Metabolic Alterations Associated With Abscisic Acid-Induced Frost Hardiness in Bromegrass Suspension Culture Cells

Metabolic alterations associated with the induction of freezingtolerance by abscisic acid (ABA) were characterized by chemicalanalysis and by [U-^l4C]sucrose partitioning into cellular constituentsin bromegrass (Bromus inermis Leyss cv. Manchar) cell suspensioncultures. ABA caused a significant elevation in dry matter,particularly in the fraction insoluble in 85% ethanol, thatwas highly correlated with enhanced frost tolerance. Cell walls,the largest component of the insoluble fraction, increased significantlyas frost tolerance increased throughout the ABA treatment period.ABA stimulated total [¹⁴C]sucrose uptake by cells from 7% onday 1 to 97% on day 7 compared to control cells. Partitioningstudies detected a significant increase in ¹⁴CO₂ evolution at3, 5 and 7 days after ABA treatment and a significantly higherincorporation of [¹⁴C]sucrose into the ethanol insoluble fractionafter 5 and 7 days of treatment. Organic acid depletion in ABA-treatedcells was also highly correlated with the increase in hardiness.The concentration of total sugars was higher in ABA-treatedcells. The results indicate that most of the metabolic changesduring ABA-induced acclimation were similar to changes reportedfor cells acclimated in response to low temperature. ¹Oregon Agricultural Experiment Station Technical Paper No.9052 ²Present address: Department of Horticulture, University ofSaskatchewan, Saskatoon, Sask. Canada S7N 0W0 (Received November 1, 1989; Accepted March 13, 1990)     

Protein Synthesis Linked to Respiration and Phosphorylation in Intact Euglena Mitochondria

Highly purified condensed mitochondria obtained from bleachedmutant. W₁₀BSmL of Euglena gracilis Klebs var bacillaris Coriincorporate [³⁵S]methionine into protein when fortified withmalate, ADP, Mg²⁺, phosphate and a sucrose osmoticum. Twentyto twenty-five polypeptide bands were found to be labeled inorganello when the labeled protein was subjected to sodium dodecylsulfatepolyacrylamide gel electrophoresis. Methionine incorporation,but not respiration or oxidative phosphorylation, was blockedby chloramphenicol and other 70S ribosomal translation inhibitorsbut cycloheximide and ribonuclease were without effect. Inhibitorsof electron transport and uncouplers of oxidative phosphorylationwere excellent inhibitors of protein synthesis. Thus, thesemitochondrial preparations carry out protein synthesis in organellothat is linked to respiration and oxidative phosphorylation. ¹Present address: VA Hospital Outpatient Clinic, 17 Court St.,Boston, MA 02115, U.S.A. ²Present address: Laboratories de Microbiologia e Inmunologia,Universidad Catolica de Chile, Casilla 114-D, Santiago, Chile. ³Present address: Botany Department, University of Massachusetts,Amherst, MA 01003, U.S.A. (Received June 17, 1985; Accepted October 28, 1985)     

A Mutant of Arabidopsis thaliana That Defines a New Locus for Glycine Decarboxylation

A novel photorespiratory mutant of Arabidopsis thaliana, designatedgld2, was isolated based on a growth requirement for abnormallyhigh levels of atmospheric CO₂. Photosynthetic CO₂ fixationwas inhibited in the mutant following illumination in air butnot in atmosphere containing 2% O₂. Photosynthetic assimilationof ¹⁴CO₂ in an atmosphere containing 50% O₂ resulted in accumulationof 48% of the soluble label in glycine in the mutant comparedto 9% in the wild type. The rate of glycine decarboxylationby isolated mitochondria from the mutant was reduced to 6% ofthe wild type rate. In genetic crosses, the mutant complementedtwo previously described photorespiratory mutants of A. thalianathat accumulate glycine during photosynthesis in air due todefects in glycine decarboxylase (glyD, now designated gld1)and serine transhydroxymethylase (stm). Because glycine decarboxylaseis a complex of four enzymes, these results are consistent witha mutation in a glycine decarboxylase subunit other than thataffected in the gld1 mutant. The two gld loci were mapped tochromosomes 2 and 5, respectively. ³Present address: Department of Crop and Soil Sciences, MichiganState University, East Lansing, MI 48824, U.S.A. ⁴Present address: Department of Applied Bioscience, Facultyof Agriculture, Hokkaido University, Kita-Ku, Sapporo, 060 Japan ⁵Present address: Department of Biology, Carnegie Institutionof Washington, 290 Panama Street, Standford, CA 94305, U.S.A.     

