Protein Kinase Inhibitors Inhibit Stimulation of Inositol Phospholipid Turnover and Induction of Phenylalanine Ammonia-Lyase in Fungal Elicitor-Treated Tobacco Suspension Culture Cells

Elicitor prepared from Phytophthora nicotianae stimulated inositolphospholipid turnover and induced phenylalanine ammonia-lyaseactivity in tobacco suspension culture cells [Kamada and Muto(1994) Plant Cell Physiol. 35: 397]. Protein kinase inhibitors,K252a and staurosporine inhibited both responses. These resultssuggest that inositol phospholipid turnover plays an importantrole in PAL induction through protein kinases. In addition,their mode of inhibition were different, proposing that severaltypes of protein kinases are involved in these elicitor-inducedresponses. ¹Present address: The Johns Hopkins University School of Hygieneand Public Health, 615 N. Wolfe St., Baltimore, Maryland 21205,U.S.A. ²Present address: Nagoya University BioScience Center and GraduateSchool of Agricultural Sciences, Nagoya University, Chikusa-ku,Nagoya, 464-01 Japan.     

Orthovanadate Suppresses Accumulation of Phenylalanine Ammonia-Lyase mRNA and Chalcone Synthase mRNA in Pea Epicotyls Induced by Elicitor from Mycosphaerella pinodes

Orthovanadate delayed accumulation of mRNAs encoding phenylalanineammonia-lyase and chalcone synthase in pea epicotyls inducedby an elicitor from Mycosphaerella pinodes. However, accumulationof mRNA for a putative P-type ATPase was not affected. The relationshipbetweenthe ATPase and defense responses is discussed. ³Present address: Plant Pathology Laboratory, School of Agriculture,Nagoya University, Chikusa, Nagoya, 464-01 Japan.     

Elicitor Activity of a Fungal Product Assessed at the Single-Cell Level by a Novel Gel-Bead Method

Elicitor from Erysiphe pisi was incorporated into gel beads.Individual beads were placed on single cells from barley coleoptiles.The elicitor induced unusual cytoplasmic responses and temporaryresistance to infection in coleoptile cells. The technique isapplicable to assessment of elicitor activity at the single-celllevel. ¹Contribution no. 118 from the Laboratory of Plant Pathology,Mie University. ²Present address: Laboratory of Plant Pathology & GeneticEngineering, College of Agriculture, Okayama University, Okayama,700 Japan     

Induction of Phenylalanine Ammonia-Lyase Activation and Isoflavone Glucoside Accumulation in Suspension-Cultured Cells of Red Bean, Vigna angularis, by Phytoalexin Elicitors, Vanadate, and Elevation of Medium pH

Treatment of suspension-cultured cells of red bean, Vigna angularis,with nigeran resulted in an accumulation of isoflavone glucosides,such as daidzein 7-O-ß-D-glucoside, daidzein 7,4'-di-O-ß-D-glucoside,and 2'-hydroxydaidzein 7,4'-di-O-ß-D-glucoside, whichwas accompanied by a transient increase in the activity of phenylalanineanimonia-lyase (PAL). Similar effects were also seen with otherphytoalexin elicitors, such as RNase A and cell wall componentsof Phytophthora megasperma var. sojae. Interestingly, the accumulation of isoflavone glucosides andthe transient increase in PAL activity were induced also byvanadate, a specific inhibitor of plasma membrane adenosinetriphosphatase. K₃PO₄ showed similar effects, but this was ascribedto the elevation of medium pH caused by adding this basic salt.In fact, merely raising the pH of the medium was found to besufficient for the induction of PAL activity. Experiments usinginhibitors showed that the induction depends on RNA and proteinsyntheses. The results are discussed in relation to the possiblemechanism of action of phytoalexin elicitors. ¹ Present address: Laboratory of Biochemistry, Faculty of Agriculture,Nagoya University, Furocho, Chikusa, Nagoya 464, Japan.     

