Indolepyruvate Pathway for Indole-3-Acetic Acid Biosynthesis in Bradyrhizobium elkanii

Indole-3-acetaldehyde (IAAId) was detected in the culture supernatantof Bradyrhizobium elkanii. Deuteriumlabelled L-tryptophan (Trp)was incorporated into IAAId and indole-3-acetic acid (IAA),suggesting that B. elkanii produces IAA via IAAId from Trp.In B. elkanii cell suspension, indole-3-pyruvic acid (IPyA)was converted to IAAId, and exogenously added IAAId was rapidlyconverted to IAA. Furthermore, the activity of indolepyruvatedecarboxylase (IPDC), which catalyzes the decarboxylation ofIPyA to produce IAAId and is a key enzyme for IPyA pathway,was detected in B. elkanii cell-free extract. The IPDC activitydepended on Mg²⁺ and thiamine pyrophosphate, cofactors of decarboxylation.This mounting evidence strongly suggests that IAA synthesisoccurs via IPyA pathway (Trp IPyA p IAAId IAA) in B. elkanii. (Received December 11, 1995; Accepted March 4, 1996)     

Concanavalin A Inhibits Auxin-Induced Elongation and Breakdown of (1 -> 3), (1 -> 4)-{beta}-D-Glucans in Segments of Rice Coleoptiles

Concanavalin A (Con A) suppresses auxin-induced elongation ofsurface-abraded segments from both dicotyledonous and poaceousplants. In coleoptile segments of rice (Oryza sativa L.), theauxin-induced decrease in the minimum stress-relaxation timeand increase in the mechanical extensibility of the cell wallswere also inhibited by Con A, indicating that the lectin suppresseselongation by inhibiting the cell wall loosening. Auxin causeda decrease in the level of (1 3), (1 4)-ß-D-glucansin the cell walls of rice coleoptile segments, and this decreasewas also inhibited by the lectin. Con A suppressed the autolytichydrolysis of the glucans, as well as their breakdown in vitroby a protein fraction that had been extracted from the cellwalls of rice coleoptiles with 1 M NaCl. Furthermore, most ofthe glucan-hydrolyzing activity of the wall proteins bound toa Con A-Sepharose column, suggesting that glycoprotein enzymesare involved in the hydrolysis. Although Con A also affectedthe hydrolysis of other wall polysaccharides, the present data,when considered in combination with the inhibitory effects ofglucan-specific or glucanasespecific antibodies, support theview that the breakdown of (1 3),(1 4)-ß-D-glucansis associated with the cell wall loosening that is responsiblefor auxin-induced elongation in Poaceae. (Received August 17, 1994; Accepted February 15, 1995)     

Incorporation of some possible radioactive intermediates into chlorogenic acid in sliced sweet potato tissue

t-Cinnamic acid-2-¹⁴C, p-coumaric acid-2-¹⁴C and caffeic acid-2-¹⁴Cwere administered to discs of sweet potato roots and incorporationof each radioactive compound into chlorogenic acid was compared.The data suggest that chlorogenic acid is synthesized througheither or both of two major pathways, phenylalanine t-cinnamate t-cinnamoyl derivative p-coumaroyl derivative chlorogenicacid and phenylalanine t-cinnamate p-coumarate p-coumaroylderivative chlorogenic acid. ¹Part 75 of the phytopathological chemistry of sweet potatowith black rot and injury. ²Present address : Department of Biology, Tokyo MetropolitanUniversity, Setagaya-ku, Tokyo. (Received December 23, 1968; )     

