Isolation, characterization and bioactivity of a region-specific pheromone, [Val8]sodefrin from the newt Cynops pyrrhogaster

Previous analysis of PCR products derived from total RNA from the abdominal gland of the male newt, Cynops pyrrhogaster, inhabiting the Nara area of Japan led to the identification of a gene encoding [Val(8)]sodefrin, as well as the female-attracting peptide pheromone, sodefrin. In this study, purification of this sodefrin variant from the abdominal glands of male newts from the Nara area was accomplished using gel-filtration chromatography and reversed-phase HPLC. Amino acid sequence analysis and mass spectrometry confirmed that the final product was [Val(8)]sodefrin. A full-length cDNA encoding the biosynthetic precursor of [Val(8)]sodefrin was cloned and characterized. The deduced amino acid sequence of prepro[Val(8)]sodefrin showed 86.2% identity with that of the sodefrin precursor. The [Val(8)]sodefrin variant potently attracted females from the Nara area, but the variant was much less or not effective in attracting females captured in the Niigata and Chiba areas. The term aonirin ("aoni" from "aoni-yoshi", the conventional epithet of Nara) is proposed to designate this region-specific pheromone. It is speculated that the coevolution of a novel pheromone and its complementary receptor in the Nara newts may lead to reproductive isolation and eventual differentiation into a separate species.     

Monte Carlo-based calculation of imaging plate response to <Superscript>90</Superscript>Sr in teeth: experimental validation of the required correction on sample thickness

Recently, a numerical method was proposed to correct the imaging plate (IP) response to ⁹⁰Sr concentration in tooth samples, depending on the sample thickness. This is important to quantify any ⁹⁰Sr concentration in teeth, which in turn is necessary to determine any ⁹⁰Sr incorporation of a person retrospectively. Although the final goal will be to evaluate the (inhomogeneous) spatial distribution of ⁹⁰Sr inside tooth samples precisely, the present study was restricted—as a first step—to the evaluation of ⁹⁰Sr in teeth assuming a uniform ⁹⁰Sr distribution. A numerical method proposed earlier was validated experimentally in the present study by measuring the IP response to standard sources of various thicknesses and ⁹⁰Sr concentrations. For comparison, the energy deposition of the β-rays emitted by ⁹⁰Sr in the IP—which is considered to be proportional to the IP luminescence signal—was calculated for the various sample thicknesses involved, by means of the MCNP-4C code. As a result, the measured IP response could be reproduced by the calculations within the uncertainties, depending on the thickness of the standard sources. Thus, the validity of the proposed numerical method to correct the IP response for sample thickness has successfully been demonstrated.     

Assessment of higher insect taxa as bioindicators for different logging-disturbance regimes in lowland tropical rain forest in Sabah, Malaysia

One of the serious environmental problems since the 1980s has been the conflict between the high rate of deforestation and maintenance of healthy ecosystem services and biological values in tropical forests. There is an urgent demand for setting up an appropriate environmental assessment to keep healthy ecosystem functions and biodiversity along with sustainable forest use based on ecology. In this study, we tried to assess logging-disturbance effects on the abundances of several flying insect groups (higher-taxon approach) in lowland tropical rain forest (Deramakot Forest Reserve, Sabah, Malaysia), while considering seasonal changes and vertical forest stratification. The season was the most important factor affecting the abundances of all the insect groups. Effects of logging disturbance were prominent in the understorey but obscure in the canopy. Changes in physical conditions caused by logging—possibly an increased evaporation due to solar radiation—may have decreased the abundance of desiccation-sensitive insects, especially in the understorey. There are also two probable reasons for the difference between events in the understorey and those in the canopy: (1) noise effects of various physical, environmental factors may have obscured insect responses to logging disturbance in the canopy; (2) higher spatio-temporal variation in quality and quantity of living food resources—such as leaves, flowers and fruits—provided in the canopy may have affected the abundance of their consumer insects independently of logging disturbance. Thus, this study suggests that the abundance of some insect groups at higher-taxon level, especially in the understorey, can be used as bioindicators for assessing effects of logging disturbance on the forest ecosystem.     

