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)     

SOME BEHAVIOURAL AND ELECTRICAL RESPONSES OF SCROBICULARIA PLANA (BIVALVIA:TELLINACEA) TO SIPHONAL WOUNDING

Water pumping, valve movements and heart rate have been recordedfrom Scrobicularia for short periods of normal behaviour andthen after siphonal wounding. Scrobicularia exhibits regularand repetitive pumping periods interrupted for only 2–3s after siphonal wounding, without the regularity of these periodsbeing affected. Wounding does not prevent animals from usingtheir inhalant siphons for deposit feeding. A preliminary investigationof neural responses to stimulation has shown that wounding thesiphon causes minimal disturbance to the animal, a brief (2s)burst of nerve activity occurs, the siphon is retracted, butvalve adduction does not occur. In contrast to this tactilestimulation of the mantle edge always elicits a large burstof impulses in the posterior adductor nerve, valve closure results,usually for 14–15 s. ¹Present address: Dept of Zoology, University of Cape Town,Rondebosch 7700, South Africa. (Received 2 February 1981;     

Changes in Steady-State Photosynthetic 14C-Incorporation into cellular Metabolites of Chlorellax pyrenoidosa during Transitions from High to Low Irradiance

Changes in the levels of ¹⁴C-labelled metabolites were monitoredin Chlorella pyrenoidosa cells during a transition from highto low irradiance, i.e., from 700 to 430 µmol quanta (400–700nm) m^–2 s^–1. Chlorella cells assimilated ¹⁴CO₂ photosynthetically(steady-state ¹⁴C-labelling) for 12 min at the high irradianceand then 10 min at the low irradiance. With the transition tolow light, the level of ¹⁴C-labelled ribulose 1,5- bisphosphate(RuBP) did not decrease, even though the rate of total ¹⁴C-incorporationdecreased by 80%. The data suggest that RuBP carboxylase deactivatesrapidly (within 1 or 2 min) on exposure to low light, causingRuBP pool sizes to be maintained (or even increased) in spiteof a decreased rate of RuBP regeneration. There was also evidenceof light modulation of other enzymes, including some enzymesinvolved in sucrose synthesis. The rate of sucrose synthesisdecreased with decrease in light intensity while the level ofuridine diphosphoglucose increased, but within a few minutes,both returned to their former levels. ¹Present address: Chemical Biodynamics, Lawrence Berkeley Laboratory,Building 3, 1 Cyclotron Road, Berkeley, CA 94720, U.S.A. (Received March 8, 1986; Accepted June 25, 1986)     

Comparison of Photosynthetic and Photorespiratory Enzyme Activities between Green Leaves and Colorless Parts of Variegated Leaves of a C4 Plant, Stenotaphrum secundatum (Walt.) Kuntze

The activities of enzymes involved in C⁴ photosynthesis andphotorespiration in colorless parts of variegated leaves ofStenotaphrum secundatum (Walt.) Kuntze were compared with thosein green leaves. Chlorophyll content of the colorless part wasonly about 0.3–3% of that of the green leaves. The activities of chloroplastic enzymes, pyruvate, P_i dikinase,NADP⁺-malic enzyme and NADP⁺-glyceraldehyde 3-phosphate dehydrogenasewere considerably lower in colorless tissue on a fresh weightor protein basis (the ratios of the activities in the green/colorlesstissues ranging from 5 to 20). A cytoplasmic enzyme, UDP-glucosepyrophosphorylase as well as aspartate and alanine aminotransferasesshowed comparable activities in the two types of tissue, whereasPEP carboxylase in the colorless tissue had only the one-thirdactivity of that in green tissue. Differences in activitieswere also observed for the glycolate pathway enzymes (the ratiosranging from 2 to 7 for glycolate oxidase, hydroxypyruvate reductaseand serine hydroxymethyltransferase, and 7 to 15 for catalase),while cytochrome c oxidase showed comparable activity in thetwo types of tissue. The results suggest that the deficiency of thylakoid developmentin the colorless tissue influences enzyme activities not onlyin plastids but also in other cellular compartments. ¹Present address: Institute of Applied Microbiology, Universityof Tokyo, Tokyo 113, Japan. (Received March 26, 1986; Accepted June 17, 1986)     