Xyloglucan and r{beta}-D-Glucan in Cell Walls of Rice Seedlings

Cell walls of 4-day old rice seedlings were extracted successivelywith ammonium oxalate-oxalic acid, 4% KOH and 24% KOH. A rß-D-glucanpreparation and a xyloglucan preparation were isolated fromthe 4% KOH extract and 24% KOH extract, respectively. Methylationanalysis and enzymic degradation studies of the polysaccharidesshowed that the former was built up predominantly of repeating-oligosaccharideunits of 3-O-rß-cellobiosyl-D-glucose and 3-O-rß-cellotriosyl-D-glucosein a molar ratio of 2.6 : 1.0, and the latter was of repeating-oligosaccharideunits of -D-xylosyl-(16)-rß-D-glucosyl-(14)-[-D-xylosyl-(16)]-rß-D-glucosyl-(14)-D-glucose,-D-xylosyl-(16)-rß-D-glucosyl-(14)-D-glucose and cellobiose. ¹ Present address: Department of Botany, Iowa State University,Ames, Iowa 50011, U.S.A. (Received August 29, 1981; Accepted January 12, 1982)     

Chlorophyll Formation in the YG-6 Mutant of Chlorella regularis: Spectral Characterization of Protochlorophyllide Phototransformation

Dark-grown YG-6 mutant cells of Chlorella regularis accumulateat least two forms of phototransformable protochlorophyllide(Pchlide) with in vivo absorption maxima at 634 nm (Pchlide634) and 650 nm (Pchlide 650). Difference spectrophotometricanalyses and the action spectra showed that Pchlide 634 is firsttransformed into the 648 nm form and then phototransformed intochlorophyllide (Chlide) 672 nm. Pchlide 650 is phototransformedinto Chlide 685 which then shifts towards short wavelength-formingChlide 667 in the subsequent dark stage (Shibata shift). Pchlide650 is regenerated at the expense of photoinactive Pchlide 632.In washed cells after the phototransformation, the Shibata shiftwas accelerated. Freezing/thawing treatment in the dark causedconversion of phototransformable Pchlide 650 into photoinactivePchlide 633, but phototransformation activity of Pchlide 634still partly remained. These results suggest that in the final step of light-dependentchlorophyll formation in the YG-6 mutant of C. regularis, twosequentially and functionally separate routes are present: (1) Pchlide 634 Pchlide 648 Chlide 672 Chlorophyll a. (2) Pchlide 650 Chlide 685 Chlide 667 Chlorophyll a. (Received June 4, 1983; Accepted November 11, 1983)     

Effects of auxin on the structure of hemicelluloses of Avene coleoptiles

Avena coleoptile hemicelluloses were fractionated into water-solublehemicelluloses I and IIB and water-insoluble hemicellulose IIA.These hemicelluloses were then subjected to glycosidic linkageanalysis by methylation technique, which revealed that hemicelluloseI was predominantly composed of arabinoxylans and ß-(l4)glucans and hemicellulose IIB was composed of arabinoxylans,ß-(l4) : (l3)-mixed linked glucans, ß-(l4)-glucansand xyloglucans. Hemicellulose IIA was mainly composed of xyloglucansand probably ß-(l4)-glucans. Methylation analysisof hemicelluloses extracted from Avena coleoptile segments treatedwith auxin in the presence of mannitol (0.15 M) indicated thatauxin apparently had no effect on the structure of arabinoxylanand caused a specific decrease in the amount of ß-(l4): (l3)-mixed linked glucan. (Received November 19, 1979; )     

Effects of illumination on absorption peak shifts in spectra of intact etiolated cotyledons of Pharbitis nil I. Existence of two kinds of shift patterns

Two patterns were found in the shifts of absorption peaks inspectra of intact etiolated Pharbitis cotyledons illuminatedat room temperature. One was a well-known pattern, P649C678C683C672,called the "high-intensity illumination pattern" in this study.The other, called the "low-intensity illumination pattern,"was P649C672. (Received June 16, 1976; )     

