A plant vacuolar protease, VPE, mediates virus-induced hypersensitive cell death

Programmed cell death (PCD) in animals depends on caspase protease activity. Plants also exhibit PCD, for example as a response to pathogens, although a plant caspase remains elusive. Here we show that vacuolar processing enzyme (VPE) is a protease essential for a virus-induced hypersensitive response that involves PCD. VPE deficiency prevented virus-induced hypersensitive cell death in tobacco plants. VPE is structurally unrelated to caspases, although VPE has a caspase-1 activity. Thus, plants have evolved a regulated cellular suicide strategy that, unlike PCD of animals, is mediated by VPE and the cellular vacuole.     

Cytokinin oxidase regulates rice grain production

Most agriculturally important traits are regulated by genes known as quantitative trait loci (QTLs) derived from natural allelic variations. We here show that a QTL that increases grain productivity in rice, Gn1a, is a gene for cytokinin oxidase/dehydrogenase (OsCKX2), an enzyme that degrades the phytohormone cytokinin. Reduced expression of OsCKX2 causes cytokinin accumulation in inflorescence meristems and increases the number of reproductive organs, resulting in enhanced grain yield. QTL pyramiding to combine loci for grain number and plant height in the same genetic background generated lines exhibiting both beneficial traits. These results provide a strategy for tailormade crop improvement.     

Antioxidant activity of peptides obtained from porcine myofibrillar proteins by protease treatment

Hydrolysates obtained from porcine myofibrillar proteins by protease treatment (papain or actinase E) exhibited high antioxidant activity in a linolenic acid peroxidation system induced by Fe(2+). Hydrolysates produced by both papain and actinase E showed higher activities at pH 7.1 than at pH 5.4. The antioxidant activity of the papain hydrolysate was almost the same as that of vitamin E at pH 7.0. These hydrolysates possessed 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity and chelating activity toward metal ions. Antioxidant peptides were separated from the papain hydrolysate by ion exchange chromatography. The acidic fraction obtained by this method exhibited higher activity than the neutral or basic fractions. Antioxidant peptides in the acidic fraction were isolated by high-performance liquid chromatography on an ODS column and shown to possess the structures DSGVT, IEAEGE, DAQEKLE, EELDNALN, and VPSIDDQEELM. The DAQEKLE peptide showed the highest activity among these peptides.     

Differentiation-inducing effects of small fruit juices on HL-60 leukemic cells

Epidemiological studies indicate that high intakes of fruits and vegetables are associated with a reduced risk of cancer, and several plant-derived drugs have been developed in medical oncology. Since only a small part of the flora has been tested for any kind of bioactivity, we chose small fruits as sources of differentiation-inducing activity against HL-60 leukemic cells. We have prepared juices from various small fruits that grow mainly in the northern part of Japan. Screening of 43 samples indicated that juices of Actinidia polygama Maxim., Rosa rugosa Thunb., Vaccinium smallii A. Gray, and Sorbus sambucifolia Roem. strongly induced differentiation of HL-60 cells to monocyte/macrophage characteristics in a concentration-dependent manner as indicated by histochemical and biochemical examinations.     

Isolation and characterization of aminopeptidase (Jc-peptidase) from Japanese cedar pollen (Cryptomeria japonica)

An aminopeptidase, Jc-peptidase, was purified from Japanese cedar pollen by seven steps, including precipitation with ammonium sulfate, ion-exchange chromatography, gel filtration, hydrophobic interaction chromatography on phenyl-agarose, and high-performance liquid chromatography. Purified Jc-peptidease has a molecular weight of 42 kDa and hydrolyzes the synthetic substrates of L-phenylalanyl-4-methylcoumaryl-7-amide (Phe-MCA) with Km = 5 x 10(-5) M, Tyr-MCA with Km = 7 x 10(-4) M, Leu-MCA with Km = 1 x 10(-3) M, and Met-MCA with Km = 1 x 10(-3) M. Other MCA analogues such as Arg-MCA or Glu-MCA failed to serve as its substrates. The activity was inhibited in the presence of phebestin, [(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoyl-L-valyl]-L-phenylalanine, with Ki = 4.7 x 10(-5) M, or bestatin, [(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoyl]-L-leucine, with Ki = 1.1 x 10(-4) M. According to amino acid sequence analysis, the N-terminal amino group seems to be blocked. The physiological function of the aminopeptidase (Jc-peptidase) has not been clarified in vivo.     