Effect of Ethylene on the Increase in Catalase Activity through Microbody Development in Wounded Sweet Potato Root Tissue

Catalase activity increases when slices of sweet potato roottissue are incubated in air. The increase is due to de novosynthesis of the enzyme protein and probably also to activationof a precursor protein [Esaka et al. (1983) Plant & CellPhysiol. 24: 615]. The activity-increase was partly depressedwhen tissue slices were incubated in ethylenecontaining air,while the immunologically determined amount of catalase proteindid not increase, rather it decreased, under the same conditions.We propose that ethylene inhibits the de novo synthesis of catalaseprotein but not the activation of precursor protein. Catalasefrom tissue slices incubated in ethylene-containing air migratedfaster on a polyacrylamide gel than that from intact tissueor tissue slices incubated in air. When either polyacrylamideor an SDS-polyacrylamide gel applied with crude extract fromtissue slices incubated in ethylene-containing air underwentimmunological blotting, the blots were much fainter than thosefor intact tissue. In addition, microbody membrane fractionfrom incubated tissue slices contained a significant amountof catalase which was sedimented at the bottom of a sucrosedensity gradient (20–70%) and was not solubilized by highconcentrations of lubrol PX. The fraction showed an exceptionallyhigh catalase activity per unit amount of immunoreactive proteinto anti-catalase antibody. We propose that ethylene causes somemodification of catalase protein which facilitates the formationof aggregates or cores. ¹Present address: Laboratory of Food Technology, Faculty ofApplied Biological Science, Hiroshima University, Fukuyama,Hiroshima 720, Japan. ²Present address: Terumo Co. Ltd., Omiya, Fujinomiya, Shizuoka418, Japan. (Received October 16, 1982; Accepted February 24, 1983)     

The occurrence and properties of dihydrofolate reductase in pea seedlings

Dihydrofolate reductase (E.C. 1.5.1.3 [EC] ) was found in pea seedlingsand was partially purified by treatments with ammonium sulfate,protamine sulfate and by DEAE-cellulose column chromatography.Some properties of the enzyme were investigated. Optimum pHfor the reaction was 6.5. In the enzyme reaction, FAH₂ and NADPH₂were specifically required. MICHAELIS constants for FAH₂ andNADPH₂ were 4.3x10^–6 M and 4.0x10^–5 M, respectively.Folate antagonists such as aminopterin, methotrexate and pyrimethaminewere potent inhibitors of this enzyme. Enzyme activity was almostcompletely inhibited at a concentration of 10^–7 M of aminopterinand methotrexate and 10^–6 M of pyrimethamine. Growth of germinating pea seeds was inhibited by aminopterin,methotrexate and pyrimethamine, and it recovered significantlywith a tetrahydro-derivative of folate, CF, but not with dihydrofolicor folic acid. These results suggest that growth inhibitionof pea seedlings by these antagonists is due to inhibition ofdihydrofolate reductase in seedlings. ¹Studies on the enzymatic synthesis and metabolism of folatecoenzymes in plants IV. (For the previous paper, Part III, seeReference (21)) . Part of this paper was presented at the AnnualMeeting of the Agricultural Chemical Society of Japan held atTokyo on April 4, 1967 (Received October 8, 1969; )     

Dependence of Plasmalemma Conductance and Potential on Intracellular Free Ca2+ in Tonoplast-Removed Cells of a Brackish Water Characeae Lamprothamnium