Induction of Cytosolic Acetyl-Coenzyme A Carboxylase in Pea Leaves by Ultraviolet-B Irradiation

Levels of subunits of two acetyl-coenzyme A carboxylases werehigh in small leaves of Pisum sativum, decreased with growth,and remained constant in fully expanded leaves. Irradiationof fully expanded leaves induced the cytosolic isozyme only.This result suggests a key role for the cytosolic enzyme inprotection against UV-B. ¹Present address: Laboratory of Molecular Genetics, BiotechnologyInstitute, Akita Prefectural College of Agriculture, 2-2 Minami,Ohgata, Akita, 010-04 Japan ²Present address: Laboratory of Plant Molecular Biology, Schoolof Agricultural Sciences Nagoya University, Nagoya, 464-01 Japan     

Studies on anaerobic biphasic growth of a denitrifying bacterium, Pseudomonas stutzeri

Growth of Pseudomonas stutzeri(VAN NIEL strain) in the presenceof a limiting amount of nitrate under anaerobic conditions ischaracterized by 2 logarithmic phases separated distinctly byan intermediate phase where the growth rate is very low. Inthe first logarithmic phase nitrate is reduced stoichiometricallyto nitrite stage, and in the second phase nitrite is reducedto nitrogen gas. The nitrite reducing activity of cells in the second growthphase is 3–4 times higher than that of cells in the firstphase. The rise in nitrite reducing activity is correlated witha remarkable increase in the content of cytochromes a₂ and c-552. ¹Present address: Department of Biochemistry, Hiroshima UniversitySchool of Dentistry, Hiroshima, Japan. ²Present address: Institute of Molecular Biology, Faculty ofScience, Nagoya University, Nagoya, Japan. (Received June 16, 1969; )     

Isolation of a new phytoalexin-like compound, ipomeamaronol, from black-rot fungus infected sweet potato root tissue, and its structural elucidation

A new phytoalexin-like compound was isolated from sweet potatoroot tissue infected by the black-rot fungus, Ceratocystis fimbriata.Its chemical structure was similar to ipomeamarone, and thecompound was identified as 14-hydroxy-ipomeamarone and calledipomeamaronol. ¹This paper constitutes Part 105 of the Phytopathological Chemistryof Sweet Potato with Black Rot and Injury. ²Present address: Institute for Biochemical Regulation, Facultyof Agriculture, Nagoya University, Nagoya. (Received November 7, 1972; )     

Effects of Exogenous Auxin on the Regulation of Elongation Growth of Excised Segments of Vigna Hypocotyls under Osmotic Stress

IAA applied simultaneously with osmotica greatly enhanced theadaptive recovery of the elongation growth of segments of Vignahypocotyls during osmotic stress irrespective of whether ornot absorbable solutes were present. IAA stimulated both thesurface pump and the xylem pump, which have been shown to bestimulated by osmotic stress and to control the yielding ofthe cell wall and the absorption of solutes. Thus, wall extensibilityand the effective turgor were further enhanced under osmoticstress in the presence of IAA. These results indicate that thesimultaneous presence of IAA can reduce the inhibition of growthby osmotic stress, and they support numerical predictions basedon the apoplast canal model. The mechanism involved in the rapidrecovery of growth is discussed. ¹ Present address: Research Centre, Guangxi Agricultural University,Xiu Ling Rd., Nanning, Guangxi 530005 China. ² Present address: Biology Institute, Department of GeneralEducation, Nagoya University, 1 Furo-cho, Chikusa-ku, Nagoya,464 Japan. ³ Present address: Graduate School of Integrated Science, YokohamaCity University, 22-2, Seto, Kanazawa-ku, Yokohama, 236 Japan.     

Metabolism of p-Hydroxybenzoate via Hydroxyquinol by Trichosporon cutaneum WY2-2: Characterization of the Pathway using Superoxide Dismutase as a Stabilizer of Hydroxyquinol

Trichosporon cutaneum WY2-2 was shown to metabolize p-hydroxybenzoatevia protocatechuate and hydroxyquinol. Using superoxide dismutaseas a stabilizer of hydroxyquinol, the conversion of protocatechuateto hydroxyquinol and the ring fission process of hydroxyquinolwere confirmed. Hydroxyquinol was chemically identified as theproduct of protocatechuate hydroxylase reaction. Partially purifiedprotocatechuate hydroxylase was highly specific for protocatechuate;its K_m values for protocatechuate and NADH were 17.6 and 12.4µM, respectively. It catalyzed equimolar CO₂ formation,NADH oxidation and O₂ consumption from protocatechuate. Hydroxyquinoldioxygenase was highly specific for hydroxyquinol, with a K_mof 2.9 µM. ¹A preliminary account of this work was presented at the 81stMeeting of the Chubu-branch of Agricultural Chemical Societyof Japan, Gifu, October, 1980. ²Present address: Biological Institute, Faculty of Science,Nagoya University, Nagoya 464, Japan. ³Present address: Shin Nihon Chemical Co. Ltd... 19-10, Showa-cho,Anjoh, Aichi 446, Japan. (Received November 15, 1985; Accepted August 27, 1986)     

