Oxygen Diffusion in Lupin Nodules: I.VISUALIZATION OF DIFFUSION BARRIER OPERATION

Root nodules of Lupinus albus (L.) cv. Multolupa were subjectedto short- and medium-term stresses by lowering rhizosphere temperaturefrom 25 to 16°C (2 h), detopping plants (3 h), darkeningplants (21 h) or exposing roots to 20 mol m^–3 KNO₃ for4 d. All experimental treatments produced increases in oxygendiffusion resistance, compared with control plants. These correlatedwith structural changes in the nodule cortex, which is describedin detail for the first time. The most noticeable change isthe occlusion of intercellular spaces by a glycoprotein whichwas identified using the monoclonal antibody MAC236. This glycoproteinwas also found surrounding bacteria in intercellular spacesof the cortex of control nodules. Key words: Oxygen diffusion resistance, glycoprotein, nodules, nitrogen fixation, Lupinus albus     

Biosynthesis,cDNA and amino acid sequences of a precursor of conglutin δ, a sulphur-rich protein from Lupinus angustifolius

The biosynthesis of conglutin has been studied in developing cotyledons of Lupinus angustifolius L. Precursors of conglutin formed the major sink for [³⁵S]-cysteine incorporated by developing lupin cotyledons, and these precursors were rapidly sequestered into the endoplasmic reticulum. The sequence of a cDNA clone coding for one such precursor of conglutin was determined. The structure of the precursor polypeptide for conglutin predicted from the cDNA sequence contained an N-terminal leader peptide of 22 amino acids directly preceding a subunit polypeptide of M _r 4520, together with a linking region of 13 amino acids and a subunit polypeptide of M _r 9558 at the C-terminus. The amino acid sequence predicted from the cDNA sequence showed minor variations from that established by sequencing of the protein purified from mature dried seeds (Lilley and Inglis, 1986). These were consistent with the existence of a multi-gene family coding for conglutin . Comparison of the sequences of conglutin with those of other 2S storage proteins showed that the cysteines involved in internal disulphide bridges between the mature subunits of conglutin , were maintained throughout this family of proteins but that little else was conserved either at the protein or DNA level.     

Characterization of Storage Proteins in Daucus carota L.: Two Novel Proteins Display Zygotic Embryo Specificity

Although maturation-related proteins are well known in the endospermof albuminous seeds, an important question is whether the zygoticembryo possesses its own maturation proteins. We report on theisolation and partial characterization of storage proteins ofcarrot (Daucus carota L. var Nandor) dry achenes and isolatedzygotic embryos, using one- and two-dimensional electrophoresistechniques, HPLC and amino acid sequencing. The presence ofa series of abundant polypeptides showing charge heterogeneity,that are rapidly degraded upon germination, was revealed inthe endosperm. These proteins consisted of glycoproteins, themost abundant of which displayed a molecular mass (M_r) of 58,000,albumins of M_r 42,000 comprising at least one rß-1,3-glucanase,and two globulins of M_r 90,000 and 50,000–55,000 respectively,the second being an oligomer composed of three subunits of M_r13,000, 20,000 and 30,000. None of these storage proteins identifiedin the endosperm were detected in zygotic embryos. In contrast,two novel proteins were isolated from zygotic embryos, namelya globulin family of M_r 50,000 and pI 6.3–6.8, which wasnamed "daucin", and a late embry-ogenesis abundant (LEA) proteinfamily of M_r 25,000 and pI6.3–6.6, named "RAB25". Sincethe latter proteins are apparently absent of the endosperm,these results suggest that the maturation of carrot zygoticembryos requires its own specific set of storage and LEA proteins. (Received July 15, 1997; Accepted October 28, 1997)     

Lectin-like activity of lupin seed conglutin {gamma}, a glycoprotein previously referred to as a storage protein

The lupin seed glycoprotein conglutin was capable of bindingvarious exogenous and endogenous N-glycosylated proteins andalso specifically bound to a Sephadex G-75 column. The bindingactivity of conglutin was abolished by either addition of 10mM EDTA or by enzymic deglycosylation and acid treatment. Inthe latter case activity could be restored by the addition ofMn and Ca ions. Immunological homology at polypeptide level of conglutin withother legume lectins was found. Key words: Conglutin , glycoprotein, legume lectin, Lupinus albus     