In response to hypotonic treatment internodal cells of the brackishwater Characeae Lamprothamnium regulate turgor pressure by releasingK⁺ and Cl^–, accompanying membrane depolarization and atransient increase in membrane electrical conductance (Okazakiet al. 1984b). The hypothesis that a transient increase in cytoplasmicfree Ca²⁺ concentration ([Ca²⁺]_c) caused by hypotonic treatmenttriggers release of K⁺ and Cl^– from the cell (Okazakiand Tazawa 1986a, b, c) was tested using tonoplast-removed cells.These cells did not regulate turgor pressure. The plasmalemmaconductance remained almost constant for a change in the intracellularfree Ca²⁺ concentration ([Ca²⁺],) from 10^–6 to 10^–2mol?m^–3. The results suggest that some cytoplasmic Ca²⁺-sensitizingsoluble components, which work as mediators to activate K⁺ and/orCl^– channels in the plasmalemma and/or the tonoplast,were lost after desintegration of the tonoplast. The plasmalemmapotential was depolarized under high [Ca²⁺]_i. However, no membranedepolarization was observed upon hypotonic treatment. Sincemembrane depolarization has been suggsted to occur under normal[Ca²⁺]_c in intact cells (Okazaki and Tazawa 1986a, b), its absencesuggests that some cytoplasmic factors, which induce the membranedepolarization in a Ca²⁺-independent manner, are lost in tonoplast-removedcells. ¹ Present address: Department of Biology, Osaka Medical College,Sawaragi-cho 2-41, Takatsuki, Osaka 569, Japan. (Received October 22, 1986; Accepted March 31, 1987)     

REGULATION OF BUD REST IN TUBERS OF POTATO SOLANUM TUBEROSUM L.: II. INHIBITION OF SPROUTING BY INHIBITOR {beta} COMPLEX AND REVERSAL BY GIBBERELLIN A3

Two acid ether extracts of inhibitor ß complex wereprepared from resting ‘Red Pontiac’ potato tuberpeelings, and buds. The inhibiting zone was separated chromatographicallyand tested on wheat coleptiles, dwarf peas, and potato "eyes."Dwarf peas did not respond. ß showed marked inhibitionof wheat coleoptile elongation and of sprouting of potato eyes.Ability to inhibit sprouting was a function of repeated application.This inhibition was reversed by GA₃. When the inhibiting eluatesof the chromatographed first extraction were stored for 6 monthsat –10°, they completely lost ability to inhibit bothcoleoptiles and potato buds. Storage of the corresponding eluatesof the second extract at 4° for 48 days resulted in partialloss of activity in the coleoptile test and complete loss inthe potato eye test. The results tend to support the conceptthat a balance of inhibitors and stimulators participates incontrolling rest, but do not support conclusively the implicationof ß in rest. ¹This study was generously supported in part by United StatesPublic Health Service Grant EF-61 ²Present address: Department of Botany, Hebrew University, Jerusalem,Israel     

Studies on the effect of certain enzymic poisons on the metabolism of storage organs III. Uptake and utilization of sucrose by radish root slices in presence of iodoacetate

Iodoacetate greatly retarded the uptake of sucrose and slightlyaffected its inversion by radish root slices. Carbohydrate content of the samples decreased substantiallyboth in water and in iodoacetate. Feeding with sucrose led tomarked accumulation of carbohydrates and supplemental additionof iodoacetate induced less accumulation of carbohydrates. Iodoacetate caused exudation of nitrogen fractions into theculture media. Protein synthesis via amino acids seems to beoperative in iodoacetate treated slices. It is also suggestedthat nitrate-N, in presence of sucrose, is converted into peptidesand proteins. Addition of iodoacetate to sucrose media inhibitedthis pathway of protein synthesis. Both sucrose and iodoacetate (4 x 10^–4M) stimulated theCO₂ output whereas 25 x 10^–4M iodoacetate did not changethe CO₂ output when compared with that of controls. Sucroseand iodoacetate (4 x 10^–4 M) when joined together maskedthe accelerating effect of each other. ¹Present address: Department of Botany, Faculty of Science,University of A'in Shams, Abbassia, Cairo, Egypt, U. A. R. (Received November 6, 1968; )     