Inhibition of Hydroxyquinol Auto-Oxidation by a Cell Extract of Trichosporon cutaneum

A cell extract from acetate-grown Trichosporon cutaneum WY2-2inhibited auto-oxidation of phenolics, especially that of hydroxyquinol.It prevented auto-oxidation of hydroxyquinol without directinteraction with hydroxyquinol. Bovine erythrocyte superoxidedismutase had similar characteristics as the cell extract, andthe elution patterns of superoxide dismutase activity and ofthe inhibitory activity to hydroxyquinol auto-oxidation froma Sephadex G-150 column coincided. These results indicate thatthe inhibitory activity in the cell extract is mainly due tosuperoxide dismutase. High activity of superoxide dismutase(20–30 unit/mg protein) and its isozyme profiles suggestan intimate relation between the regulation of superoxide dismutaseand catabolism of phenolics via hydroxyquinol. ¹Present address: Biological Institute, Faculty of Science,Nagoya University, Nagoya 464, Japan. ²Present address: Shin Nihon Chemical Co. Ltd., 19-10, Showa-cho,Anjoh, Aichi 446, Japan. (Received November 15, 1985; Accepted July 3, 1986)     

Localization and Binding Characteristics of a High-Affinity Binding Site for N--Acetylchitooligosaccharide Elicitor in the Plasma Membrane from Suspension-Cultured Rice Cells Suggest a Role as a Receptor for the Elicitor Signal at the Cell Surface

A high-affinity binding site for N-acetylchitooligosac-chlarideelicitor was found to localize in the plasma membrane from suspension-culturedrice cells. Binding kinetics as well as the specificity of thisbinding site corresponded well with the behavior of the ricecells to the editor. These characteristics suggest that thebinding site represents a functional receptor for N-acetylchitooligosaccharideelicitor in rice. ²Present address: Okinawa Prefectural Livestock ExperimentalStation, 2009-5 Shoshi, Nakijin-son, Okinawa, 905-04 Japan. ³Present address: School of Hygiene and Public Health, The JohnsHopkins University, 615 North Wolfe Street, Baltimore, Maryland,21205 U.S.A. ⁴Present address: University of Tenessee, Microbiology, knoxville,Tennessee, 37996 U.S.A.     

Synergistic Function of rolB, rolC, ORF13 and ORF14 of TL-DNA of Agrobacterium rhizogenes in Hairy Root Induction in Nicotiana tabacum

Comparison of the frequency of rooting in the tobacco leaf segmentsinoculated with Agrobacterium tumefaciens harboring variouscombinations of rolB, rolC, ORF13 and ORF14 of TL-DNA of Riplasmid (pRiHRI) revealed that the genes differ in their functionto stimulate adventitious root induction. A single gene rolBinduced roots, while rolC, ORF13 and ORF14 independently promotedthe root induction by the rolB gene. The effects of these geneson the rolB-mediated rooting were in the order of ORF13>rolCORF14. Present address: Laboratory of Phylogenetic Botany, Departmentof Biology, Chiba University, 1-33 Yayoi-cho, Inage-ku, Chiba,263-8522 Japan. ² Present address: Department of Chemical and Biological Sciences,Faculty of Science, Japan Women's University, 2-8-1 Mejirodai,Bunkyo-ku, Tokyo, 112-8681 Japan.     

Distribution of Fallover in the Carboxylase Reaction and Fallover-Inducible Sites among Ribulose 1,5-Bisphosphate Carboxylase/Oxygenases of Photosynthetic Organisms