Changes in Levels of Phosphoenolpyruvate Carboxylase with Induction of Crassulacean Acid Metabolism in Mesembryanthemum crystallinum L.

Expanded leaves of Mesembryanthemum crystallinum L. performingC₃ photosynthesis were induced to perform pronounced Crassulaceanacid metabolism (CAM) by exposing the plant roots to higherNaCl concentration. Levels of phosphoenolpyruvate (PEP) carboxylaseactivity increased 10-fold during the 7-day induction period.Densitometric analysis of Coomassie-stained sodium dodecyl sulfate(SDS) polyacrylamide gradient slab gels of leaf extracts, preparedduring the course of CAM induction, revealed that at least fivebands of polypeptides increased in content (kilodalton valuesof 98, 91, 45, 41, 38). Higher levels of three additional polypeptides(kilodalton values of 102, 76, 33) became apparent after tissuehad been grown for 2 weeks at 400 mM NaCl. Of these polypeptides,that having a mass of 98 kilodaltons was identified as the subunitof PEP carboxylase by comparison with the corresponding bandfrom partially purified PEP carboxylase from the same tissue.Only a faint 98 kilodalton band was evident on SDS gels fortissue operating in the C₃ mode; staining intensity at thislocation increased with increasing NaCl-salinity in the rootingmedium until CAM was fully induced. These data provide evidencefor net synthesis of PEP carboxylase and several other proteinsduring the induction of CAM in M. crystallinum. ¹ Present address: USDA, P. O. Box 867 Airport Rd., Beckley,WV. 25801, U.S.A. ² Present address: Department of Botany, Washington State University,Pullman, Washington 99164, U.S.A. ³ Present address: Botanisches Institut der Universit?t, MittlererDallenbergweg 64, 8700 W?rzburg, W.-Germany. (Received October 27, 1981; Accepted March 15, 1982)     

Comparison of carbon dioxide fixation and the fine structure in various assimilatory tissues of Amaranthus retroflexus L

The pattern for primary products of CO₂-fixation and the chloroplaststructure of Amaranthus retrqflexus L., a species which incorporatescarbon dioxide into C₄ dicarboxylic acids as the primary productof photosynthesis, were compared in various chlorophyll containingtissues,i.e., foliage leaves, stems, cotyledons and pale-greencallus induced from stem pith. Despite some morphological differencesin these assimilatory tissues, malate and aspartate were identifiedas the major compounds labelled during a 10 sec fixation of¹⁴CO₂ in all tissues. Whereas, aspartate was the major componentin C₄-dicarboxylic acids formed in foliage leaves, malate predominatedas the primary product in stems, cotyledons and the pale-greencallus. The percentage of ¹⁴C-radioactivity incorporated intoPGA and sugar-P esters increased and ¹⁴C-sucrose was detectedin the prolonged fixation of ¹⁴CO₂ in the light, not only infoliage leaves, but also in stems and cotyledons. ¹ This work was supported by a Grant for Scientific ResearchNo. 58813, from the Ministry of Education, Japan. ² Present address: Institute of Applied Microbiology, Universityof Tokyo, Tokyo, Japan. ³ Present address: Department of Biochemistry, University ofGeorgia, Athens 30601. Georgia, U. S. A. (Received July 10, 1971; )     

THE RATIO OF CALCAREOUS AND ORGANIC SHELL COMPONENTS OF FRESHWATER SPHAERIID CLAMS IN RELATION TO WATER HARDNESS AND TROPHIC CONDITIONS

Shells from 14 populations of sphaeriid clams including Sphaeriumstriatinum, S. simile, Pisidium walkeri, Musculim partumeiumand M. iransversum were analyzed for organic carbon (µgCmg^–1 shell), nitrogen (µg,N mg^–1 shell) andCaCO_s (%CaCO₃ of total clam dry weight). Habitat waters wereassessed for total hardness (expressed as ppm CaCo₃), ppm Ca,ppm Mg, conductivity (µmho) and suspended organic Carbon(µgCl^–1). For all populations, shell organic C andN are positively correlated and there is an inverse relationshipbetween the amounts of shell CaCO₃ and shell organic carbon.Trophic considerations give the best correlation with shelltype at the generic level of consideration since species ofMusculium are found at the opposite end of the trophic scale(eutrophic) from all other populations studied. For S. striatinum,the most extensively studied species, the amount of shell CaCO₃is inversely related to water hardness. The selection of shellsin the Sphaeriidae is discussed in relation to structural-functionalneeds and habitat conditions ¹ Present Address: Department of Biology, Syracuse University,Syracuse, New York 13210, U.S.A. ² Present Address: Department of Zoology, Miami University,Oxford, Ohio 45056, U.S.A. (Received 5 December 1978;     