Quantitative Isolation of Undegraded Polysomes from Aged Pea Tissue in the Absence of Contaminants and Artefacts

Undegraded polysomes were isolated successfully from aged peaepicotyls by grinding frozen tissue in at least 10 volumes ofbuffer A (0.2 M Tris-HCl, pH 8.5; 0.2 M sucrose; 60 mM KCl;30 mM MgCl₂), taking care to prevent the tissue from thawingprior to homogenization. Supposedly pure polysomes, derivedfrom the membrane-bound polysome fraction, were apparently contaminatedwith membranes, and contained polysomes clumped together vianascent chains. Problems with contaminants and artefacts werepartially alleviated by the use of polyoxyethylene tridecylether as a detergent replacing Triton X-100; further alleviatedby the use of large volumes of detergent-containing buffer toresuspend the membrane-bound polysome; and almost completelyeliminated by brief treatment of resuspended polysomes withprotease K. Optimal conditions for isolating polysomes fromaged tissue are given. ¹ Present address: Institute of Agricultural Environment Control,College of Agriculture, Ehime University, 3-5-7 Tarumi, Matsuyama790, Japan. (Received April 24, 1985; Accepted September 2, 1985)     

Inhibiting Effect of Auxin on Glucosamine Incorporation into Cell Walls of Rice Coleoptiles

Auxin induced growth and decreased the hexosamine content ofthe cell walls of rice coleoptile sections. Indole-3-aceticacid (IAA) at 10^–5 M inhibited the incorporation of ¹⁴C-glucosamineinto the cell walls. IAA did not affect the ¹⁴C-incorporationinto the cytoplasm, while inhibitors of glycoprotein synthesis,unicamycin and monensin, suppressed the incorporation into boththe cytoplasm and the cell walls. The radioactivity due to labeledglucosamine in the cell walls increased during the chase, butthis increase was inhibited by IAA. Among the cell wall fractions,the increase in radioactivity and its inhibition by IAA wereconspicuous in the hemicellulose I fraction. The inhibitoryeffect of IAA on glucosamine incorporation into the cell wallswas observed even in the presence of 0.15 M mannitol solutionwhich completely suppressed the IAA-induced growth. These resultssuggest that auxin induces growth at least partly by inhibitingthe transport of asparagine-linked glycoproteins from the cytoplasmto the cell walls. ¹ Present address: Department of Biology, Faculty of Science,Osaka City University, Sumiyoshi-ku, Osaka 558, Japan (Received July 23, 1986; Accepted December 22, 1986)     

Different Intracellular Localization of Two Cytochrome P-450 Systems, Ipomeamarone 15-Hydroxylase and Cinnamic Acid 4-Hydroxylase, in Sweet Potato Root Tissue Infected with Ceratocystis fimbriata

Ipomeamarone 15-hydroxylase activity was mainly recovered inthe pellet fraction between centrifugations at 10,000 and 100,000?gfrom a crude extract of Ceratocystis fimbriata-infected sweetpotato root tissue, whereas cinnamic acid 4-hydroxylase activitywas found between centrifugations at 300 and 10,000?g. Whenparticles in the crude extract were fractionated by sucrosedensity gradient centrifugation, the rough-surfaced microsomeswere distributed over a wide density range from 1.09 to 1.14g cm^–3, judging from the distributions of protein, RNAand NADPH-cytochrome c reductase activity. Phosphorylcholine-glyceridetransferase activity was only in the lighter half of the microsomalfraction (density: 1.09–1.11 g cm^–3). Ipomeamarone15-hydroxylase activity was found in heavier half of the microsomalfraction (density: 1.10–1.14 g cm^–3). We proposethat this tissue has two rough-surfaced endoplasmic reticulumspecies, only one of which carries phosphorylcholine-glyceridetransferase, and that the cytochrome P-450 system is localizedon the species lacking the enzyme. Cinnamic acid 4-hydroxylaseactivity was mainly found in a fraction that had densities of1.17–1.19 g cm^–3 and contained vesicular particlesof various sizes. ¹ Present address: Laboratory of Food Hygienics, Faculty ofAgriculture, Kagawa University, Miki-cho, Kida-gun, Kagawa 761-07,Japan. (Received September 6, 1984; Accepted December 27, 1984)     

