Role for piRNAs and noncoding RNA in de novo DNA methylation of the imprinted mouse Rasgrf1 locus

Genomic imprinting causes parental origin-specific monoallelic gene expression through differential DNA methylation established in the parental germ line. However, the mechanisms underlying how specific sequences are selectively methylated are not fully understood. We have found that the components of the PIWI-interacting RNA (piRNA) pathway are required for de novo methylation of the differentially methylated region (DMR) of the imprinted mouse Rasgrf1 locus, but not other paternally imprinted loci. A retrotransposon sequence within a noncoding RNA spanning the DMR was targeted by piRNAs generated from a different locus. A direct repeat in the DMR, which is required for the methylation and imprinting of Rasgrf1, served as a promoter for this RNA. We propose a model in which piRNAs and a target RNA direct the sequence-specific methylation of Rasgrf1.     

Element interconnections in Lotus japonicus: A systematic study of the effects of element additions on different natural variants

Lotus japonicus was used to study the distribution and interconnections of 15 elements in plant tissues, including essential and non-essential elements: boron (B), sodium (Na), magnesium (Mg), potassium (K), calcium (Ca), manganese (Mn), iron (Fe), nickel (Ni), copper (Cu), zinc (Zn), arsenic (As), strontium (Sr), molybdenum (Mo), cadmium (Cd) and cesium (Cs). Large amounts of B and Ca accumulated in plant leaves, while Fe, Na, Ni, As and Cd tended to mainly occur in the roots, and Mo was the only element to accumulate in the stems. The elemental compositions within plants were severely disturbed by treatment with toxic elements. Competition between element pairs in the same group (e.g. K and Cs; Ca and Sr) was not found. Iron, Cu and Zn accumulation were induced by Cd and Ni addition. When natural variants grew in a nutrition solution with subtoxic levels of As, Cd, Cs, Ni, Mo and Sr, intriguing relationships between the elements (such as Fe, As and K; Mg and Ni; Mn and Ca) were revealed using principal-component analysis. This study on the plant ionome offers detailed information of element interactions and indicates that chemically different elements might be closely linked in uptake or translocation systems.     

Several possible spawning sites of the Japanese eel determined from collections of their eggs and preleptocephali

The spawning area of the Japanese eel is located at the southern part of the West Mariana Ridge in the western North Pacific, but their spawning events have not been observed. To further understand Japanese eel spawning ecology, an interdisciplinary research survey by the R/V NATSUSHIMA (NT14-09, 14 May–4 June 2014) was conducted to detect spawning sites based on the seamount, salinity front, new moon and third quadrant (spawning south of front, west of ridge) hypotheses. Attempts were made to film spawning events with underwater camera systems and to consider if eels might be detected in hydroacoustic observations. Although no Japanese eels or spawning events were video-recorded and no eel aggregations could be clearly identified acoustically, three eggs were collected at two stations in the third quadrant region at or just south of 13° N on 26 and 27 May. Three or four days later, newly hatched preleptocephali were collected at two stations far to the south, including 224 at a station > 160 km southwest of the egg catches, and a few preleptocephali were caught at two stations closer to the egg stations. The eggs and southern preleptocephali were from discrete spawning events, which indicated that at least two spawning sites occurred in May 2014.

     

Relationship between <Superscript>137</Superscript>Cs concentration and potassium content in stem wood of Japanese cedar (<Emphasis Type="Italic">Cryptomeria japonica</Emphasis>)

To utilize forest resources in areas affected by fallout from the Fukushima Daiichi Nuclear Power Plant accident, it is important to understand the mechanisms of ¹³⁷Cs movement through the stem wood of contaminated trees. Understanding the mechanism of absorption and migration of ¹³⁷Cs to stem wood is necessary for clues to the future prediction of the transition of ¹³⁷Cs to xylem. In the present study, radial variations in ¹³⁷Cs concentration were investigated in Japanese cedar (Cryptomeria japonica D. Don) trees collected 1 year and 10 months after the accident. Additionally, the relationship between ¹³⁷Cs concentration and potassium (K) content was established. Trees with a higher moisture content and lower lightness value in heartwood tended to have a higher ¹³⁷Cs concentration in the heartwood. In these trees, ¹³⁷Cs concentration peaked at the heartwood–sapwood boundary and gradually decreased toward the pith. By contrast, K content within the heartwood remained nearly constant along the radial direction. The heartwood-to-sapwood ratio of ¹³⁷Cs concentration was significantly positively correlated with that of K content. Based on these results, we suggest that ¹³⁷Cs movement from sapwood to heartwood might be related to the K content ratio of heartwood and sapwood.     