The biphasic reaction course, fallover, of carboxyla-tion catalysedby ribulose 1,5-bisphosphate carboxylase/ox-ygenase (RuBisCO)has been known as a characteristic of the enzyme from higherland plants. Fallover consists of hysteresis in the reactionseen during the initial several minutes and a very slow suicideinhibition by inhibitors formed from the substrate ribulose-l,5-bisphosphate(RuBP). This study examined the relationship between occurrenceof fallover and non-catalytic RuBP-binding sites, and the putativehysteresis-inducible sites (Lys-21 and Lys-30S of the largesubunit in spinach RuBisCO) amongst RuBisCOs of a wide varietyof photosynthetic organisms. Fallover could be detected by followingthe course of the carboxylase reaction at 1 mM RuBP and thenon-catalytic binding sites by alleviation of fallover at 5mM RuBP. RuBisCO from Euglena gracilis showed the same linearreaction course at both RuBP concentrations, indicating an associationbetween an absence of fallover and an absence of the non-catalyticbinding sites. This was supported by the results of an equilibriumbinding assay for this enzyme with a transition state analogue.Green macroalgae and non-green algae contained the plant-type,fallover enzyme. RuBisCOs from Conjugatae, Closterium ehrenbergii,Gona-tozygon monotaenium and Netrium digitus, showed a muchsmaller decrease in activity at 1 mM RuBP than the spinach enzymeand the reaction courses of these enzymes at 5 mM RuBP werealmost linear. RuBisCO of a primitive type Conjugatae, Mesotaeniumcaldariorum, showed the same linear course at both RuBP concentrations.Sequencing of rbcL of these organisms indicated that Lys-305was changed into arginine with Lys-21 conserved. ⁷ On leave from Research and Development Center, Unitika Ltd.,23 Kozakura, Uji, Kyoto, 611 Japan. ⁸ Present address: Department of Applied Biological Chemistry,Faculty of Agriculture, Tohoku University, Tsutsumidori-Ama-miyamachi, Sendai, 981 Japan. ⁹ Present address: National Institute for Basic Biology, Myodaiji,Okazaki, 444 Japan. ¹⁰ Present address: Department of Environmental Biology, TokyoPharmaceutical University, Hachioji, Tokyo, 192-03 Japan.     

Purification of Sulphite-Cytochrome c Reductase of Thiobacillus novellus and Reconstitution of Its Sulphite Oxidase System with the Purified Constituents

Sulphite-cytochrome c reductase (sulphite: ferricytochrome coxidoreductase, EC 1.8.2.1 [EC] ) derived from Thiobacillus novelluswas purified by chromatography on a DEAE-cellulose column andby gel filtration with a Sephadex G-100 column. Although thereductase thus purified moved as a single band both in gel filtrationand in isoelectric focusing it was always split into two bandsby polyacrylamide gel electrophoresis; the one had the enzymaticactivity and showed absorption spectrum of cytochrome, whilethe other had no activity and was colourless, in contrast withthe results reported by Charles and Suzuki [(1966) Biochim.Biophys. Acta 128: 522]. The enzymatic properties of the purifiedreductase were almost the same as those of the enzyme obtainedby Charles and Suzuki. Cytochrome c-551 free of the reductase activity was obtained.Its molecular weight was determined to be 23,000 by polyacrylamidegel electrophoresis in the presence of sodium dodecyl sulphate.The cytochrome seemed to exist in the organism as a complexwith the reductase or a subunit of the enzyme. In the stateof the complex with the enzyme, the cytochrome was reduced veryquickly on addition of sulphite, while the cytochrome free ofthe reductase activity was hardly reduced by the enzyme withsulphite. A sulphite oxidase system was reconstituted with the reductase,cytochrome c-550 and cytochrome oxidase highly purified fromthe bacterium. ¹ Present address: Water Research Institute, Nagoya University,Nagoya 464, Japan ² Present address: Institute for Biological Science, SumitomoChemical Co., Ltd., Takarazuka, Hyogo 665, Japan (Received January 23, 1981; Accepted March 9, 1981)     

Phytochrome Participation in Induction of Haustoria in Cuscuta japonica, a Holoparasitic Flowering Plant

Seedlings of Cuscuta japonica pasted on an acrylic plate developedmany haustoria in response to far-red light after pre-irradiationwith white light. The effect of far-red light was cancelledcompletely by red light irradiated immediately after the far-redlight. In order to elucidate the photoreceptor(s) for photocontrolin the induction of haustoria, action spectra for the photo-inductionand its reversion were determined in the wavelength region from340 nm to 800 nm using the Okazaki Large Spectrograph. The action spectrum for the induction of haustoria had a pronouncedpeak at 740 nm and a small peak at 420 nm, while the actionspectrum for reversal of the induction had a pronounced peakat 660 nm and a small peak at 380 nm. These results indicatethat phytochrome is involved in the photocontrol of inductionof haustoria in Cuscuta japonica. Considering the far-red /redreversibility, it was suggested that phytochrome B is the photorecepter.This is the first reliable evidence of phytochome participationon development of haustoria in parasitic plants. ⁴Present address: Graduate School of Science Division of BiologicalScience, Nagoya University, Nagoya, 464-01 Japan.     