Manganese Nutrition of Lupinus spp. Especially in Relation to Developing Seeds

Patterns of transport and accumulation of manganese were studiedin Lupinus albus L. and Lupinus angustifolius L. in a wide rangeof availability levels in the rooting medium. The recently described‘split seed’ disorder, involving discolouration,splitting, and deformity of seeds, was reproduced in sand cultureusing critically low levels of manganese. The disorder was preventedby maintaining adequate manganese in the medium and its incidencein field and glasshouse was quantitatively related to the managneselevel in seed and fruit phloem sap. The use of phloem sap analysisfor early diagnosis of the disorder is suggested. High levelsof manganese in parent seed is suggested to afford protectionagainst the disorder by improving early vegetative growth ina manganese deficient situation. Direct carry-over of manganesefrom one seed generation to the next was insignificant. Manganese proved to be fully mobile in xylem but only sparinglymobile in phloem from vegetative structures to seed. It wasaccumulated in massive amounts in leaves and fruits when availabilitywas high. Seed manganese content increased 80–100 foldas the level in the rooting medium was increased from 0•1to 500 mg Mn 1^–1. L. albus was superior to L. angustifoliusin accumulating manganese in leaves and pods, and more efficientin translocating the element to its seeds. These differenceswere greatest at low or moderate manganese levels. Xylem intakeby a fruit was small relative to phloem intake when manganeseavailability was low, but became increasingly important as thesupply in the rooting medium was raised.     

Phloem and Xylem Transport in the Supply of Minerals to a Developing Legume (Lupinus albus L.) Fruit

The economy of carbon, nitrogen, water and mineral elementsin fruits of Lupinus albus L. was studied by measuring accumulationof these quantities in the developing fruit and estimating itstranspirational losses and CO₂ exchanges. Combining this informationwith data on levels of mineral elements in the xylem sap andphloem sap supplying the fruit, it was possible to test whethertransport based on mass inflow through xylem and phloem wouldaccount for the observed intake of elements. A model of transportbased on water and carbon intake suggested that vascular intakeduring the fruit's life matched the recorded increment for mineralsto within ± 15 per cent for N, Na, Zn, Fe and Cu, andto within ± 23 per cent for P, K and S. However, estimatedvascular intake of Ca, Mg and Mn accounted for less than one–thirdof the recorded intake by the fruit, inadequacy of vascularintake being especially great early in growth. Transport inphloem accounted for more than 80 per cent of the fruit's vascularintake of C, N and S, and 70–80 per cent of its P, K,Mg and Zn. Xylem contributed 68 per cent of the vascular inputof Ca, 59 per cent of the Na, and 34–38 per cent of theFe, Mn and Cu. Enclosure and darkening of fruits reduced levelsof Ca and Fe but increased levels of N, P, K and Zn in fruitdry matter relative to unenclosed, illuminated fruits. Resultswere related to previous observations on fruit functioning. Lupinus albus, legume fruit, mineral supply, phloem, xylem     

Stability of the Nitrogen Concentration of Inoculated White Lupins during Pod Development

The general pattern of decrease of the 'critical' plant N concentration(i.e. minimum concentration required for maximum growth rate)during growth has been described for several C₃ non grain-legumespecies, and this can be used as a reference curve for diagnosisof N nutrition in these species. The present study was undertakento investigate changes of N concentration during growth of agrain legume, in different conditions of N nutrition. Whitelupin (Lupinus albus L.) was grown for six crop seasons in fieldtrials in which inoculation with Rhizobium lupini, nitrogenfertilizer rate, cultivar and plant density were density weremanipulated. The yield and dry matter production of noninoculatedplants were lower than, or at the best similar to, those ofinoculated plants, whatever the level of N supply. From anthesisto the beginning of seed filling, the N concentration of shootsof inoculated plants was found to be remarkably stable betweenyears, N fertilization regimes, cultivars, and for individualplants within a plot. Nitrogen concentration only varied withplant density. By contrast, the N concentration of noninoculatedplants was highly variable and generally lower than that ofinoculated plants, whatever the level of N supply. The highand stable N concentration of inoculated plants did not appearto be necessary for maximum growth rate but seemed to be requiredfor maximum production of seed dry matter and N. The potentialuse of these results to diagnose, in any white lupin crop, aninefficiency of the lupin-R. lupini interaction is evaluated.Copyright1993, 1999 Academic Press Lupinus albus L., white lupin, N₂ concentration, inoculated plants, non-inoculated plants, N₂ fixation efficiency, diagnosis     

