Inhibition of Adventitious Root Growth in Tradescantia by Calmodulin Antagonists and Calcium Inhibitors

When cuttings of Tradescantia fluminensis stem were incubatedin distilled water, the buds located at the node grew into adventitiousroots. The root growth could be inhibited by calmodulin antagonists,trifluoperazine, chlorpromazine, compound 48/80 and calmidazolium,in a concentration-dependent manner. The divalent cation chelatorethyleneglycol-bis-(ß-aminoethyl ether)-N, N, N, N-tetraaceticacid had no effect, however, the intracellular chelator TMB-8completely inhibited root growth. The growth was also inhibitedby calcium ionophore A23187 [GenBank] , lanthanum, a competitive inhibitorof Ca²⁺ uptake and verapamil, a calcium channel inhibitor. AWestern blot of the adventitious root extract followed by immunostainingwith an anti-spinach calmodulin antibody clearly showed thepresence of calmodulin in this tissue. These results stronglysuggest the involvement of calmodulin and calcium in the growthof Tradescantia advenitious roots. ¹A part of this work has been published in abstract form in"Molecular and Cellular Aspects of Calcium in Plant Development"(Editedby A.J.Trewavas, Plenum Publishing Co. 1986. ³Present address: Plant Laboratory , Kirin Brewerry Co. Ltd.,Kitsuregawa-cho, Tochigi-ken 329-14 ,Japan (Received May 2, 1987; Accepted September 17, 1987)     

Glutamine synthesis in germinating seeds of Cucurbita moschata

During germination, an increase in glutamine and glutamine synthetase[L-glutamate: ammonia ligase (ADP), EC 6.3.1.2 [EC] ] occurred inthe cotyledons reaching a maximum at 4 to 6 days. The enzymehad a Km of 4.5 nun for L-glutamate, and 0.67 mu for hydroxylamine.Hydroxylamine exhibited substrate inhibition kinetics. The enzymewas inhibited by calcium ion, fluoride ion and p-hydroxymercuribenzoatebut not by EDTA, sodium ion or chloride ion. The sulf hydrylinhibition was reversed by dithiothreitol. In vivo synthesisof glutamine-¹⁴C from glutamate-¹⁴C was found to parallel theincrease in glutamine content and the in vitro glutamine synthetaseactivity during germination. ¹ Present address: Department of Biology, Mercyhurst College,Erie, Pennsylvania 16501, U.S.A. (Received June 12, 1971; )     

STUDIES ON THE PRODUCTION AND PHYSIOLOGICAL EFFECTS OF ASYMMETRIN

In shake-flask culture asymmetrin was produced by PenicIlliumthomiiaud Byssochlamys niveaduring autolysis. A bioassay wasdevised to estimate relative concentrations of this compound.Culture filtrates of P. thomiicontaining asymmetrin caused adecrease in wet weight, dry weight, and total nitrogen of Phaseolusvulgarisup to 18 days after treatment. At certain intervalsafter treatment with culture filtrate of B. nivea,respirationwas inhibited, and changes were observed in isotope ratio valuesof plants supplied with glucose-¹⁴ C. Plants treated previouslywith culture filtrates of B. nivearesponded but little to gibberelhcacid. High concentrations of mdoleacetic acid inhibited growthof control plants but stimulated growth of plants treated withculture filtrate of B. nivea. ¹Present address: United Fruit Company, Norwood, Massachusetts.Supported in part by a Public Health Service Fellowship No.GF 13,776 from the division of General Medical Sciences, PublicHealth Service, National Institutes of Health, and Grant G 20989from the National Science Foundation. ²Journal Paper No. 2170 of the Purdue Agricultural ExperimentStation.     