Changes in the milk composition of okapi (Okapia johnstoni) during the first 6 months of lactation

We collected 17 milk samples at 1 day to 6 months post partum from one captive okapi (Okapia johnstoni). Milks were examined for nutrient composition, including water, fat, crude protein, carbohydrates, ash and five minerals. The average values of the concentrations were 79.2 ± 3.0% SD water, 8.5 ± 3.4% fat, 7.5 ± 1.1% crude protein, 3.6 ± 0.6% carbohydrate, 1.2 ± 0.1% ash, 263.7 ± 47.3 mg/100 g calcium, 183.0 ± 40.0 mg/100 g phosphorus, 95.2 ± 17.6 mg/100 g sodium, 74.4 ± 34.3 mg/100 g potassium and 28.4 ± 4.4 mg/100 g magnesium, respectively. The protein concentration increased at late lactation (P < 0.01). The contents of sodium and potassium were rather high in the early lactation period, while those of calcium and phosphorus were rather low in the early lactation period. During the course of lactation in the first week post partum and the protein concentration was high at 1 day post partum. We compare our findings with the American Association of Zoos & Aquariums's Okapi Species Survival Plan Hand‐rearing Protocol, and recommend that nutrient composition of the formula for okapi calves is developed using the present study as a guide.     

Regolith migration and sorting on asteroid Itokawa

High-resolution images of the surface of asteroid Itokawa from the Hayabusa mission reveal it to be covered with unconsolidated millimeter-sized and larger gravels. Locations and morphologic characteristics of this gravel indicate that Itokawa has experienced considerable vibrations, which have triggered global-scale granular processes in its dry, vacuum, microgravity environment. These processes likely include granular convection, landslide-like granular migrations, and particle sorting, resulting in the segregation of the fine gravels into areas of potential lows. Granular processes become major resurfacing processes because of Itokawa's small size, implying that they can occur on other small asteroids should those have regolith.     

Long-range interactions within a nonnative protein

Protein folding and unfolding are coupled to a range of biological phenomena, from the regulation of cellular activity to the onset of neurodegenerative diseases. Defining the nature of the conformations sampled in nonnative proteins is crucial for understanding the origins of such phenomena. We have used a combination of nuclear magnetic resonance (NMR) spectroscopy and site-directed mutagenesis to study unfolded states of the protein lysozyme. Extensive clusters of hydrophobic structure exist within the wild-type protein even under strongly denaturing conditions. These clusters involve distinct regions of the sequence but are all disrupted by a single point mutation that replaced residue Trp62 with Gly located at the interface of the two major structural domains in the native state. Thus, nativelike structure in the denatured protein is stabilized by the involvement of Trp62 in nonnative and long-range interactions.     

Clinico-pathological findings in peripartum dairy cows fed anion salts lowering the dietary cation-anion difference: involvement of serum inorganic phosphorus, chloride and plasma estrogen concentrations in milk fever

In our previous study, it was demonstrated that the administration of anion salts, which slightly lower the dietary cation-anion difference (DCAD), in the prepartum period is safe and effective for preventing milk fever in multiparous cows. In the present study, several clinico-pathological constituents in serum and urine, which might be related to milk fever, were analyzed using stored samples from the previous study to identify clinico-pathological parameters for easily evaluating the efficacy of lowering DCAD and to further investigate the mechanism by which lowering DCAD prevents milk fever. Among the parameters analyzed in the present study, inorganic phosphorus (iP) was involved in milk fever because the serum concentration and urinary excretion of iP were significantly higher in the group of primiparous cows (heifer group), which did not develop hypocalcemia, than those in other groups of multiparous cows. Serum chloride concentrations in the heifer group and the group of multiparous cows fed anion salts (anion group) tended to remain higher than those in other control groups of multiparous cows suggesting that serum chloride concentration may be utilized for evaluating the status of metabolic acidosis and the efficacy of lowerng DCAD in dairy cows fed anion salts. In addition, plasma estradiol-17beta concentration in the heifer group tended to be lower at parturition compared with that in other multiparous groups suggesting that estrogen known as a potent inhibitor of bone resorption may be involved in developing milk fever.     