Identification and Characterization of the Seed Storage Proteins from Alfalfa (Medicago sativa)

The major seed storage proteins in alfalfa are medicagin (alegumin-like globulin), alfin (a vicilin-like globulin) anda family of Lower Molecular Weight albumins (LMW₁₃). These comprise30%, 10% and 20%, respectively, of the total extractable proteinfrom cotyledons of mature seeds. Alfin is a heterogeneous oligomericprotein (Mr 150 kD) composed of polypeptides ranging in sizefrom Mr 50 to 14 kD (₁,-₆; 50, 38, 32, 20, 16 and 14 kD, respectively).Medicagin is also a high molecular weight oligomeric protein,but requires high concentrations of salt for solubilization.It is comprised of a family of individually distinct subunits,each composed of an acidic polypeptide (A₁–A₉; Mr 49 to39 kD) linked via disulphide bond(s) to a basic polypeptide(B₁, B₂, B₃; Mr 24, 23 and 20 kD, respectively). This pairingis highly specific and two families are recognizable on thebasis of the B polypeptide (B₃ or B₁/B₂). Subunits (M_r 50–65kD) are assembled as trimers (8S) or larger oligomers (12S–15S)in mature seeds. The lower molecular weight albumins (LMW₁–₃)are acidic (pl<6), and consist of sets of disulphide-bondedpolypeptides (Mr 15 and 11 kD). Key words: Medicago sativa, seed storage proteins, alfin, medicagin     

Allocation of Organ Biomass in Perfect and Imperfect Flowers

Perianth M_P, gynoecium M_G, and androecium M_A dry-weight biomass(in g) of 39 species of perfect flowers was measured. Thesedata were pooled with published data from an additional 51 speciesand used to determine size-dependent variations in (M_G and M_A)in terms of the hypothesis that the quotient of M_G and M_A exceeds1·0 for out-breeding (xenogamous) species and less than1·0 for in-breeding (autogamous) species. Ordinary leastsquare regression of the pooled data (n = 90) showed M_G = 0·118M^0·916_P (r² = 0·884) and M_A = 0·186 M^0·975_P(r² = 0·865), indicating that the biomass of the gynoeciumproportionally decrease as floral size increases. The exponentsof these regressions indicate that the ratio of gynoecial toandroecial biomass decreased with increasing floral size suchthat comparatively small flowers (M_P < 0·0021 g) hadM_G/M_A > 1·0 (predicted for 'out-breeders') while comparativelylarger flowers (M_P > 0·0021 g) had M_G /M_A < 1·0(predicted for 'in-breeders'). Thus, on average, the type ofbreeding system was a size-dependent phenomenon. To test whether the biomass of a floral organ-type is a legitimateindicator of gender reproductive effort, the biomass (in g)of stamen filaments M_m and anther sacs M_AS of 39 species wasdetermined. Least square regression of these data showed M_AS= 0·188 M^0·854_fil (r² = 0·967), indicatingthat species with larger stamen filaments, on the average, boreproportionally smaller anther sacs and thereby cautioning againstthe uncritical use of the allocation of biomass to floral organ-typeas a strict gauge of gender-function investment. To determine whether the loss of one gender-function resultsin proportional reallocation of biomass to the remaining gender-function,the size-dependency of androecial and gynoecial biomass wasdetermined for a total of 33 perfect and imperfect flowers ofCucumis melo. Regression of the data obtained from perfect flowersyielded M_A = 0·402 M^1·47_P (r² = 0·898)and M_G = 4·63 M^1·36_P (r² = 0·842). SinceM_G/M_A M^0·11_P , the biomass allocation to the gynoeciumrelative to the androecium decreased with increasing floralsize. This result was consistent with the broad interpecificcomparison based on 90 species with perfect flowers . Regressionof the data for imperfect flowers yielded M_A = 0·151M^1·02_P (r² = 0·675) and M_G = 4·68 M^1·47_P(r² = 0·996), indicating a near allometric relation forthe androecium and a strong positive anisometry for the gynoecium.Thus, for flowers of comparable size, a loss of female genderobtains a modest to significant again in androecial biomasswhereas the loss of male gender yields only a slight increasein gynoecial biomass. Collectively, the results of these studies indicate that biomassallocation patterns are size-dependent phenomena whose complexitieshave been largely ignored in the literature.Copyright 1993,1999 Academic Press Allometry, floral biomass, reproduction     

Immunocytochemical localization of conglutin γ and legumin-like globulin in developing and mature seeds ofLupinus albus L.