Possible Involvement of Abscisic Acid in Increases in Activities of Two Vacuolar H+-Pumps in Barley Roots under Aluminum Stress

Levels of abscisic acid (ABA) in barley roots increased upontreatment with AlCl₃. Treatment with AlCl₃ or ABA increasedboth ATP-dependent and PP_i-dependent H⁺-pumping activities intonoplast-enriched membrane vesicles. Increase in the H⁺-pumpingactivities caused by aluminum stress could result from increasedlevels of ABA. ¹Present address: Department of Botany, Faculty of Science,Hirosaki University, Hirosaki, Aomori, 036 Japan     

Preferential Loss of Chloroplast Proteins in Nitrogen Deficient Euglena

Two dimensional gel electrophoresis was used to follow changesin the relative amounts of over 500 cellular proteins duringnitrogen deficiency and in light limited nitrogen sufficientstationary phase Euglena cultures. Of 53 polypeptides whoserelative amount decreased in nitrogen deficient cells, 37 werechloroplast proteins and only 11 were mitochondrial proteins.This corresponds to a decrease in the relative amounts of 77%of the chloroplast proteins and 31% of the mitochondrial proteinsfound on the two dimensional gel map. Over a similar time period,the relative amounts of only 1 chloroplast and 1 mitochondrialprotein decreased in light limited nitrogen sufficient stationaryphase cultures. Many of the chloroplast proteins whose leveldeclined during nitrogen deficiency were proteins whose lightinduced accumulation is independent of chlorophyll synthesis,photosynthetic CO₂ fixation and the developmental status ofthe chloroplast. Taken together, these results indicate thatnitrogen deficiency triggers a preferential loss of chloroplastproteins which can not simply be explained through a dependenceof protein stability on chlorophyll levels or the developmentalstatus of the chloroplast. ¹Present address: The Mycology Center, Washington UniversitySchool of Medicine, Box 8050 St. Louis, MO 63178, U.S.A. ²Present address: Department of Biology, University of Tampa,Tampa, Florida 33060, U.S.A. (Received March 23, 1988; Accepted June 20, 1988)     

Characterisation of Non-Uniform Photosynthesis Induced by Abscisic Acid in Leaves Having Different Mesophyll Anatomies

The effects of abscisic acid (ABA) on photosynthesis in leavesof Helianthus annuus L. were compared with those in leaves ofVicia faba L. After the ABA treatment, the response of photosyntheticCO₂ assimilation rate, A, to calculated intercellular partialpressure of CO₂, P_i, (A(p_i) relationship) was markedly depressedin H. annuus. A less marked depression was also observed inV.faba. However, when the abaxial epidermes were removed fromthese leaves, neither the maximum rate nor the CO₂ responseof photosynthetic oxygen evolution was affected by the applicationof ABA. Starch-iodine tests revealed that photosynthesis was not uniformover the leaves of H. annuus treated with ABA. The starch contentwas diffferent in each bundle sheath extension compartment (thesmallest subdivision of mesophyll by veins with bundle sheathextensions, having an area of ca. 0.25 mm² and ca. 50 stomata).In some compartments, no starch was detected. The distributionof open stomata, examined using the silicone rubber impressiontechniques, was similar to the pattern of starch accumulation.In V.faba leaves, which lack bundle sheath extensions, distributionof starch was more homogeneous. These results indicate that the apparent non-stomatal inhibitionof photosynthesis by ABA deduced from the depression of A(pⁱ)relationship is an artifact which can be attributed to the non-uniformdistribution of transpiration and photosynthesis over the leaf.Intercellular gaseous environment in the ABA-treated leavesis discussed in relation to mesophyll anatomy. ¹ Present address: Department of Botany, Duke University, Durham,NC 27706, U.S.A. (Received September 30, 1987; Accepted January 13, 1988)     

Changes in the Translatable RNA Population during Abscisic Acid Induce Freezing Tolerance in Bromegrass Suspension Culture