Effect of intermittent drainage in reduction of methane emission from paddy soils in Hokkaido,northern Japan

ABSTRACT

Emission of methane (CH₄), a major greenhouse gas, from submerged paddy soils is generally reduced by introducing intermittent drainage in summer, which is a common water management in Japan. However, such a practice is not widely conducted in Hokkaido, a northern region in Japan, to prevent a possible reduction in rice grain yield caused by cold weather. Therefore, the effects of intermittent drainage on CH₄ emission and rice grain yield have not been investigated comprehensively in Hokkaido. In this study, we conducted a three-year field experiment in Hokkaido and measured CH₄ and nitrous oxide (N₂O) fluxes and rice grain yield to elucidate whether the reduction in CH₄ emission can be achieved in Hokkaido as well as other regions in Japan. Four experimental treatments, namely, two soil types [soils of light clay (LiC) and heavy clay (HC) textures] and two water management [continuous flood irrigation (CF), and intermittent drainage (ID)], were used, and CH₄ and N₂O fluxes were measured throughout the rice cultivation periods from 2016 to 2018. Cumulative CH₄ emissions in 2016 were markedly low, suggesting an initially low population of methanogens in the soils presumably due to no soil submergence or crop cultivation in the preceding years, which indicates a possible reduction in CH₄ emission by introducing paddy-upland crop rotation. Cumulative CH₄ emissions in the ID-LiC and ID-HC plots were 21–91% lower than those in the CF-LiC and CF-HC plots, respectively, whereas the cumulative N₂O emissions did not significantly differ between the different water managements. The amount of CH₄ emission reduction by the intermittent drainage was largest in 2018, with a comparatively long period of the first drainage for 12 days in summer. Rice grain yields did not significantly differ between the different water managements for the entire 3 years, although the percentage of well-formed rice grains was reduced by the intermittent drainage in 2018. These results suggest that CH₄ emission from paddy fields can be reduced with no decrease in rice grain yield by the intermittent drainage in Hokkaido. In particular, the first drainage for a long period in summer is expected to reduce CH₄ emission markedly.     

Results of hyperamplification of centrosomes in naturally developing tumors of dogs.

OBJECTIVE: To evaluate results of centrosome hyperamplification in naturally developing tumors of dogs. SAMPLE POPULATION: Tumor specimens from 9 dogs with tumors (rhabdomyosarcoma, osteosarcoma, chondrosarcoma, myxosarcoma, and mammary gland tumor) and 2 canine osteosarcoma cell lines. PROCEDURE: 3 antibodies for centrosome proteins (ie, anti-gamma-tubulin, anti-BRCA1, and anti-pericentrin) were used for immunohistochemical analysis. Double immunostaining for centrosomes was used to confirm the specificity of these antibodies for centrosomes. Mutational analysis of the canine p53 gene was carried out by polymerase chain reaction-single-strand conformation polymorphism analysis, and expression of canine MDM2 protein was evaluated by use of immunohistochemical analysis, using anti-MDM2 antibody. RESULTS: Immunohistochemical analysis of dog osteosarcoma cell lines with apparent aneuploidy revealed frequent hyperamplification of centrosomes in the osteosarcoma cell lines. Similar hyperamplified centrosomes were detected in the tumor tissues from all of the 9 tumors. The frequency of cells with hyperamplified centrosomes (3 to 20/cell) in each tumor tissue ranged from 9.50 to 48.1%, whereas centrosome hyperamplification was not observed in normal lymph nodes from these dogs. In 8 of the 9 tumors, mutation of p53 gene or overexpression of MDM2, or both, was detected. CONCLUSIONS AND CLINICAL RELEVANCE: Various types of naturally developing tumors in dogs often have hyperamplification of centrosomes associated with chromosome instability. Hyperamplification of centrosomes is a novel tumor marker for use in cytologic and histologic examinations of clinical specimens obtained from dogs.     

Structural proteins of equine infectious anemia virus and their antigenic activity

Using purified equine infectious anemia (EIA) virus labeled with 3H-glucosamine or 14C-protein hydrolysate, structural proteins were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. As a result, 2 glycoproteins and 10 proteins with molecular weights (mol wt) ranging from 12,000 to 115,000 daltons were demonstrated. Of 12 structural proteins, 3 proteins, namely a glycoprotein with mol wt of 76,000 (gp76) and 2 proteins with mol wt of 25,000 (p25) and 12,000 (p12), respectively, had distinct antigenic activity from one another in immunodiffusion. Development of antibodies against gp76 and p25 was compared in infected horses. The antibody to gp76 appeared earlier and stronger than to p25 in horses infected with the homologous virus strain. The fraction with glycoproteins was found to have hemagglutinating activity which was inhibited by the serum sample from horses infected with equine infectious anemia virus.