A mitogen-activated protein kinase NtMPK4 activated by SIPKK is required for jasmonic acid signaling and involved in ozone tolerance via stomatal movement in tobacco

The mitogen-activated protein kinase (MAPK) cascade is involved in responses to biotic and abiotic stress in plants. In this study, we isolated a new MAPK, NtMPK4, which is a tobacco homolog of Arabidopsis MPK4 (AtMPK4). NtMPK4 was activated by wounding along with two other wound-responsive tobacco MAPKs, WIPK and SIPK. We found that NtMPK4 was activated by salicylic acid-induced protein kinase kinase (SIPKK), which has been isolated as an SIPK-interacting MAPK kinase. In NtMPK4 activity-suppressed tobacco, wound-induced expression of jasmonic acid (JA)-responsive genes was inhibited. NtMPK4-silenced plants showed enhanced sensitivity to ozone. Inversely, transgenic tobacco plants, in which SIPKK or the constitutively active type SIPKK(EE) was overexpressed, exhibited greater responsiveness to wounding with enhanced resistance to ozone. We further found that NtMPK4 was expressed preferentially in epidermis, and the enhanced sensitivity to ozone in NtMPK4-silenced plants was caused by an abnormal regulation of stomatal closure in an ABA-independent manner. These results suggest that NtMPK4 is involved in JA signaling and in stomatal movement.     

Antioxidants in the serum of children with insulin-dependent diabetes mellitus

To determine whether alteration in serum antioxidant status is related to the increased oxidative stress as a cause of diabetic angiopathy, we measured both the antioxidant activity (AOA) and total peroxyl radical-trapping antioxidant parameter (TRAP), and their component individual antioxidants in serum of children with insulin-dependent diabetes mellitus (IDDM). The AOA was measured as the ability to inhibit lipid autoxidation in brain homogenates. TRAP was assayed as the ability to delay lipid peroxidation induced by an azo initiator. Antioxidants measured were ceruloplasmin, transferrin, and albumin components of AOA; and ascorbic acid, uric acid, protein sulfhydryl, and alpha-tocopherol as components of TRAP. Serum AOA appeared to be decreased in the diabetics in relation to poor glycemic control, corresponding to the decrease in transferrin and albumin. Serum haptoglobin level was also decreased in the diabetics. Similarly, the directly measured TRAP value was decreased in the diabetic serum mainly due to the decreased contribution of unidentified chain-breaking antioxidants, despite the increase in ascorbic acid and alpha-tocopherol. The decrease in both types of antioxidant activity in the diabetic serum, as new findings, suggests that a defective serum antioxidant status contributes to the increased oxidative stress in IDDM.     

Divalent cation-induced surface tension increase in acidic phospholipid membranes: Ion binding and membrane fusion

Chelation binding of divalent cations to phospholipid membranes may cause deformation in the headgroup regions of these lipid molecules. This deformation may be responsible for the observed large increase in surface tension of acidic phospholipid membranes induced by divalent cations. On the other hand, simple binding of monovalent cations without being followed by such a deformation of membrane molecules, does not result in a large surface tension increase in the membrane. A theoretical explanation for the above situation is given and the divalent cation-induced acidic phospholipid membrane fusion as well as other lipid membrane fusions are discussed in terms of the increased surface energy of membranes.     

Productivity and mineral cycling in an oak coppice forest 1. Structure and phytomass of the forest

Ecological studies were made on the structure and phytomass of the secondary coppice forest near Tokyo, which was dominated by a deciduous oak,Quercus serrata Thunb. Average height of dominant trees was about 10 m. The shoot density at the beginning of the study was 4,600 ha⁻¹ of which 89.5% belonged toQ. serrata. During the subsequent two growing seasons 8.3% of shoots, mainly small ones, died. All the tree shoots in a 10 m×10 m quadrat were cut and most of the underground parts were dug out. The phytomass calculated by the allometric relations of the dry weight of each plant organ to the square of DBH (D ²) agreed fairly well with the values directly weighed. The average phytomass of the overstory trees was 906 kg a⁻¹. The leaf area index (LAI) of the canopy was 3.85. Phytomass of the undergrowth, mainly a dwarf bamboo,Pleioblastus chino Makino, was 91.8 kg a⁻¹ with LAI of 3.46. The dead phytomass in the stand was 177 kg a⁻¹, so the phytomass alive and dead amounted to 1,170 kg a⁻¹. Heavy self-thinning of the coppice forest is discussed in relation to the rapid development of the log-normal distribution of tree sizes and to the large allometric constants for leaf and branch weight toD ².     