Characterization of the Hydrolytic Activity of Avocado Cellulase

The cellulase produced by ripening avocado fruits (Persea americanaMill cv. Fuerte) was isolated and purified using chromatofocusing(pH 7–4) and gel filtration on a Bio- Gel P-100 column.Characteristics of the cellulase were assessed by using, assubstrates, a range of polysaccharides containing various sugarresidues and varying types of linkages between the residues.Only those substrates containing (14)-ß-glucosyl linkageswere hydrolyzed by the purified enzyme. Two polysaccharidesthat were extensively hydrolyzed by the cellulase were carboxymethylcelluloseand (13),(14)-ß-D-glucans such as from Avena endospermcell walls. Characterization of the activity in the degradationof the mixed linked glucan of Avena and cellodextrins indicatedthat the enzyme has a limit recognition-hydrolytic site of four(l4)-ß linked glucose residues. It was also foundthat the enzyme could cleave only (14)-ß-linkagesthat were adjacent to other (l4)-ß-D-glucosyl linkages.Activity of the cellulase against isolated avocado fruit cellwalls indicated that the purified enzyme was incapable of appreciablysolubilizing the cellulosic components of these walls. ¹Supported in part by National Science Foundation Research GrantPCM 7818588. ²USDA-ARS, Dairy Forage Research Center, University of Wisconsin,Madison, WI 53706. ³Department of Vegetable Crops, University of California, Davis,CA 95616. (Received September 14, 1985; Accepted February 12, 1986)     

Methionine as a Dominant Precursor of Phytosiderophores in Graminaceae Plants

Nine ¹⁴C-labeled amino acids and ¹⁴C-acetic acid from root tipsof Fe-deficient Graminaceae plants (Hordeum vulgare, Oryzaesativa and Avena saliva) were surveyed to determine the precursoramino acid of phytosiderophores. The dominant precursor wasmethionine, which was incorporated into avenic acid deoxymugineicacid mugineic acid epihydroxymugineic acid and/or hydroxymugineicacid in this order. Methionine sulfoxide or methionine sulfonemay be important intermediates in going from methionine to avenicacid. (Received May 6, 1987; Accepted June 12, 1987)     

Long chain fatty acid synthesis in developing castor bean seeds II. Operation of the path from sucrose to long chain fatty acids in a subcellular particulate system

Particles spun down at 10,000 ? g from developing castor beanseeds were capable of synthesizing LFAs from sucrose, a physiologicalprecursor transferred from leaves as a photosynthetic product.Tracer experiments, in combination with inhibitor effects, intermediatedilutions and cofactor requirements, indicated the operationof the following path: sucroseUDPGG-1-PG-6-PGAPpyruvateacetyl-CoAmalonyl-CoALFA.The whole path appears to be associated with 10,000 ? g particles,because repeated washings were unsuccessful in dissociatinga partial path from the particles, depsite of disorganizingthe structure of the particles. Based on the occurrence of freehexose(s) and the utilization of UDPG similar to that of sucroseor G-1-P in this reaction, it is probable that hexose phosphateis formed from sucrose via UDPG and fructose, although the conversionof sucrose to hexose phosphates via glucose and furctose isnot ruled out. Inhibitor experiments showed that ATP is self-supportingover the whole path, the ATP formed in the glycolytic path beingconsumed in a acetyl-CoA carboxylation step. Since oxidizedpyridine nucleotides are as available as reduced ones for LFAsynthesis, they seem to shuttle between the reduction in theconversion of sucrose to acetyl-CoA and the oxidation in LFAsynthesis from acetyl-CoA. From the pattern of the LFAs synthesized,NAD⁺ is available for the synthesis of saturated LFAs (18: 0,16: 0). whereas NADP⁺ is available for that of unsaturated LFAs(18: 1, 16: 1). (Received July 23, 1973; )     

In vivo chlorophyll forms in higher plants and algae sensitive to treatment with lutein