Different effects of galactose and mannose on cell proliferation and intracellular soluble sugar levels in <Emphasis Type="Italic">Vigna angularis</Emphasis> suspension cultures

Plant cells utilize various sugars as carbon sources for growth, respiration and biosynthesis of cellular components. Suspension-cultured cells of azuki bean (Vigna angularis) proliferated actively in liquid growth medium containing 1% (w/v) sucrose, glucose, fructose, arabinose or xylose, but did not proliferate in medium containing galactose or mannose. These two latter sugars thus appeared distinct from other sugars used as growth substrates. Galactose strongly inhibited cell growth even in the presence of sucrose but mannose did not, suggesting a substantial difference in their effects on cell metabolism. Analysis of intracellular soluble-sugar fractions revealed that galactose, but not mannose, caused a conspicuous decrease in the cellular level of sucrose with no apparent effects on the levels of glucose or fructose. Such a galactose-specific decrease in sucrose levels also occurred in cells that had been cultured together with glucose in place of sucrose, suggesting that galactose inhibits the biosynthesis, rather than uptake, of sucrose in the cells. By contrast, mannose seemed to be metabolically inert in the presence of sucrose. From these results, we conclude that sucrose metabolism is important for the heterotrophic growth of cells in plant suspension-cultures.     

MAMO, a maternal BTB/POZ-Zn-finger protein enriched in germline progenitors is required for the production of functional eggs in Drosophila

A hallmark of germline cells throughout the animal kingdom is their ability to execute meiosis. However, despite its prime importance, little is known about how germline progenitors acquire this ability. In Drosophila, the primordial germ cells (PGCs) are characterized by the inheritance of germ plasm, which contains maternal factors that have sufficient ability to direct germline development. Here, we show that a novel maternal factor, MAMO, is autonomously required in PGCs to produce functional gametes. MAMO protein which contains both a BTB/POZ (Broad Complex, Tramtrack, Bric-a-brac/Pox virus and Zinc finger) domain and C(2)H(2) zinc finger motifs is enriched in PGCs during embryogenesis. The PGCs with reduced maternal MAMO activity are able to undergo oogenesis, but fail to execute meiosis properly. In the resulting oocytes, meiosis-specific chromosomal configurations are impaired. We additionally show that the decondensation of fertilized sperm nuclei is also affected in the eggs. We propose that maternal MAMO activates downstream genes to promote specialized morphological changes of both female meiotic chromosomes and the sperm nucleus, which are critical in zygote formation.     

Expression mapping of cytotoxic T-lymphocyte antigen-2α gene transcripts in mouse brain

Cytotoxic T-lymphocyte antigen-2alpha (CTLA-2alpha), an inhibitor peptide homologous to the proregion of mouse cathepsin L, was originally discovered and expressed in mouse-activated T-cells and mast cells. Expressed recombinant CTLA-2alpha is shown to exhibit selective inhibition to cathepsin L-like cysteine proteinases. However, its in vivo targets in mammalian tissues are yet to be identified. We carried out in situ hybridization studies to examine the expression pattern of CTLA-2alpha mRNA and determine the specific cell types synthesizing CTLA-2alpha in the mouse brain. CTLA-2alpha mRNA was detected in various neuronal populations within the telencephalon in cerebral cortices, olfactory system, septum, basal ganglia, amygdala and highest levels were observed in the hippocampus. Within the diencephalon high density of positive cells was found in mediodorsal and lateral posterior thalamic nuclei and medial habenular nucleus (MHb). In the hypothalamus, high density of CTLA-2alpha mRNA labeling was seen in the suprachiasmatic nucleus (Sch), optic tract, arcuate nucleus, and median eminence. The fasciculus retroflexus and its termination in the mesencephalic interpeduncular nucleus were also densely labeled. Other mesencephalic expression sites were the superior colliculus, periaqueductal gray, paramedian raphe nucleus, and inferior colliculus. In the rhombencephalon, strong labeling was detected in the pontine, vestibular, and reticular nuclei. Intense expression was also noted within cerebellar cortex in Purkinje neurons and at a moderate level in granule cell layer, stellate, and basket cells. A possible function of this novel inhibitor peptide in relation to learning, memory, and diseases is discussed.     