Intake of green-plant membrane with dietary oil suppresses postprandial hypertriglyceridemia in rats via promoting excretion of bile acids

Green-plant membrane is a phytonutrient present in green leafy vegetables at high concentration. Postprandial increases in blood triglyceride levels result in insulin resistance and type 2 diabetes. Additionally, dietary life and eating order also affect postprandial hypertriglyceridemia. In this study, the effects of once-daily intake of green-plant membrane with dietary oil on postprandial hypertriglyceridemia were investigated in vitro and in vivo. In vitro, green-plant membrane bound hydrophobic bile acids but did not inhibit pancreatic lipase activity. Following the administration, green-plant membrane with dietary oil in rats, oral fat tolerance tests, increases in serum triglycerides levels were significantly reduced. Moreover, fecal total lipid and bile acid volumes were significantly increased in rats that administered 200 mg/mL green-plant membrane. These results suggest that green-plant membrane with dietary oil inhibits dietary fat absorption via promotion of bile acid excretion in feces and the effectiveness of eating green-plant membrane, such as green leafy vegetables, with meals.     

Radiation and fermentation treatment of cellulosic wastes

The effect of radiation pasteurization of sugar cane bagasse and rice straw and fermentation using various strains of fungi were studied for upgrading of cellulosic wastes. The initial contamination by fungi and aerobic bacteria both in bagasse and straw was high. The doses of 30 kGy for sterilization and 8 kGy for elimination of fungi were required. Irradiation effect showed that rice straw contained comparatively radioresistant microorganisms. It was observed that all the fungi (Hericium erinacium, Pleurotus djamor, Ganoderma lucidum, Auricularia auricula, Lentinus sajor-caju, Coriolus versicolor, Polyporus arcularius, Coprinus cinereus) grow extending over the entire substrates during one month after inoculation in irradiated bagasse and rice straw with 3% rice bran and 65% moisture content incubated at 30°C. Initially, sugar cane bagasse and rice straw substrates contained 39.4% and 25.9% of cellulose, 22.9% and 26.9% of hemicellulose, and 19.6% and 13.9% of lignin + cutin, respectively. Neutral detergent fibre (NDF) values decreased significantly in sugar cane bagasse fermented byG. lucidum, A. auricula andP. arcularius, and in rice straw fermented by all the 8 strains of fungi. Acid detergent fibre (ADF) values also decreased in bagasse and rice straw fermented by all the fungi.P. arcularius, H. erinacium, G. lucidum andC. cinereus were found to be the most effective strains for delignification of sugar cane bagasse.     

Atkinsiella awabi sp. nov. isolated from stocked abalone,Haliotis sieboldii

A fungal disease in the abalone,Haliotis sieboldii, stocked in Yamaguchi Prefecture, Japan, showed external signs of infection of tubercle-like swelling on the mantle and melanized lesions on the peduncle. The fungus responsible was isolated by inoculating materials taken from the lesions onto PYGS agar with streptomycin sulphate and ampicillin, and incubation at 20°C. For morphological observation and spore formation study, the fungus was transferred respectively into PYGS broth and sterilized artificial seawater and incubated at 20°C. Resulting, hyphae were stout, irregular, branched, 16–140µm diam, sporadically consisting of dense cytoplasmic swollen hyphae. Sporangia were formed through the formation of septa and lateral or terminal discharge tubes which were wavy or coiled. Zoospores were pyriform, biflagellate and diplanetic. The encysted spore generally developed a hairlike filament with globular enlarged tip in PYGS broth. Direct germination without filament formation also occurred occasionally. This fungus was identified as belonging to the genusAtkinsiella, and was designatedAtkinsiella awabi sp. nov. The fungus was exclusively a marine fungus and grew best in shrimp extract medium at 20°C. Five chemicals were tested for their effects against fungal zoospores.     