Abscisic acid (ABA) has been shown to increase freezing toleranceof bromegrass (Bromus in-ermis Leyss cv. Manchar) cell suspensioncultures from a LT₅₀ (the temperature at which 50% cells werekilled) of –7 to – 30?C in 5 days at 23?C. Our objectivewas to study the qualitative changes in the translatable RNApopulation during ABA induced frost tolernace. In vitro translationproducts of poly(A)⁺ RNA isolated from bromegrass cells withor without 75 µM ABA treatment for various periods oftime were separated by 2D-PAGE and visualized by fluorography.SDS soluble proteins from the same treatments were also separatedby 20-PAGE. After 5 days treatment, at least 22 new or increasedabundance SDS soluble polypeptides were observed. From fluorographs,29 novel or increased abundance in vitro translation productscould be detected. The pattern of changes between ABA inducedSDS-soluble proteins and translation products from the 2D gelswere similar. A time course study (0–7 days) showed that17 of the 29 translation products were detected after 1 dayABA treatment, and at least 14 were present after 1 h. Coldtreatment (+4?C) induced fewer changes in the pool of translatableRNA than with ABA treatment. Three translation products inducedby cold appear to be similar to 3 of the ABA induced translationproducts. The majority of the ABA inducible translatable RNAsappeared at 10 µM or higher which coincides with the inductionof freezing tolerance. Many of these ABA inducible RNAs persisted7 days after ABA was removed from the media and correspondinglythe LT₅₀ (–17?C) was still well above the control level(–17?C). The results suggest that ABA alters the poolof translatable RNAs during induction of freezing tolerancein bromegrass suspension culture cells. ¹Oregon Agricultural Experiment Station Technical Paper No.9256. (Received August 3, 1990; Accepted October 18, 1990)     

Thermal Characteristics of C4 Photosynthetic Enzymes from Leaves of Three Sugarcane Species Differing in Cold Sensitivity

Phosphoenolpyruvate carboxylase, NADP-malate dehydrogenase andNADP-malic enzyme in desalted extracts from the leaves of threesugarcane species differing in cold sensitivity were relativelystable at cold temperatures, and their Arrhenius plots appearedas straight lines. Pyruvate,P_i dikinase (PPDK) from the threespecies was cold-inactivated, and its Arrhenius plots exhibiteda clear break-point around 10.6°C. Analysis of cold labilityof PPDK using deuterium oxide and Triton X-100 showed that theinteractions between the subunits possibly involve hydro-phobicbonds which would lead to cold lability. There were no apparentdifferences among the three sugarcane species in the thermalproperties of the four C₄ photosynthetic enzymes. The resultssuggest that the differences in cold sensitivity of sugarcanephotosynthesis may not relate to the thermal properties of C₄photosynthetic enzymes per se. ¹ Present address: Department of Biochemistry, University ofNebraska, Lincoln, NE 68588-0664, U.S.A.     

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)     

Membrane Potentials in Excitable Cells of Aldrovanda vesiculosa Trap-Lobes

The resting membrane potential in excitable cells of Aldrovandatrap-lobes is composed of diffusion and electrogenic potentials.The diffusion potential, about –100 mV in artificial pondwater, was determined from the external K⁺ and Na⁺ concentrations.The permeability ratio, P_Na/P_K of the membrane was estimatedto be about 0.3. The electrogenic potential hyperpolarized themembrane to about –140 mV. The peak value of the actionpotential increased by +26 mV with a tenfold increase in theexternal Ca²⁺ concentration. The action potential was blockedby an application of the Ca²⁺ chelater or the Ca channel blocker,LaCl₃. Cells showed additional Ca²⁺ influx (7.8 pmole/cm² impulse)during membrane excitation. These facts suggest that the transientincrease in Ca²⁺ influx causes the action potential presentin cells of Aldrovanda trap-lobes. ¹ Present address: Jerry Lewis Neuromuscular Research Center,School of Medicine, University of California Los Angeles, LosAngeles, CA90024, U.S.A. ² Present address: Biological Laboratory, Kyoritsu Women's University,Hachioji 193, Japan. (Received September 21, 1983; Accepted September 7, 1984)     

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; )     

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.     

Specific binding of gibberellin A1 to aleurone grain fractions from wheat endosperm

A subcellular fraction enriched in aleurone grains isolatedin glycerol from aleurone layers of wheat endosperm specificallyand reversibly bound GA₁-(³H). Specific binding of GA₁ to otherfractions including spherosomes, nuclei, mitochondria, and plasmamembranes was negligible. The K_d of binding to aleurone grainswas 1.5 µM and the number of specific binding sites 0.45pmoles per mg protein. The presence of Ca⁺⁺ ions was absolutelyrequired for binding. Abscisic acid which inhibits giberellinaction in vivo prevented specific GA₁-binding in vitro. GA₁-bindingto aleurone grains is important to the primary action of thehormone which may involve mobilization of reserves from thealeurone grain-spherosome complex for utilization in membranebiogenesis. ¹ Present address: Section of Cytology, Yale University Schoolof Medicine, New Haven, CT 06510, U.S.A. ³ Present address: Laboratoire de Biologie V?g?tale, Ecole NormaleSup?rieure, 24 rue Lhomond, 75231 Paris, France. (Received March 28, 1977; )     