Putative mechanism of the itch-scratch circle: repeated scratching decreases the cutaneous level of prostaglandin D2, a mediator that inhibits itching

In atopic dermatitis, scratching of the skin as a reaction to itching causes injury to the skin, which, in turn, further increases the itching resulting in the establishment of the so-called itch-scratch circle. We have shown that prostaglandin (PG) D2 plays an inhibitory role against pruritus in mice with atopic-like dermatitis; therefore, we examined the relationship between scratching and the cutaneous PGD2 level using an artificial scratching model with a wire brush. Mechanical scratching induced a temporary increase of the skin PGs levels (PGE2, PGD2, 6-ketoPGF1alpha, PGF2alpha). The skin PGD2 level and the ability of PGD2 production decreased at 48 h after repeated scratch, compared to that of normal skin, not so after single scratch. Immunohistochemical analysis and Western blotting revealed a decrease in the levels of cyclooxygenase-1 (COX-1) and hematopoietic PGD synthase in mechanically scratched skin. The reduced ability of the skin for PGD2 production following mechanical scratching could be caused by this decrease in the expression levels of COX-1 and PGD2 synthase. The results suggest that repeated scratching in mice decreases the ability of the skin to produce PGD2, which is an endogenous mediator that inhibits pruritus, resulting in the establishment of the itch-scratch circle.     

Development of a package-free transparent disposable biosensor chip for simultaneous measurements of blood constituents and investigation of its storage stability

A package-free transparent disposable biosensor chip was developed by a screen-printing technique. The biosensor chip was fabricated by stacking a substrate with two carbon electrodes on its surface, a spacer consisting of a resist layer and an adhesive layer, and a cover. The structure of the chip keeps the interior of the reaction-detecting section airtight until use. The chip is equipped with double electrochemical measuring elements for the simultaneous measurement of multiple items, and the reagent layer was developed in sample-feeding path. The sample-inlet port and air-discharge port are simultaneously opened by longitudinally folding in two biosensor units with a notch as a boundary. Then the shape of the chip is changed to a V-shape. The reaction-detecting section of the chip has a 1.0 microl sample volume for one biosensor unit. Excellent results were obtained with the chip in initial simultaneous chronoamperometric measurements of both glucose (r=1.00) and lactate (r=0.998) in the same samples. The stability of the enzyme sensor signals of the chip was estimated at ambient atmosphere on 8 testing days during a 6-month period. The results were compared with those obtained for an unpackaged chip used as a control. The package-free chip proved to be twice as good as the control chip in terms of the reproducibility of slopes from 16 calibration curves (one calibration curve: 0, 100, 300, 500 mg dl(-1) glucose; n=3) and 4.6 times better in terms of the reproducibility of correlation coefficients from the 16 calibration curves.     

Characteristics of copper tolerance in Yarrowia lipolytica

We discovered that a mutant strain of the dimorphic yeast Yarrowia lipolytica could grow in the yeast form in high concentrations of copper sulfate. The amount of metal accumulated by Y. lipolytica increased with increasing copper concentrations in the medium. Washing with 100 mM EDTA released at least 60% of the total metal from the cells, but about 20–25 μmol/g DW persisted, which represented about 30% of the soluble fraction of cultured cells. The soluble fraction (mainly cytosol) contained only about 10% of the total metal content within cells cultured in medium supplemented with 6 mM copper. We suggest that although a high copper concentration induces an efflux mechanism, the released copper becomes entrapped in the periplasm and in other parts of the cell wall. Washing with EDTA liberated not only copper ions, but also melanin, a brown pigment that can bind metal and which located at the cell wall. These findings indicated that melanin participates in the mechanism of metal accumulation. Culture in medium supplemented with copper obviously enhanced the activities of Cu, Zn-SOD, but not of Mn-SOD.