Identification of OS-2 MAP kinase-dependent genes induced in response to osmotic stress, antifungal agent fludioxonil, and heat shock in Neurospora crassa

Two-component signal transduction comprising of OS-1 (histidine kinase), OS-4 (MAPKK kinase), OS-5 (MAPK kinase), and OS-2 (MAP kinase) plays an important role in osmotic regulation in Neurospora crassa. To identify the genes regulated downstream of OS-2 MAP kinase, quantitative real-time RT-PCR analysis was conducted in selected genes based on Hog1 MAP kinase regulated genes in yeast. In response to osmotic stress and fludioxonil, expression of six genes that for glycerol synthesis (gcy-1, gcy-3, and dak-1), gluconeogenesis (fbp-1 and pck-1), and catalase (ctt-1) was activated in the wild-type strain, but not in the os-2 mutant. A heat shock treatment also induced their expression in the same way. Consisting with the gene expression, the enzyme activity of glycerol dehydrogenase, but not glycerol-3-phosphate dehydrogenase, was increased in response to osmotic stress and fludioxonil in the wild-type strain. OS-2 was phosphorylated by the OS-1 cascade in response to relatively low osmotic stress and fludioxonil. However, OS-2 phosphorylation by heat shock and a higher osmotic stress was found in the os-1 mutant normally but not in the os-4 and os-5 mutants. These results suggested that non-OS-1 signaling activates OS-2 in an OS-4-dependent manner in such conditions.     

Photosynthetic electron transport inhibition by 2-substituted 4-alkyl-6-benzylamino-1,3,5-triazines with thylakoids from wild-type and atrazine-resistant Chenopodium album

The effect of 2-benzylamino-1,3,5-triazines on photosynthetic electron transport (PET) was measured with thylakoids isolated from atrazine-resistant, wild-type Chenopodium album, and spinach to find novel 1,3,5-triazine herbicides bearing a strong PET inhibition. The PET inhibition assay with Chenopodium (wild-type and resistant), yielded a resistance ratio (R/W = I50 (resistant)/I50 (wild-type)) of 324 for atrazine while for benzylamino-1,3,5-triazine derivatives of diamino-1,3,5-triazines a R/W of 11 to 160 was found. The compounds having a benzylamino group at one of the amino groups in the diamino-1,3,5-triazines have a resistant ratio down to one half to 1/30 of the atrazine value. The average resistance ratio of 21 benzylamino derivatives of monoamino-1,3,5-triazines was found to be about 4.0. The inhibition of 21 benzylamino-1,3,5-triazines assayed with atrazine-resistant Chenopodium thylakoids, indicated by pI50 (R)-values, correlated well with the PET inhibition pI50 (W) of wild-type thylakoids from Chenopodium.     

Diverse reaction behaviors of artificial ubiquinones in mitochondrial respiratory complex I

The ubiquinone (UQ) reduction step catalyzed by NADH-UQ oxidoreductase (mitochondrial respiratory complex I) is key to triggering proton translocation across the inner mitochondrial membrane. Structural studies have identified a long, narrow, UQ-accessing tunnel within the enzyme. We previously demonstrated that synthetic oversized UQs, which are unlikely to transit this narrow tunnel, are catalytically reduced by native complex I embedded in submitochondrial particles but not by the isolated enzyme. To explain this contradiction, we hypothesized that access of oversized UQs to the reaction site is obstructed in the isolated enzyme because their access route is altered following detergent solubilization from the inner mitochondrial membrane. In the present study, we investigated this using two pairs of photoreactive UQs (pUQ_m-1/pUQ_p-1 and pUQ_m-2/pUQ_p-2), with each pair having the same chemical properties except for a ∼1.0 Å difference in side-chain widths. Despite this subtle difference, reduction of the wider pUQs by the isolated complex was significantly slower than of the narrower pUQs, but both were similarly reduced by the native enzyme. In addition, photoaffinity-labeling experiments using the four [¹²⁵I]pUQs demonstrated that their side chains predominantly label the ND1 subunit with both enzymes but at different regions around the tunnel. Finally, we show that the suppressive effects of different types of inhibitors on the labeling significantly changed depending on [¹²⁵I]pUQs used, indicating that [¹²⁵I]pUQs and these inhibitors do not necessarily share a common binding cavity. Altogether, we conclude that the reaction behaviors of pUQs cannot be simply explained by the canonical UQ tunnel model.