Lamella preparations of spinach, Chlorella, Phaeodactylum, Anabatnaand Porphyra were treated with a hydrophobic reagent, lutein,and the absorption and fluorescence spectra in the red regionbefore and after treatment were compared for changes causedby the treatment. Absorption spectra of all these preparationsunderwent the same spectral change, transformation of a bandat 684 nm into a band at 666 nm. The longer the maximum wavelengthof the red peak, the greater was the fractional absorbance decreaseat 684 nm. The content of C684 (the chlorophyll form responsiblefor the 684 nm band) in the lamellae was estimated from thefractional decreases as being progressively higher in the orderof Phaeodactylum, Porphyra, Anabatna, Chlorella and spinach.The fluorescence spectra at liquid nitrogen temperature beforetreatment showed two bands. The longer wavelength band was transformedby the treatment into a shorter wavelength band(s), as describedbelow, according to the maximum wavelengths: spinach, F735F695(or F686); Chlorella, F715F700 (or F686); Phaeodactylum, anunidentified componentF690; Anabaena, F732F685 (or F695); Porphyra,F726F683. These chlorophyll forms with fluorescence maxima between715 and 735 nm were, therefore, designated C684 based on absorptionspectrophotometry, and are considered to play a role in photosystemII. (Received August 15, 1972; )     

AGMATINE AND N-CARBAMYLPUTRESCINE AS INTERMEDIATES IN THE FORMATION OF NICOTINE BY TOBACCO PLANTS

Agmatine-G-³H and N-carbamylputrescine-l,4-¹⁴C were effectivelyincorporated into the nicotine of tobacco plants. This resultmay indicate a route that the pyrrolidine ring of nicotine isformed from putrescine by the following pathway: arginineagmatineN-carbamylputrescineputrescinepyrrolidinering. (Received February 7, 1966; )     

A PHENOLIC PIGMENT REACTIVE TO L-LACTATE CYTOCHROME C REDUCTASE OF YEAST

1. A phenolic pigment was extracted from baker's yeast. The pigmentis slowly autooxidizable, and rapidly oxidized with Rhus-laccaseor polyphenol oxidase and reduced by dithionite.
2. The pigmentdissolved in ethylether had an absorption peak at258 mµ,shoulders at 289 and 382 mµ and a plateauat 450–500mµ. The difference spectrum between oxidizedand reducedforms of the pigment showed a wide plateau around500 mµ.
3. The pigment supported the oxygen uptake by reconstructed enzymesystem: L-lactate, L-lactate cytochhrome c reductase and Rhuslaccaseor polyphenol oxidase. In its absence, no oxygen uptake tookplace. The pigment was replaced successfully with p-quinone,catechol and menadione, but not with ubiquinone. The sequenceof hydrogen transport can be represented: L-lactate L-lactatecytochrome c reductase "phenolic pigment" oxidase oxygen.

(Received August 11, 1967; )     

Rhizoid differentiation in Spirogyra I. Basic features of rhizoid formation

Several types of rhizoids occurring in the process of differentiationin Spirogyra sp. are described and their interrelation was elucidated.There are two differentiation sequences: P_pPRh_ros or PpPRh_rodRh_ros(for explanation of abbreviations see p. 533), although undersome conditions the sequences ceased halfway through. The initiationtime for rhizoid formation had no relation to the cell cyclestage. The difference in growth patterns between the rhizoidand ordinary filament cells was demonstrated with Calcofluor-stainingand centrifugation. The optimum temperature and pH of the culture medium for rhizoiddifferentiation were 20?C and pH 7, respectively. A contactstimulus was not necessary for induction. Of the several environmental factors examined, light was themost important, for rhizoid formation, since a rhizoid was inducedonly when light was given after cutting the filament. (Received December 14, 1972; )     

Mycobacterial arabinan biosynthesis: the use of synthetic arabinoside acceptors in the development of an arabinosyl transfer assay