Summary The distribution of two seed proteins, namely conglutin and a legumin-like globulin, in developing and mature seeds ofLupinus albus L. has been examined by immunocytochemistry and the concomitant modifications of their constituent polypeptides followed by SDS-PAGE. Both proteins were found within vacuolar protein bodies in various tissues of the cotyledons, although with some differences in the distribution patterns. The legumin-like protein was found to be deposited within the large storage parenchyma cells of the cotyledons in a manner similar to that reported for other storage proteins; little or no immunolabelling was associated with the cotyledonary epidermal and vascular parenchyma cells. In contrast conglutin was present in all cell types.A precursor of the legumin-like protein accumulated transiently in the developing cotyledon, but was subsequently modified by proteolytic cleavage. The onset of such modification was concomitant with a transition in the predominant vacuolar forms within the storage parenchyma cells. No precursor molecules of conglutin have been detected in this study, thus indicating that this protein is deposited in the protein bodies in its mature form.Abbreviations LM light microscopy - EM electron microscopy - DAF days after flowering - SDS-PAGE sodium dodecyl sulphate polyacrylamide gel electrophoresis - GAR goat antirabbit antiserum     

Density Studies on Lupins. II. Components of Seed Yield

Components of seed yield of cv. Ultra (Lupinus albus L.) andcv. Unicrop (L. angustifolius L.) were measured when grown atthree densities. The low density (10 plants m^–2) Unicropyield (34 g seed per plant) was 1.8 times that of Ultra as ithad more branches, pods and seeds per pod. Ultra seeds (310mg per seed) were heavier than Unicrop seeds (180 mg). The branchingpattern of Ultra was less dependent on plant density, henceat 93 plants m^–2 it gave a higher per plant yield (7.4vs 6.4 g) than Unicrop at lower densities (83 plants m^–2).Density had most influence on pod formation and only small effectson seeds per pod and seed weight. Yield components on the main-steminflorescence were influenced less by density than componentson branch inflorescences. Later formed, higher order generationsof inflorescences were most affected by increased inter- andintra-plant competition. Pod numbers on the main-stem were similarfor both species. Pods formed at higher flower nodes in Unicrop,but the lower flower nodes were less fertile than those in Ultra.Node position of flowers had no influence on seed set in main-stemUnicrop pods, but pods from higher nodes in Ultra formed fewerseeds. Seed weights in Unicrop were similar among main-stemnodes but in Ultra seed weights tended to increase at highernodes. Lupinus spp, lupins, seed yield, planting density     

Thylakoid Membrane Development and Differentiation: Assembly of the Chlorophyll a-b Light-Harvesting Complex and Evidence for the Origin of Mr=19, 17.5 and 13.4 kDa Proteins