Involvement of Calcium in Adventitious Bud Initiation in Torenia Stem Segments

In Torenia stem segments cultured in vitro, active meristematicdivisions are induced in the epidermis by treatment with cytokinin,resulting in the formation of adventitious buds. Applicationof the calcium ionophore A23187 [GenBank] was found to induce meristematicdivisions in the absence of cytokinin. The induction by A23187 [GenBank] was inhibited by simultaneous addition of auxin, but not byanti-cytokinin. A two hour pre-treatment with A23187 [GenBank] was alsoeffective, but only when it was applied to the explants justafter their excision from mother plants. The A23187 [GenBank] -inducedmeristematic zones developed into dome-shaped structures, butnot into complete adventitious buds. Complete elimination ofcalcium from the culture medium caused 50% inhibition of A23187 [GenBank] -and/or cytokinin-induced initiation of meristematic divisions.When the explants were preincubated with EGTA and then culturedon a Ca-free medium containing EGTA, cytokinin failed to inducebud initiation. Similar inhibition was also obtained by lanthanum,a calcium antagonist, by verapamil, a calcium channel inhibitor,and by trifluoperazine and chlorpromazine, calmodulin inhibitors.These results support the idea that adventitious bud initiationinduced by cytokinin in Torenia stem segments may be mediated,at least partially, by an increase in the level of intracellularCa²⁺. ¹Bioscience Research Center, Mitsui Petrochemical IndustriesLtd., Waki-cho, Kuga-gun, Yamaguchi 740, Japan. (Received May 9, 1985; Accepted October 5, 1985)     

The nature of the dual effect of auxin on root formation in Azukia cuttings

In disbudded Azukia stem cuttings, auxin exerted a dual effecton root formation. The first phase of auxin action is identifiedwith the acceleration of cell division, especially longitudinaldivision. In cuttings treated with auxin during the first 24hr, longitudinally divided cells were observed in all 12 rootprimordia, while in water-treated cuttings, such cells wereobserved only in 8 root primordia. The second phase is the promotionof the reaction in which root primordia unable to develop furtherwithout auxin supply develop into roots. Irrespective of thetreatment during the first 24 hr, the auxin-treatment duringthe second 24 hr increased the number of roots protruding fromthe cuttings. Portulal applied during the first 24 hr increased the numberof root primordia which contained longitudinally divided cells.Gibberellin applied during the first 24 hr inhibited both transverseand longitudinal divisions in root primordia. ¹ Supported in part by Grant No. 139011 from the Ministry ofEducation, Japan. ² Present address: Junior College of Toyo University, Hakusan,Bunkyo-ku, Tokyo 112, Japan. (Received June 13, 1978; )     

Trifluralin and root growth

Trifluralin inhibited root elongation and induced root tip swellingof Zea mays and Triticum aeslivum. Time-course experiments showedthese effects occurred within 6 hr of treatment. As the rootstreated with trifluralin enlarged, there was a concomitant increasein root growth inhibition. Bioassays were devised to quantitativelymeasure the radial enlargement of trifluralin-treated roots.Histological observations indicated that swollen root tissuewas growing in a non-polar manner. The root swelling effectof trifluralin was inhibited 70% by the reducing agent, 2,3-dimercaptopropanol.Trifluralin had no effect on SH content of root tip proteinof Z.mays. ¹ Published with the approval of the Director of die West VirginiaAgricultural Experiment Station as Scientific paper No. 1177. ² Present address: Department of Biology, Mercyhurst College,Erie, Pennsylvania 16501, U.S.A. (Received March 19, 1971; )     

Studies on the change of the respiratory system in roots in relation to the growth of plants II. Activity of iron-containing oxidases in roots of cereal crops with the aging of plants

Distribution of iron-containing oxidases in aging nodal rootsof rice and wheat was studied. Activities of cytochrome c oxidase(1.9.3.1 [EC] , cytochrome c : O₂ oxidoreductase), catalase (1.11.1.6 [EC] ,H₂O₂: H₂O₂ oxidoreductase) and peroxidase (1.11.1.7 [EC] , donor:H₂O₂ oxidoreductase) in wheat roots were comparatively higherthan were those in rice roots at corresponding stages. Cytochromec oxidase in roots remained active throughout the lives of therice and wheat crops. In rice roots, catalase seemed to playa distinct role around the panicle formation stage. Decay ofcatalase activity took place earlier than did that of peroxidaseand cytochrome c oxidase activities. In wheat roots similarenzyme activity changes were not observed. Data may suggestthat the high activity of iron containing oxidases at the panicleformation stage (I) may be chiefly due to catalase activityin rice roots. ¹Paper presented at the 14th Annual Meeting of the Society ofthe Science of Soil and Manure, Japan (1968). (Received November 21, 1968; )     