Information on the biosynthesis of the D-arabinans of the cellwall of Mycobacterium tuberculosis is rapidly emerging, withthe promise of new targets for drug development against tuberculosis.Accordingly, arabinosyl transferase assays were developed utilizingsynthesized [1–¹⁴C]-β-D-arabinofuranosyl-1-monophosphoryldecaprenolas donor and a variety of O- and S-alkyl arabinosides as acceptors.These were: -D-Araf-(15)--D-Araf-O- and -S-alkyl di-arabinosidesand -D-Araf-(15)--D-Araf-(15)--D-Araf-O- and -S-alkyl triarabinosides.Whereas the O- and S-alkyl monosaccharide acceptors were inactive,the O- and S-alkyl disaccharide and the O- and S-alkyl trisaccharideacceptors (<C12) possessed considerable acceptor activity,and the trisaccharide acceptors were more potent than the correspondingdisaccharides. The O-alkyl disaccharide acceptors with a C8alkyl chain were more active than those containing the C6 orC10 analogs. Chemical analysis of the enzymatically synthesizedproducts of the reactions demonstrated that β-D-arabinofuranosyl-1-monophosphoryldecaprenolwas an effective donor for two of the three potential arabinosyltransferases: β-D-arabinofuranosyl-1-monophosphoryldecaprenol:arabinan (15) arabinosyl transferase and β-D-arabinofuranosyl-1-monophosphoryl-decaprenol:arabinan β(12) arabinosyl transferase. The β(12) arabinosyltransferase activity was more in evidence in the presence ofthe O-alkyl disaccharide acceptor, whereas both transferaseswere about equivalent in the presence of the S-alkyl trisaccharideacceptor. The tuberculosis drug, ethambutol, a known mycobacterialarabinosyl transferase inhibitor, was inactive within thesearabinosyl transferase/acceptor based assay systems, supportingother evidence that a third activity, responsible for the formationof 13 linkage, is the drug target. acceptor arabinan biosynthesis glycosyltrans-ferase assay mycobacteria     

Carbohydrate Metabolism in the Developing Endosperm of Rice Grains

The metabolism of carbohydrates in developing rice endospermwas characterized by a comparison of levels of activities of33 major enzymes between the endosperm and green leaves of rice.Activities of ADPglucose pyrophosphorylase, starch synthaseand branching enzyme (Q-en-zyme), compared on the basis of solubleprotein content, were markedly higher in endosperm than in greenleaves. The high levels of Q-enzyme may be responsible for theefficient production of starch in the rice endosperm. The measurementof levels of metabolic intermediates and the localization ofkey enzymes in isolated amyloplasts from rice endosperm supportthe view that sucrose is metabolized in the cytoplasm via thepathway: sucroseUDPglucosehexose-PFBPtriose-P. Triose-P thenenters the amyloplast, where it is converted to G1P via FBPand, finally, G1P is converted to starch by the concerted reactionsof ADPglucose pyrophosphorylase, starch synthase and Q-enzyme. ¹Present address: Yamagata Prefectural Agricultural Experi mentStation, Minorigaoka, Yamagata, 990-02 Japan. ²Present address: Institute of Biological Sciences, The University of Tsukuba, Tsukuba Science City, Ibaraki, 305 Japan. (Received February 15, 1989; Accepted June 10, 1989)     

Hydrogen Peroxide is Scavenged by Ascorbate-specific Peroxidase in Spinach Chloroplasts

Intact spinach chloroplasts scavenge hydrogen peroxide witha peroxidase that uses a photoreductant as the electron donor,but the activity of ruptured chloroplasts is very low [Nakanoand Asada (1980) Plant & Cell Physiol. 21 : 1295]. Rupturedspinach chloroplasts recovered their ability to photoreducehydrogen peroxide with the concomitant evolution of oxygen afterthe addition of glutathione and dehydroascorbate (DHA). In rupturedchloroplasts, DHA was photoreduced to ascorbate and oxygen wasevolved in the process in the presence of glutathione. DHA reductase(EC 1.8.5.1 [EC] ) and a peroxidase whose electron donor is specificto L-ascorbate are localized in chloroplast stroma. These observationsconfirm that the electron donor for the scavenging of hydrogenperoxide in chloroplasts is L-ascorbate and that the L-ascorbateis regenerated from DHA by the system: photosystem IferredoxinNADPglutathione.A preliminary characterization of the chloroplast peroxidaseis given. (Received April 16, 1981; Accepted June 3, 1981)     