Pea plants were grown under intermittent illumination (ImL)conditions. The low dosage of light given to ImL plastids limitedthe rate of chlorophyll (Chl) a and Chl b biosynthesis and,therefore, it retarded the rate of photosynthetic unit formationand thylakoid membrane development. Depending on the developmentalstage of the photosynthetic unit, ImL plastids had variableChl a/Chl b ratios (2.7 <Chl a/Chlb<20) and showed distinctintermediates in the assembly of the chlorophyll a–b light-harvestingcomplex (LHC) of photosystem-II (PSII). The results are consistentwith a step-wise increment in the PSII antenna size involvingthree distinct forms of the PSII unit: (i) a PSII-core formwith about 37 Chl a molecules; (ii) a PSIL_ß form containingthe PSII-core and the LHC-II-inner antenna with a total of about130 Chl (a + b) molecules, and (iii) the mature PSII_a form containingPSII_ß and the LHC-II-peripheral antenna with a totalof 210–300 Chl (a + b) molecules. The thylakoid membranecontained polypeptide subunits b, c and d (the Lhcb1, 2 and3 gene products, respectively) when only the LHC-II-inner waspresent. Polypeptide subunit a, (the apoprotein of the chlorophyll-proteinknown as CP29), along with increased amounts of b and c appearedlater in the development of thylakoids, concomitant with theassembly of the LHC-II-peripheral. The results suggest thatpolypeptide subunit d has priority of assembly over subunita. It is implied that, of all LHC-II constituent proteins, subunitd is most proximal to the PSII-core complex and that it servesas a linker in the transfer of excitation energy from the bulkLHC-II (subunits b and c) to the PSII-core. The work also addressesthe origin of low-molecular-weight proteins (M_r = 19, 17.5 and13.4 kDa) which co-isolate with intact developing plastids andwhose abundance decreases during plastid development. Aminoacid compositional and immunoblot analyses show a nuclear histoneorigin for these low-molecular-weight proteins and suggest co-isolationof histone-containing nuclear vesicles along with intact developingplastids. ¹Present address: Plant Physiology Research Group, The Universityof Calgary, Department of Biological Sciences, 2500 UniversityDrive N.W., Calgary, Alberta CANADA T2N 1N4.     

Comparative Studies of Acid Glycosidases from Three Legumes

The major isoenzymes of -mannosidase (EC 3.2.1.24 [EC] ) and ß-galactosidase(ECf 3.2.1.23 [EC] ) have been separated from cotyledons of gardenpea, Pisum sativum L. (Vicieae), chick pea, Cicer arietinumL. (Cicereae), and cowpea, Vigna unguiculata (L.) Walp. (Phaseoleae).Some of their properties have been determined, including pHoptima, K_m values for p-nitrophenyl glycosidc substrates, andthe effects of several inhibitors. Swainsonine, an indolizidinealkaloid, was the most effective inhibitor of mannosidase 1,with I₃₀ values of 5.6 x 10^–8 M (cowpea), 1x 10^–7M (chick pea) and 2.9 x 19^–7 M (pea). The most effectiveinhibitor of ß-galactosidase 2 from all sources wasD-galactonic acid-1,4-lactonwe (-lactone), with K_i values rangingbetween 3.0 and 3.9x 10^–3 M. An inhibitor of the E. coliß-galactosidose, p-aminophenyl thio-ß-D-galactopyranoside,did not inhibit any of the legume ß-galctosidases;rather it enhanced the activites of the enzymes from chick peaand cowpea cotyledons. Etiolated hull and seed tissues frompea pods developing in darkness contained similar acid glycosidaseactivities to normal green tissues, thus the chloroplast isan unlikely location for ß-galactosidase 2. The majorß-galactosidasesdetected with an indigogenic substrate (5-bromo-4-chloro-3-indoxyl-ß-D-galactopyranoside)following gel electrophoresis of extracts from pea hull, seedcoats and cotyledons appeared to be different from ß-galactosidase2. Acid glycosidase, cotyledon, isoenzyme, -lactone, legume, swainsonine     

Glycosidases from Cotyledons of Pisum sativum L.

-Mannosidase and ß-N-acetylglucosaminidase were purifiedfrom extracts of cotyledons of germinating Pisum sativum L.A 13-fold purification of a-mannosidase free from ß-N-acetylglucosaminidaseactivity was achieved by precipitation in ammonium sulphate,column chromatography on DEAE-cellulose, and treatment with2 M pyridine. ß-N-Acetylglucosaminidase was purified200-fold by the use of (NH₄)₂SO₄, and chromatography on ConcanavalinA¹-Sepharose and Sephacryl-200. This preparation showed no measurablecontamination by -mannosidase activity. Both glycosidases appearto be glycoproteins and demonstrate optimal activity at pH valuesof 4.0–4.5. Both glycosidases appear to have very similarmolecular weights, with -mannosidase being slightly larger thanß-N-acetylglucosaminidase. An extensive search forthe activity of aspartylglycosylamine amido hydrolase in peacotyledons proved unsuccessful.     