Changes in Activities of Enzymes Involved in General Phenylpropanoid Metabolism during the Induction and Reduction of Anthocyanin Synthesis in a Carrot Suspension Culture as Regulated by 2,4-D

The activities of enzymes involved in general phenylpropanoidmetabolism were followed in a carrot suspension culture duringthe induction and reduction of anthocyanin synthesis regulatedby 2,4-D. When no anthocyanin synthesis occurred in a mediumcontaining 2,4-D (+2,4-D medium), the activities of phenylalanineammonia-lyase (PAL) and 4-coumarate:CoA ligase (4CL) increased1 day after transfer due to the transfer effect, but subsequentlydecreased and remained at a low level. Cinnamate-4-hydroxylase(C4H) activity showed a low level throughout culture. When cellswere transferred to a medium lacking 2,4-D (–2,4-D medium),the activities of PAL, C4H and 4CL increased and maximum activitiesof these enzymes were observed 6–7 days after transfer,when anthocyanin was most rapidly synthesized. When cells were cultured in the –2,4-D medium, the additionof 2,4-D immediately reduced the induced activity of PAL. PALactivity was super-induced by the transfer effect, while anthocyaninsynthesis decreased. The addition of intermediates of generalphenylpropanoid metabolism, with 2,4-D, to the medium 6 daysafter transfer to the –2,4-D medium did not promote anthocyaninsynthesis, whereas dihydroquercetin did promote it. Regulationof anthocyanin synthesis by 2,4-D is discussed in relation tochanges in enzyme activities involved in general phenylpropanoidmetabolism. ¹ Present address: Cell Science and Technology Division, FermentationResearch Institute, Agency of Industrial Science and Technology,Yatabe-machi, Ibaraki 305, Japan. ² Present address: Biological Institute, Faculty of Science,Tohoku University, Sendai 980, Japan.     

Effect of Nasal Dilators on Perceived Odor Intensity

Subjects wearing nasal dilators rated olfactory stimuli as beingmore intense compared with ratings done without nasal expansion.The results support a perceptual constancy model in olfaction.Chem. Senses 22: 177–180, 1997. ¹Present address: Biology Department, St Lawrence UniversityCanton, NY 13617, USA ²Present address: PO Box 802, Drew University, Madison, NJ 07940,USA     

Effect of Salicylhydroxamic Acid on Respiration, Photosynthesis, and Peroxidase Activity in Various Plant Tissues

Earlier reports from our laboratory described salicylhydroxamicacid (SHAM) stimulation of O₂ uptake by expanded soybean leavesor older green cotyledons. This stimulation could not be interpretedin terms of engagement or capacity of the cytochrome and alternativerespiratory pathways. In this report, we tested the possibilitythat a soluble peroxidase, which can be easily eluted from soybeanleaves and cotyledons, might be responsible for SHAM stimulationin whole tissue. The peroxidase catalyzes oxidation of NADHby O₂, is strongly stimulated by SHAM and benzhydroxamic acid(BHAM) and inhibited by KCN, propyl gallate and gentisic acid.This peroxidase, however, does not seem to be responsible forSHAM-stimulated O₂ uptake in whole, green tissue. In our earlier work reporting SHAM-stimulated respiration ingreen tissue, the samples had not been shielded from room light(10–20 µmol photons m^–2.s^–1). In thisreport, we show that O²-uptake rates of controls measured indarkness were always greater than those measured in room light.SHAM stimulation was not observed in the dark or in tissue withoutchlorophyll. We also found that CO₂ uptake of whole leafletsin saturating light was completely inhibited by SHAM fed throughthe transpiration stream. SHAM, therefore, is a potent inhibitorof photosynthesis. We conclude that the SHAM-stimulated respirationof green tissues we reported earlier likely was due to verylow rates of photosynthesis occurring under room light. ³Present address: SANDOZ Ltd., Agrobiological Research Station,4108 Witterswil, Switzerland ⁴Present address: WTC 1A3, Weyerhaeuser Co., Tacoma, WA 98477,U.S.A. (Received June 23, 1989; Accepted October 20, 1989)     