EVIDENCE ON THE SITE OF ACTION OF GROWTH RETARDANTS

1. The ability of five growth retardants to inhibit the GA-inducedand endogenous growth of Avena leaf sections has been investigated.The retardants vary in effectiveness. The order, from most effectiveto least, is Phosfon D, Amo-1618, C011, CCC and B995. 2. The inhibition of growth caused by Phosfon D and Amo-1618is not reversed by GA. It is apparent that the retardants donot compete with GA at the site of GA-action. 3. Addition of IAA will partially reverse the inhibition inducedby Phosfon D or Amo-1618. It is concluded that the retardantsact in part in Avena leaf sections by interfering with the auxinmetabolism of the tisssue. ¹ Supported in part by grants G-14578 and GB-1950 from the NationalScience Foundation ² Present address: Department of Botany, University of Washington,Seattle, Washington     

Elongation and Gravireactivity of Roots from an Agravitropic Maize Mutant: Implications of Growth Inhibitors

Growth of agravitropic (AGT) mutant (c.v. Kys) maize roots wassignificantly higher in comparison to that of two normal gravitropiccultivar roots. Vertical halfdecapitated roots bent towardsthe remaining half tip. This result can be explained by thefact that cap cells produced growth inhibitors (GI) moving inbasipetal direction. The curvatures of half-decapitated rootsin Kys AGT root were significantly smaller than in normal cultivarroot. One essential consequence of this AGT mutation was broughtby low GI content inside the extension zone. ¹ In honour of Professor J. Ashida (Received August 27, 1982; Accepted January 17, 1983)     

The Application of 31P Nuclear Magnetic Resonance to Higher Plant Tissue: II. DETECTION OF INTRACELLULAR CHANGES

^3IP nuclear magnetic resonance (NMR) spectra are used to monitorintracellular changes in sections of potato tuber (Solariumtuberosum), maize root tips (Zea mays) and the roots of singlemaize seedlings. Intracellular changes that result from theuptake of inorganic phosphate, D-mannose, the spectroscopicbroadening probe Mn²+ and 2, 4-dinitrophenol are described.It is concluded that NMR provides a powerful, direct methodfor following intracellular changes in plant tissues.     

Effects of Sodium on Photosynthesis in Panicum coloratum

Foliar application of NaCl to sodium-deficient Panicum coloratumstimulated photosynthesis, as did application via roots. Effectsof sodium on photosynthetic responses to internal concentrationsof CO₂ under different light intensities and initial productsof ¹⁴CO₂ fixation suggested that CO₂ fixation and aminationof oxalacetate were limited by sodium deficiency. ² Present address: Institute for Life Science Research, NihonNohyaku Co., Ltd., Kawachi-Nagano, Osaka, 586 Japan.     

Intracellular Ca2+ Concentration and Cytoplasmic Streaming in Vallisneria Mesophyll Cells

Intracellular Ca²⁺ concentration regulating the cytoplasmicstreaming in Vallisneria mesophyll cells was estimated. Theleaf segment was cut open at the middle of the mesophyll celllayers and the exposed mesophyll cells were treated with testsolutions of various Ca²⁺ concentrations in the dark. This allowedA23187 [GenBank] , a calcium ionophore, to exert its full effect on thecell membrane. The streaming was induced or maintained in solutions which containedCa²⁺ at lower than 10^–6M. However, Ca²⁺ at concentrationshigher than 10^–5M had a definite, inhibitory effect. Theinduction and cessation of streaming could be repeated by alternatelychanging the solutions. (Received March 14, 1986; Accepted May 15, 1986)     

Solubilization of nitrate reductase from the blue-green alga Anabaena cylindrica

Nitrate reductase was solubilized and purified from Anabaenacylindrica by Triton X-100 treatment of particulate preparationsfollowed by adsorption on calcium phosphate gel. Reduced methylviologen, FAD or FMN, but not ferredoxin, served as an effectiveelectron donor for the nitrate reduction by solubilized nitratereductase. ¹This work was supported by a grant (4061) from the Ministryof Education (Received June 25, 1970; )     

A cross-cultural study of taste discrimination with Australians and Japanese

As part of a larger cross-cultural study of the chemoreceptiveabilities and preferences of Australians and Japanese, thisstudy investigated the differential taste sensitivity of subjectsfrom both cultures. The results indicate that there are no significantdifferences between the abilities of subjects from the two culturesto discriminate small increments (just noticeable differences)in the intensities of the four common tastants: sucrose, sodiumchloride, citric acid and caffeine. Furthermore, the Weber ratiosfound with each tastant were within the range of those reportedin the literature. These findings represent further evidencefor the view that differences in responses of subjects fromdifferent cultures to chemosensory stimuli are restricted topreference behaviour arising from experience rather than fromgenetically-based influences. ³Present address: Faculty of Science and Technology, Universityof Western Sydney, Bourke Street, Richmond, NSW 2753, Australia ⁴Present address: CSIRO Biometrics Unit, Institute of AnimalProduction and Processing, PO BOx 52 North Ryde NSW 2113, Australia     