The presence of gibberellin A20 in Stevia rebaudiana and its significance for the biological activity of steviol

Gibberellin-like substances of stems and leaves from Steviarebaudiana were analyzed and gibberellin A₂₀ was identifiedby gas chromatography-mass spectrometry. The presence of GA₂₀in S. rebaudiana is significant for the interpretation of thegibberellin activity of steviol. It indicates that steviol,the C-13 hydroxykaurenoic acid, may function as a precursorfor C-13 hydroxy-gibberellins and not as a gibberellin analog. ¹ This work was supported by National Science Foundation GrantGB 17304 to M. R. and by a Research Grant-in-Aid from SigmaXi to L. M. A. The research described is from a dissertationsubmitted by L. M. A. in partial fulfillment of the requirementsfor the Ph.D. degree. ² Present address: Laboratory of Plant Morphogenesis, Departmentof Biology, Manhattan College, Bronx, N. Y. 10471, U. S. A. (Received June 12, 1978; )     

Effects of Osmotic Stress, Ionic Stress and IAA on the Cell-Membrane Resistance of Vigna Hypocotyls

The cell-membrane resistance (R_m) of Vigna hypocotyls was examined,and the effects of osmotic stress, ionic stress and IAA on R_mwere investigated. R_m decreased by 64 to 77% under osmotic stressin the presence of absorbable solutes (40 mM sorbitol, 15 mMKC1, 30 mM sucrose; or 40 mM sorbitol, 15 mM KC1, 30 mM sucroseplus 10^–4 M IAA) or under ionic stress (50 mM NaCl or50 mM KC1). R_m was not changed by perfusion with 10^–4M IAA. Therefore, the hyper-polarizations of the membrane potentialobserved in both cases should be ascribed totally to the activationof the electrogenic proton pump. Although R_m showed an increaseof 1.6 fold when the hypocotyls were subjected to osmotic stress(100 mM sorbitol or 100 mM sorbitol plus 10^–4 M IAA),83.6% or 92.4% of the hyperpolarization of the membrane potential(V_px was also the result of the activation of the pump. Theresults, calculated on the basis of the current source model,support the viewpoint that the hyperpolarization of the cellmembrane potential of Vigna hypocotyls under osmotic stress,ionic stress or in the presence of IAA is an expression of theactivation of the proton pump, and is not caused by an increasein R_m. ¹ Present address: Researchers and Planners of Natural Environment, Yotsugi Bldg. (2F), 1-5-4 Horinouchi, Suginami-Ku, Tokyo,166 Japan ² Present address: Graduate School of Integrated Science, YokohamaCity University, 22-2 Seto, Kanazawa-Ku, Yokohama, 236 Japan (Received February 14, 1991; Accepted July 24, 1991)     

PEP Carboxylases from Two C4 Species of Panicum with Markedly Different Susceptibilities to Cold Inactivation

Phosphoenolpyruvate carboxylase (PEPC) (EC 4.1.1.31 [EC] ) assayedin extracts of Panicum maximum Jacq. loses up to 50% of itsactivity after incubation for 60 minutes at 0C while the enzymefrom P. miliaceum L. is completely stable under these conditions.Following dilution at room temperature the enzyme from P. maximumis labile, while that from P. miliaceum is stable. The P. maximumenzyme can be largely stabilized against dilution and againstcold-inactivation by D₂O which stabilizes hydrophobic bondsand the compatible solutes proline, betaine and trimethylamine-N-oxide.Mineral ions, previously demonstrated to be protective againstcold inactivation of pyruvate, P_i dikinase from maize, provideno protection of P. maximum PEPC against either cold or dilution.The chaotropic ion SCN^- causes partial inactivation of the enzymefrom P. miliaceum in the cold. The possible interrelationshipbetween inactivation by dilution and inactivation by cold isdiscussed. The enzyme from both species, when assayed withoutpreincubation at low temperature, exhibits similar, slightlycurvilinear Arrhenius plots; and no differences were found betweenthe two species in the temperature dependence of photosynthesis. ¹Present address: Botany Dept., University of California, DavisCA 95616 U.S.A.