Mating reaction in Saccharomyces cerevisiae VI. Effect of 2-deoxyglucose on conjugation

The effect of 2-deoxyglucose (2-dG) on the mating reaction ofSaccharomyces cerevisiae was investigated and the followingresults were obtained. 1) The cell fusion process of the mating reaction was completelyinhibited by 0.05% 2-dG added to a culture medium containing2% D-glucose. This inhibition was partially reversed by raisingthe glucose concentration in the medium. 2) Sexual cell agglutination was hardly affected by 2-dG. 3) 2-dG at concentrations inhibiting cell fusion considerablysuppressed the incorporation of ¹⁴C-glucose into the cell wallpolysaccharides, glucan and mannan. 4) Glucose uptake and protein synthesis were only slightly inhibitedby 2-dG. 5) No enhancement of bulk polysaccharide synthesis was detectedduring mating. ¹Present address: Biological Institute, Faculty of Science,Nagoya University, Chikusa-ku, Nagoya 464, Japan. (Received April 20, 1974; )     

COMPARATIVE STUDIES ON GROWTH, RESPIRATION, PHOTOSYNTHESIS AND PIGMENT CONTENT IN RHODOPSEUDOMONAS PALUSTRIS

1. Comparative studies were performed on growth, photosyntheticand respiratory activities, and pigment content in Rhodopseudomonaspalustris.
2. The growth of the organism, as influenced by variousculturalconditions such as light, aerobiosis, anaerobiosisand nutritionalfactors was investigated.
3. The respiratoryactivity of the bacterium was found to be higherin dark-growncells than in cells grown in the light. The photosyntheticactivitydid not significantly depend on the growth conditionsof theculture. Cells of younger cultures were found to be moreactivethan those of older cultures, with respect both to respirationand photosynthesis.
4. The pigment content was found to be higherin the light-growncells than in the dark-grown ones. The ratiophotosyntheticactivity/bacteriochlorophyll was significantlyhigher in thelatter than in the former.
5. Light, as well asvarious nutritional factors, was found toexert a marked accelerationon pigment formation, although ithas not yet been possibleto culture cells completely lackingin photosynthetic pigmentsand accordingly in photosyntheticactivity.

¹ Present address: Division of Dermatology and Urology, TokyoMetropolitan Hiroo Hospital, Tokyo. ² Present address: Department of Biology, Saitama University,Urawa. ³ Present address: Department of Biochemistry, School of Medicine,Yokohama University, Yokohama. ⁴ Present address: Department of Biophysics and Biochemistry,Faculty of Science, University of Tokyo, Tokyo. (Received July 23, 1961; )     

THE INCORPORATION OF 32P ORTHOPHOSPHATE INTO THE PHOSPHOLIPIDS OF ELONGATING AVENA COLEOPTILES

Four phospholipids of Avena coleoptile tissue were identifiedas phosphatidyl inositol, phosphatidyl glycerol, phosphatidylethanolamine and phosphatidyl choline. IAA caused an increase in total uptake of ³²P and incorporationof ³²P in phospholipids. IAA also caused a shift in the proportionsof identified ³²P phospholipids. Incorporation of ³²P into phosphatidylcholine was greater while incorporation into phosphatidyl glyceroland phosphatidyl ethanolamine was less in IAA-treated tissuecompared with untreated control tissue. ¹Contribution No. 338 from the Department of Botany, PennsylvaniaState University and 3001 from the Pennsylvania AgriculturalExperiment Station. ²Present address: Juniata College, Huntingdon, Pennsylvania,U.S.A.     

Light-Dependent Expression of Protochlorophyllide Oxidoreductase Gene in the Liverwort, Marchantia paleacea var. diptera

A cDNA encoding the NADPH:protochlorophyllide oxidoreductase(EC 1.6.99.1 [EC] ) was isolated from suspension-cultured cells ofthe liverwort, Marchantia paleacea var. diptera. In contrastto the situation in most higher plants, the liverwort gene wasexpressed in a light-dependent manner. ²Present address: Department of Biological Science, Facultyof Science, Kumamoto University, Kurokami, Kumamoto, 860-8555Japan.