Thermosensitivity in Spring White Lupin

Spring morphotypes of white lupin (Lupinus albus L.) are veryresponsive to cool temperatures during seedling developmentas expressed in a number of subsequent developmental characters.Thermosensitivity was examined in a number of studies whereseedlings were incubated at various stages of development andat differing temperatures. Plants were scored for thermosensitivity,as expressed by the number of mainstem and first order lateralvegetative nodes, at flowering. The stages of radicle emergenceand split seed coat were observed to be the developmental stageswhen mainstem and first-order lateral meristems were most sensitiveto incubation temperature. These data show that the optimumwindow for cold treating spring lupin seed is very narrow andoccurs prior to seedling emergence. Seedlings at radicle emergencestage were incubated at temperatures ranging from 1 to 27 °Cfor 0 to 14 d in 2 d increments. For temperatures less than12 °C, mainstem node number decreased curvilinearly withduration of incubation. For temperatures between 12 and 17 °C,mainstem node number decreased linearly with duration of incubation.However, mainstem node number increased linearly with durationof incubation at temperatures greater than 17 °C. The datashow that 'warm' temperatures during early seedling developmenthave a profound influence on subsequent plant morphology. Responseto 'warm' temperatures is greater than to 'cool' temperaturesand it is suggested that the differential response to warm vs.cold incubation temperatures could account for morphologicaland phenological differences observed within genotypes in fieldstudies among sites and years. The lack of temperature responsein one genotype, 'Start' may indicate thermoneutrality in thisgenotype.Copyright 1995, 1999 Academic Press Lupinus albus L., lupin, vernalization, thermosensitivity, vegetative development     

Purification and Characterization of Arginine Decarboxylase from Soybean (Glycine max) Hypocotyls

Arginine decarboxylase (EC 4.1.1.19 [EC] ) was purified from soybean,Glycine max, hypocotyls by a procedure which includes ammoniumsulfate fractionation, acetone precipitation, gel filtrationchromatography, and affinity chromatography. Using this procedure,ADC was purified to one band in non-denaturing PAGE. The purifiedADC has an M_r of 240 kDa based on gel filtration chromatographyand is a trimer of identical subunits which has an estimatedM_r of 74 kDa based on SDS-PAGE. ADC is active between 30 and50°C and has a K_m value of 46.1 µM. ADC is very sensitiveto agmatine or putrescine but not to spermidine or spermine.In the presence of 0.5 mM agmatine (or putrescine), the enzymeactivity was inhibited by 70%. However, at the same concentrationof spermidine (or spermine), the enzyme activity was inhibitedby only 10–20%. (Received April 2, 1997; Accepted August 18, 1997)     

The Rate of Morphogenesis of Embryos and Seeds in Four Species of Grain Legumes

Comparative embryo development has been studied histologicallyin Lupinus albus, Lupinus mutabilis, Vicia faba, Pisum sativumand Latkyrus latifolius. The detailed histology of the stagesof embryo formation up to the early differentiation of tissuesof the seed is reported. The rate of embryogenesis has beentimed through 15 stages of development from anthesis and comparativerates of tissue formation established between the species. Themain observation was the slow rate of morphogenesis of embryosand seeds in Lupinus albus in comparison with the very rapidrate observed in Pisum sativum. A long period at the globularembryo stage, when embryo morphogenesis was inactive contributedto the extended development time of embryos and seeds in Lupinusalbus. Slow differentiation of reproductive tissues in L. albusdetermines late maturity in seeds and pods. Lupinus albus, white lupin, L. mutabilis, tarwi, Vicia faba, faba bean, Pisum sativum, pea, Lathyrus latifolius, everlasting pea, embryo development     

Effect of NaCI Salinity on Flows and Partitioning of C, N, and Mineral Ions in Whole Plants of White Lupin, Lupinus albusL.

