was confirmed that the crystalline phases of the Y₂O₃ and Er₂O₃ particles were homogeneous. In addition, the elemental distribution of the particles was uniform. These results indicate that dense, spherical particles of Y₂O₃ and Er₂O₃ have been synthesized with EDTA·Y·H and EDTA·Er·H complexes, respectively.     

Microbial participation in iodine volatilization from soils

The roles of microorganisms in iodine volatilization from soils were studied. Soils were incubated with iodide ion (I-), and volatile organic iodine species were determined with a gas chromatograph. Iodine was emitted mainly as methyl iodide (CH3I), and CH3I emission was sometimes enhanced by the addition of glucose. Soils were then incubated with a radioactive iodine tracer (125I), and radioiodine emitted from soils was determined. The emission of iodine was enhanced in the presence of yeast extract but was inhibited by autoclaving of soils. The addition of streptomycin and tetracycline, antibiotics that inhibit bacterial growth, strongly inhibited iodine emission, while a fungal inhibitor cycloheximide caused little effect. Forty bacterial strains were randomly isolated from soils, and their capacities for volatilizing iodine were determined. Among these, 14 strains volatilized significant amounts of iodine when they were cultivated with iodide ion. Phylogenetic analysis based on 16S ribosomal DNA sequences showed thatthese bacteria are widely distributed through the bacterial domain. Our results suggest that iodine in soils is methylated and volatilized as CH3I by the action of soil bacteria and that iodine-volatilizing bacteria are ubiquitous in soil environments. The pathway of iodine volatilization by soil bacteria should be important for understanding the biogeochemical cycling of iodine as well as for the assessment of long-lived radioactive iodine (129I) in the environment.     

Purification and characterization of intracellular and extracellular, thermostable and alkali-tolerant alcohol oxidases produced by a thermophilic fungus, Thermoascus aurantiacus NBRC 31693

Intracellular and extracellular alcohol oxidases (AO int and AO ext) were purified from the liquid and solid cultures of a thermophilic fungus, Thermoascus aurantiacus NBRC 31693, as electrophoretically and isoelectrophoretically homogeneous proteins, respectively. Both enzymes contained a flavin adenine dinucleotide (FAD) cofactor and were stained with Schiff's reagent. The molecular weight of AO int was estimated to be about 320 kDa and its subunit was 75 kDa. The molecular weight of AO ext was about 560 kDa, and it was composed of two types of subunits (75 kDa and 59 kDa). The pIs of AO int and AO ext were 5.88 and 6.08, respectively. AO int and AO ext were stable up to 60 degrees C and 55 degrees C, respectively. The enzymes were stable over a wide range of pH from 6 to 11. AO int oxidized short straight-chain alcohols (K(m) for methanol, 13.5 mM and K(m) for ethanol, 15.8 mM). On the other hand, AO ext could oxidize secondary alcohols and aromatic alcohols (veratryl alcohol and benzyl alcohol) in addition to straight-chain alcohols (K(m) for methanol, 0.5 mM and K(m) for ethanol, 10.2 mM).     

Evaluation of pretreatment with Pleurotus ostreatus for enzymatic hydrolysis of rice straw

The effects of biological pretreatment of rice straw using four white-rot fungi (Phanerochaete chrysosporium, Trametes versicolor, Ceriporiopsis subvermispora, and Pleurotus ostreatus) were evaluated on the basis of quantitative and structural changes in the components of the pretreated rice straw as well as susceptibility to enzymatic hydrolysis. Of these white-rot fungi, P. ostreatus selectively degraded the lignin fraction of rice straw rather than the holocellulose component. When rice straw (water content of 60%) was pretreated with P. ostreatus for 60 d, the total weight loss and the degree of Klason lignin degraded were 25% and 41%, respectively. After the pretreatment, the residual amounts of cellulose and hemicellulose were 83% and 52% of those in untreated rice straw, respectively. By enzymatic hydrolysis with a commercial cellulase preparation for 48 h, 52% holocellulose and 44% cellulose in the pretreated rice straw were solubilized. The net sugar yields based on the amounts of holocellulose and cellulose of untreated rice straw were 33% for total soluble sugar from holocellulose and 32% for glucose from cellulose. The SEM observations showed that the increase in susceptibility of rice straw to enzymatic hydrolysis by pretreatment with P. ostreatus is caused by partial degradation of the lignin seal. When the content of Klason lignin was less than 15% of the total weight of the pretreated straw, enhanced degrees of enzymatic solubilization of holocellulose and cellulose fractions were observed as the content of Klason lignin decreased.     

Identification of auxotrophic mutants of the yeast Kluyveromyces marxianus by non‐homologous end joining‐mediated integrative transformation with genes from Saccharomyces cerevisiae

The isolation and application of auxotrophic mutants for gene manipulations, such as genetic transformation, mating selection and tetrad analysis, form the basis of yeast genetics. For the development of these genetic methods in the thermotolerant fermentative yeast Kluyveromyces marxianus, we isolated a series of auxotrophic mutants with defects in amino acid or nucleic acid metabolism. To identify the mutated genes, linear DNA fragments of nutrient biosynthetic pathway genes were amplified from Saccharomyces cerevisiae chromosomal DNA and used to directly transform the K. marxianus auxotrophic mutants by random integration into chromosomes through non‐homologous end joining (NHEJ). The appearance of transformant colonies indicated that the specific S. cerevisiae gene complemented the K. marxianus mutant. Using this interspecific complementation approach with linear PCR‐amplified DNA, we identified auxotrophic mutations of ADE2, ADE5,7, ADE6, HIS2, HIS3, HIS4, HIS5, HIS6, HIS7, LYS1, LYS2, LYS4, LYS9, LEU1, LEU2, MET2, MET6, MET17, TRP3, TRP4 and TRP5 without the labour‐intensive requirement of plasmid construction. Mating, sporulation and tetrad analysis techniques for K. marxianus were also established. With the identified auxotrophic mutant strains and S. cerevisiae genes as selective markers, NHEJ‐mediated integrative transformation with PCR‐amplified DNA is an attractive system for facilitating genetic analyses in the yeast K. marxianus. Copyright © 2013 John Wiley & Sons, Ltd.