Planktonic ciliates in the northwestern Indian Ocean: their abundance and biomass in waters of contrasting productivity

The abundance and biomass of planktonic ciliates in the northwesternIndian Ocean ranged from 31 l^–1 and 0.1 µg C l^–1in oligotrophic open-ocean waters to 823 l^–1 and 1.2 µgC l^–1 in more productive waters of the equator, northernArabian Sea and Gulf of Oman. ³Present address: British Antarctic Survey, High Cross, MadingleyRoad, Cambridge, CB3 OET, UK     

Cell elongation in the soybean root: The influence of inhibitors of RNA and protein biosynthesis

A possible requirement for RNA and protein synthesis duringcell elongation of intact seedling tissue was studied usingthe soybean seedling foot with the elongating zone being delineatedby India ink marks at 2 and 7 mm back of the root tip. In contrastto most excised plant tissues, there was marked net synthesisof RNA and protein during cell elongation of the intact root.AD and CH were potent inhibitors of cell elongation in the soybeanroot. CH essentially eliminated protein synthesis, whether measuredby net accumulation of protein or by ¹⁴C-leiicine incorporation,while completely inhibiting cell elongation after a short lag.AD, on the other hand, only partially inhibited protein synthesiswhile causing almost total inhibition of cell elongation aftera lag. The capacity of the tissue to synthesize protein in thepresence of AD was correlated with the maintenance of functionalpolyribosomes, thus suggestive that m-RNA associated with theregulation of cell elongation is more unstable (i.e., a shortermean life) than total root m-RNA. FU did not inhibit cell elongation,protein synthesis or the level of functional polyribosomes.The requirement for RNA synthesis during cell elongation ofthe seedling root, as in excised plant tissues, appears to berestricted to the AMPrich species of RNA presumed to be m-RNA. ¹This research was supported by NIH grant GM 10157. ²Purdue University AES paper No. 3359. ³Present address: Dept. of Botany, National Taiwan University,Taipei, Taiwan.     

Some properties of shikimate: NADP oxidoreductase produced in sweet potato root tissue after slicing

The activity of shikimate: NADP oxidoreductase [EC 1. 1. 1.25] in sweet potato root tissue increased soon after slicing.Enzyme preparations obtained from both sliced tissue and fromfresh tissue probably contained a single enzyme component, andthey showed identical chromatographical behaviour. K_m values of the enzyme for NADP and shikimate were 1.0x10^–4Mand 1.3 x 10^–3M, respectively. Enzyme activity was potentlyinhibited by SH-inhibitors such as p-chloromercuribenzoate andoxidized glutathione. Enzyme activity was affected neither by mononucleotides suchas ATP, ADP and AMP, divalent cations, Mg⁺⁺, Ca⁺⁺ and Mn⁺⁺,nor by metabolites such as tryptophan, phenylalanine, tyrosineand t-cinnamic acid which are involved in aromatic compoundsyntheses. The enzyme rapidly lost its activity. This inactivation reactionshowed a time course consisting of two steps of the first-orderreaction. The inactivated enzyme preparation was not reactivatedby thiol compounds such as cysteine, 2-mercaptoethanol and glutathione,although these reagents, to a certain extent, protected theenzyme from inactivation. The results suggest that denaturationof the enzyme protein was involved in inactivation of the enzyme. ¹Part 74 of the phytopathological chemistry of sweet potatowith black rot and injury. ²Present address: Department of Biology, Faculty of Science,Tokyo Metropolitan University, Setagaya-ku, Tokyo. (Received August 5, 1968; )