Preventive effect of mildly altering dietary cation-anion difference on milk fever in dairy cows

In the present study, we examined whether mildly altering dietary cation-anion difference (DCAD) contributes to the prevention of milk fever in dairy cows. Thirty multiparous cows and ten primiparous cows (heifer group) were used in this study and the multiparous cows were randomly divided into three groups of ten animals each (anion, non-anion and control groups). The cows in the anion group were given supplemental salts that slightly lowered DCAD. These salts consisted of 115 g of CaCO3, 42 g of CaHPO4, 65 g of MgSO4 x 7 H2O and 80 g of CaCl2 x 2 H2O as a daily dose for each cow, using a catheter from 21 days before the expected date of parturition until parturition. The cows in the non-anion group were given only the same Ca, Mg and ip supplement but no sulfate and chloride salts as that in the anion group. The cows in the control and heifer groups were not given any additional supplement. The incidence of hypocalcemia in the anion group decreased to approximately half of those in the non-anion and control groups, while the heifer group did not develop hypocalcemia at all. In addition, the number of days spent for the treatment of hypocalcemia and the number of drug bottles (calcium borogluconate solution) used for the treatment decreased to less than half in the anion group compared with those in the non-anion and control groups. At parturition, the serum Ca concentration in the control (6.2 +/- 1.9 mg/dl, mean +/- standard deviation) and non-anion groups (6.4 +/- 1.7 mg/dl) were significantly lower than that in the heifer group (8.3 +/- 0.4 mg/dl), and the level in the anion group was intermediate (7.3 +/- 1.3 mg/dl). The change in ionized Ca concentration was almost the same as that in serum Ca concentration, but only the concentration in the anion group tended to increase slightly from a week before parturition and was significantly higher than that in all other groups three days before parturition. Urinary pH in the anion group was maintained at a mildly acidic level (6.8-7.0) for the last two weeks before parturition, compared with those in the control (7.3-7.5) and non-anion groups (7.9-8.1), and similar to that in the heifer group (6.3-7.3). The urinary Ca excretion was the highest in the anion group among all groups during the prepartum period. There were no specific changes in the excretion of parathyroid hormone and 1,25-dihydroxyvitamin D in all groups of multiparous cows while the levels of these hormones remained low in the heifer group throughout the experimental period. The data in the present study indicates that the administration of anion salts that slightly lowered DCAD in the preparum period was effective for preventing milk fever in multiparous cows. Safe and mild metabolic acidosis induced by the anion salts could be evaluated by urinary pH (6.8-7.0), and might increase the responsiveness to Ca requirement at parturition through some complex mechanisms unrelated to the excretion of Ca-related hormones. In addition, it was clarified that primiparous cows have a high potential to respond to sudden Ca demand unrelated to hormone excretion, and their Ca metabolism was in some respects similar to that in multiparous cows fed anion salts. Therefore, manipulating mildly DCAD is expected to be an effective, safe and natural method for preventing milk fever in dairy cows.     

Increase in soil pH due to Ca-rich organic matter application causes suppression of the clubroot disease of crucifers

Clubroot disease of cruciferous plants caused by the soil-borne pathogen Plasmodiophora brassicae is difficult to control because the pathogen survives for a long time in soil as resting spores. Disease-suppressive and conducive soils were found during the long-term experiment on the impact of organic matter application to arable fields and have been studied to clarify the biotic and abiotic factors involved in the disease suppression. The fact that a large amount of organic matter, 400 t ha⁻¹ yr⁻¹ farmyard manure (FYM) or 100 t ha⁻¹ yr⁻¹ food factory sludge compost (FSC), had been incorporated for more than 15 yr in the suppressive soils and these soils showed higher pH and Ca concentration than the disease conducive soil led us to hypothesize that an increase in soil pH due to the long-term incorporation of Ca-rich organic matter might be the primary cause of the disease suppression. We have designed a highly reproducible bioassay system to examine this hypothesis. The suppressive and conducive soils were mixed with the resting spores of P. brassicae at a rate of 10⁶ spore g⁻¹ soil, and Brassica campestris was grown in a growth chamber for 8 d. The number of root hair infections was assessed on a microscope. It was found that the incorporation of FYM and FSC at 2.5% (w/w) to the conducive soil suppressed the infection and that the finer particles (?5 mm) of FSC inhibited the infection and increased soil pH more effectively. Neutralization of the conducive soil by Ca(OH)₂, CaCO₃ and KOH suppressed the infection, but the effectiveness of KOH was less than those of Ca(OH)₂ and CaCO₃. Acidification of the suppressive soils by H₂SO₄, promoted the infection. The involvement of soil biota in the disease suppression was investigated using the sterilized (γ-ray irradiation) suppressive soils with respect to soil pH. The γ-ray irradiation promoted the infection at pH 5.5, but no infection was observed at pH 7.4 irrespective of the sterilization status. All these observations suggest that soil pH is a major factor in disease suppression by organic matter application and that Ca and soil biota play certain roles in the suppression under the influence of soil pH.