Isolation of Subunits of Coupling Factor 1 from Maize and Spinach Chloroplasts and Properties of Combinations of Subunits with ATPase Activity

Subunits (, ß, ) and mixtures of subunits ( ß, , ß , ß ) were isolated without denaturationfrom a chloroform extract of chloroplast coupling factor 1 (CF₁)from maize (Zea mays var. Ushiku 5-4) and from spinach by fastprotein liquid chromatography (FPLC), on an anion-exchange columnof Mono-Q in the presence of n-octylglucoside (OG) and on achromatofocusing column of Mono-P. The ß -subunitcomplex (CF¹ ß ) was the minimum unit required forATPase activity, as was confirmed by the reconstituted complexof ß and subunits. An subunit isolated from maizeinhibited the ATPase activity of CF₁ ß from bothmaize and spinach. CF₁ ß was found to contain anOG-dependent Mg²⁺-ATPase. The ATPase activity of CF₁ ß required divalent cations, such as Mg²⁺ or Mn²⁺, for its expressionin the presence of OG; its optimum pH was 8.0 and it was markedlyinhibited by NaN₃. The enzyme hydrolyzed ATP in prefernece toGTP but not CTP, UTP, ADP, AMP or pNPP. Lineweaver-Burk plotsof its activity were curvilinear in the range of 0.6–0.7mM ATP.Mg²⁺. ¹Present address: Department of Biology, School of Education,Waseda University, Shinjuku-ku, Tokyo, 160 Japan. (Received February 15, 1989; Accepted April 20, 1989)     

Metabolism of Choline Chloride and Its Analogs in Wheat Seedlings

The incorporation rate of choline chloride and allylcholinebromide into wheat protoplasts were rapid compared with theincorporation rate of benzylcholine bromide. Choline chloridewas metabolized via two pathways: choline betaine and choline phosphorylcholine phos-phatidylcholine. Allylcholine bromidewas metabolized via only one pathway: allylcholine phosphorylallylcholine phosphatidylallylcholine, and benzylcholine bromide was notmetabolized at all. These results suggest that the stimulationof photosynthesis (Hyeon et al. 1988) by these compounds iscaused directly by these choline analogs and not by their metabolites. (Received June 29, 1989; Accepted October 20, 1989)     

Function of the {alpha} Subunit of Rice Heterotrimeric G Protein in Brassinosteroid Signaling

The subunit of plant heterotrimeric G proteins (G) plays pivotalroles in multiple aspects of development and responses to planthormones. Recently, several lines of evidence have shown thatG participates in brassinosteroid (BR) responses in Arabidopsisand rice plants. In this study, we conducted a comprehensiveanalysis of the roles of the rice G in the responses to BR usinga defective mutant of the G gene, T65d1. Decreased sensitivityto 24-epi-brassinolide (24-epiBL) in the T65d1 mutant was observedin many processes examined, e.g. in the inhibition of root growthand the promotion of coleoptile elongation. The T65d1 mutantalso showed similar phenotypes to those of BR-deficient mutants,such as the specifically shortened second internode and theconstitutive photomorphogenic growth phenotype under dark conditions.However, a negative feedback effect by 24-epiBL on the expressionof BR biosynthetic genes was observed in the T65d1 mutant, andthe levels of BR intermediates did not fluctuate in this mutant.To determine the epistatic relationship between the T65d1 mutantand d61-7, a weak allele of a rice BR receptor mutant, the twomutants were crossed. The T65d1/d61-7 double mutant showed noepistasis in the elongation inhibition of the internodes, theinternode elongation pattern, the leaf angle and the morphologicalabnormality of leaf, except for the vertical length of seedand the seed weight. Our results suggest that the rice G affectsthe BR signaling cascade but the G may not be a signaling moleculein BRI1-meditated perception/transduction.