Plants of Lupinus albus L., cv. Ultra, were grown hydroponicallywith NO₃^–-nutrition for 51 d under control (0.05 mol m^–3Na⁺ and 10 mol m^–3 Cl^–) and saline (40 mol m^–3NaCI) conditions. Plants were harvested 41 and 51 d after germinationand analysed for content and net increment of C, N and the mineralcations K⁺, Na⁺, Mg²⁺, and Ca²⁺ and the anions Cl^–, NOJ,malate, phosphate, and SO₄^2–. Roots, stem interaodes,petioles and leaflets were analysed separately. During the studyperiod net photosynthesis, respiratory losses of CO₂ from shootand root and the composition of the spontaneously bleeding phloemsap and the root pressure xylem exudate were also determined.Using molar ratios of C over N in the transport fluids, incrementsof C and N, and photosynthetic gains as well as respiratorylosses of C, the net flows of C and N in the xylem and phloemwere then calculated as in earlier studies (Pate, Layzell andMcNeill, 1979a). Knowing the carbon flows, the ratios of ionto carbon in the phloem sap, and ion increments in individualorgans, net flows of K⁺, Na⁺, and Cl^– over the study periodwere also calculated. Salt stress led to a general decrease of all partial componentsof C and N partitioning indicating that inhibitions were notdue to specific effects of NaCI salinity on photosynthesis oron NO₃ uptake. However, there were differences between variouslyaged organs, and net phloem export of nitrogenous compoundsfrom ageing leaves was substantially enhanced under saline conditions.In addition, NO₃^–reduction in the roots was specificallyinhibited. Uptake and xylem transport of K⁺ was more severelyinhibited than photosynthetic carbon gain or NO₃^– uptakeby the root. K⁺ transport in the phloem was even more severelyrestricted under saline conditions. Na⁺ and Cl^– flowsand uptake, on the other hand, were substantially increasedin the presence of salt and, in particular, there were thenmassive flows of Na^– in the phloem. The results are discussedin relation to the causes of salt sensitivity of Lupinus albus.The data suggest that both a restriction of K⁺ supply and astrongly increased phloem translocation of Na⁺ contribute tothe adverse effects of salt in this species. Restriction ofK⁺ supply occurs by diminished K⁺ uptake and even more by reducedK⁺ cycling within the plant. Key words: Lupinus albus, salt stress, phloem transport, xylem transport, partitioning, carbon, nitrogen, K⁺, Na⁺, CI^–     

A Protein from Immature Raspberry Fruits which Inhibits Endopolygalacturonases from Botrytis cinerea and other Micro-organisms

A polygalacturonase-inhibiting protein (PGIP) was purified fromimmature raspberry fruits using ion exchange chromatography.The protein was composed of a single polypeptide chain withM_r of 38·5 kDa and a pI residing above pH 10. Kineticstudies suggested that the inhibition was of a non-competitivenature. The PGIP inhibited two endopolygalacturonases (endo-PG)purified from Botrytis cinerea and an endo-PG produced by Aspergillusniger to varying degrees but did not inhibit two exo-PGs purifiedfrom B. cinerea, bacterial endopectate lyases and bacterialendo-PGs. The concentration of PGIP at various stages of flowerand fruit development was determined. The inhibitor was notdetected in the flower, but reached a maximum of 69 units g^–1in the immature green fruit decreasing to 9 units g^–1as fruits matured. The N-terminal amino-acid sequence was determined. Key words: Polygalacturonase-inhibiting protein, Rubus idaeus, red raspberry, Botrytis cinerea, pectinases     

Time-course of changes involved in the operation of the oxygen diffusion barrier in white lupin nodules

Nodulated white lupins (Lupinus albus L. cv. Multolupa) weresubject to either darkening for 12 h, followed by 24 h recoveryin light, or to 50% O₂ for 30 min. For each treatment, noduleswere harvested at intervals for analysis by light and electronmicroscopy and determination of glycoprotein content using EnzymeLinked Immunosorbent Assays (ELISA). This allowed for an analysisof the sequence of events causing an increase in intercellularspace occlusion within the inner cortex. The temporal sequencein response to darkening appears to be: (1) an initial rapidincrease in the detectable levels of intracellular glycoprotein,due to either a state change or de novo synthesis, (2) a concomitantincrease in the volume of thickened cell walls, causing a reductionof intercellular space volume and (3) after 1–3 h a releaseof glycoprotein into the intercellular space network of theinner cortex, accompanied (and possibly spread) by the continuedconstriction of the spaces due to cell wall and cell contentexpansion. The results for exposure to 50% O₂ showed a similar,but much more rapid, sequence of events, operating within 15–30min. The main difference between the two sequences was the lackof expansion of thickened cell walls with increased pO₂. Also,it was possible to detect glycoprotein within cell walls followingexposure to 50% O₂ but not following darkening. These observationsare discussed in relation to proposed mechanisms for the operationof a variable oxygen diffusion barrier in legume nodules. Key words: Oxygen diffusion resistance, glycoprotein, nodules, Lupinus albus