Effect of a Growth Inhibitor on the Hydroxyproline Level in Cell Wall of Zea Primary Roots

Evidence supporting the view that there is an inverse relationshipbetween the hydroxyproline-protein level in the cell wall andthe ability of a cell to undergo rapid cell elongation was obtained.A growth inhibitor extracted from Zea primary roots acceleratedthe incorporation of radioactivity derived from ¹⁴C-prolineinto the sodium lauryl sulfate (SLS)-insoluble cell wall fraction.However, the inhibitor had no effect on the ratio of hydroxyprolineto proline that was incorporated into the SLS-insoluble fraction. We have discussed what this growth inhibitor may mean in thegeotropic curvature of Zea primary roots. ¹ Present address: Faculty of Education, University of Yamagata,Yamagata 990, Japan (Received May 9, 1981; Accepted August 8, 1981)     

ISOLATION AND IDENTIFICATION OF ASYMMETRIN

Asymmetrin was isolated as the sodium salt from culture filtrateof Penicillium thomii. By comparison with hadacidin (N-formylhydroxyamino-acetic acid) we concluded that the two compoundsare identical. ¹Journal Paper No. 2329 of the Purdue Agricultural ExperimentStation. Supported in part by Grant G-20989 from the NationalScience Foundation to R. W. CURTIS. ²Present address: United Fruit Company, Norwood, Massachusetts.     

Na+/H+ Antiporter in Tonoplast Vesicles from Rice Roots

The Na⁺/H ⁺ antiporter in vacuolar membranes transports Na⁺from the cytoplasm to vacuoles using a pH gradient generatedby proton pumps; it is considered to be related to salinitytolerance. Rice (Oryza sativa L.) is a salt-sensitive crop whosevacuolar antiporter is unknown. The vacuolar pH of rice roots,determined by ³¹P-nuclear magnetic resonance (NMR), increasedfrom 5.34 to 5.58 in response to 0.1 M NaCl treatment. Transportof protons into the tonoplast vesicles from rice roots was fluorometricallymeasured. Efflux of protons was accelerated by the additionof Na⁺. Furthermore, the influx of ²²Na⁺ into the tonoplastvesicles was accelerated by a pH gradient generated by proton-translocatingadenosine 5'-triphosphatase (H⁺-ATPase) and proton-translocatinginorganic pyro-phosphatase (H⁺-PPase). We concluded that thisNa⁺/H⁺antiporter functioned as a Na⁺ transporter in the vacuolarmembranes. The antiporter had a K_m of 10 mM for Na⁺ and wascompetitively inhibited by amiloride and its analogues. TheKⁱ values for 5-(N-methyl-N-isobutyl)-amiloride (MIA), 5-(N-ethyl-N-isopropyI)-amiloride(EIPA), and 5-(N, N-hexamethylene)-amiloride (HMA) were 2.2,5.9, and 2.9 µ M, respectively. Unlike barley, a salt-tolerantcrop, NaCl treatment did not activate the antiporter in riceroots. The amount of antiporter in the vacuolar membranes maybe one of the most important factors determining salt tolerance. ¹This work was supported by a grant from Bio-Media Project ofthe Japanese Ministry of Agriculture, Forestry and Fisheries(BMP96-III-1).     

Effects of External Ca2+ Concentration and of the lonophore A23187 on Development of Oogonia, Oospores and Gemmae in Saprolegnia diclina

In the oomycete Saprolegnia diclina, the number of oogonia formedper unit area of culture decreased as the Ca²⁺ concentrationin the culture medium was increased. The calcium ionophore A23187 [GenBank] decreased the number of oogonia formed and increased the numberof gemmae. In the presence of 0·08 µM Ca²⁺, oogoniumand oospore abortion rates were increased by A23187 [GenBank] with a peakeffect at 0·2 nM. In the presence of 10 µM Ca²⁺,abortion rates were unaffected by A23187 [GenBank] . Saprolegnia diclina, calcium, A23187, oogonia, oospores, gemmae, development, abortion