Accumulation of a glycine rich protein in auxin-deprived strawberry fruits

Growth of strawberry (Fragaria ananassa Duch. cv. Ozark Beauty) receptacles is regulated by auxin supplied from the achenes. The receptacle growth can be stopped at any stage by deachening the fruits, and can be resumed by exogenous application of auxin. In our earlier studies we demonstrated auxin regulated polypeptide changes at different stages of strawberry fruit development. Removal of achenes from fruits and growing the receptacles without auxin resulted in the time-dependent accumulation of 52,000 Mr polypeptide. Amino acid analysis revealed that the protein is rich in glycine. Our studies, with normal and variant strawberry receptacles, indicate that the synthesis and accumulation of this glycine-rich protein correlates with cessation of receptacle growth. These results suggest a possible role for the glycine-rich protein in cessation of growth.     

Fruit Set and Growth in Strawberry, Fragaria x ananassa Duch.

The application of GA₃in aqueous solution to leaves or flowersof hermaphrodite cultivars of strawberry, Redgauntlet and Rabunda,prevented growth of the receptacle despite hand pollination.This inhibitory effect occurred only when GA₃ was applied priorto anthesis. Although viable pollen was produced and germinatedto grow down the styles of treated plants, no seeds were formed.Receptacle growth failed underneath the unfertilized carpels,but the basal region devoid of carpels enlarged and ripened.The effect of GA₃ was the same in vivo and for flowers grownin vitro. ABA and BAP also inhibited growth of pollinated flowersin vitro, but neither substance stimulated growth of the baseof the receptacle. 2-NOA stimulated receptacle growth of pollinatedflowers but did not overcome the inhibitory effect of GA₃. Removal of fertile carpels 9 days after pollination preventedfurther receptacle growth. GA₃ treatment of the bare receptaclere-started growth but was less effective than 2-NOA. No growth substance treatment induced parthenocarpic developmentin these cultivars when unopened buds were emasculated and culturedwithout pollination, although GA₃ induced some swelling of thereceptacle base. Fragaria x ananassa Duch., strawberry fruit set, fruit growth, growth regulators     

Polyamines in normal and auxin-induced strawberry fruit development

The possible involvement of polyamines during strawberry ( Fragaria × ananassa Duch.) fruit development was investigated. Putrescine, spermidine, and spermine were identified in strawberry receptacles and achenes at all stages of development. Total (free) polyamine levels decreased from a maximum of 485 nmol g⁻¹ fresh weight at pollination to a minimum of 55 nmol g⁻¹ fresh weight in ripe receptacles. Total polyamine concentrations during corresponding stages of development were consistently higher in achenes than in receptacles, and ranged from 891 to 203 nmol g⁻¹ fresh weight. Removal of achenes from the surface of developing receptacles 10 days after pollination reduced receptacle growth, and re-initiation of growth by application of 1 m M α-naphtaleneacetic acid (α-NAA) was accompanied by a rapid increase in polyamine concentrations 24 h after treatment. Polyamine content per receptacle increased >3-fold in normally developing receptacles and in de-achened, auxin-treated receptacles 10 days after removal of achenes, but did not increase during this period in de-achened receptacles not treated with exogenous auxin. α-NAA increased growth and polyamine levels to a greater extent than the structurally related, but less effective auxin, β-NAA. Polyamine concentrations in receptacles with intact achenes remained similar to those of auxin depleted (de-achened) receptacles, implying that the concentration of these compounds may not be limiting following achene removal.     

Polyamines in normal and auxin-induced strawberry fruit development

The possible involvement of polyamines during strawberry ( Fragaria x ananassa Duch.) fruit development was investigated. Putrescine, spermidine, and spermine were identified in strawberry receptacles and achenes at all stages of development. Total (free) polyamine levels decreased from a maximum of 485 nmol g⁻¹ fresh weight at pollination to a minimum of 55 nmol g⁻¹ fresh weight in ripe receptacles. Total polyamine concentrations during corresponding stages of development were consistently higher in achenes than in receptacles, and ranged from 891 to 203 nmol g⁻¹ fresh weight. Removal of achenes from the surface of developing receptacles 10 days after pollination reduced receptacle growth, and re-initiation of growth by application of 1 m M α-naphtaleneacetic acid (α-NAA) was accompanied by a rapid increase in polyamine concentrations 24 h after treatment. Polyamine content per receptacle increased >3-fold in normally developing receptacles and in de-achened, auxin-treated receptacles 10 days after removal of achenes, but did not increase during this period in de-achened receptacles not treated with exogenous auxin. α-NAA increased growth and polyamine levels to a greater extent than the structurally related, but less effective auxin, β-NAA. Polyamine concentrations in receptacles with intact achenes remained similar to those of auxin depleted (de-achened) receptacles, implying that the concentration of these compounds may not be limiting following achene removal.     

The Development of Embryo, Endosperm, and Nucellus Tissues in Relation to Receptacle Growth in the Strawberry

Maleic hydrazide was applied to strawberry plants in an attemptto inhibit the growth of specific tissues within the developingachenes so as to trace their function as sources of stimulito the developing receptacles. Whole plants were sprayed at 1, 500 p.p.m. at various intervalsbefore and after the opening of the primary flower on the truss.Regular twice weekly sprays with various concentrations of 2-naphthoxyaceticacid at concentrations between 50 and 5oop.p.m. were also combinedwith the maleic hydrazide treatments. Observations were madeon the development and final weight of all fruits, and valuesobtained for the percentages of achenes in which there wereviable embryos at maturity. Achenes treated with maleic hydrazideat various intervals were examined to determine the effectsof the treatment on the development of the embryo, endosperm,and nucellus. If treatment with maleic hydrazide was delayeduntil the third day after anthesis the receptacles were ableto develop and to ripen successfully even though all the acheneson their surface were devoid of viable embryos. Maleic hydrazideat 1, 500 p.p.m. was totally inactive as an inhibitor of receptacleexpansion if treatment was delayed till later than the 10thday after anthesis. It is concluded that the embryo is not the source of the growthstimulus which promotes the development of the receptacle, andthat the endosperm probably does not become active as a majorsource of auxin-like substances until it has become cellularbetween 10 and 14 days after anthesis. The possible identityof the tissue controlling receptacle-growth in the period immediatelyfollowing pollination and fertilization is discussed.     

Auxin-regulated polypeptide changes at different stages of strawberry fruit development

The pattern of polypeptides at different stages of strawberry (Fragaria ananassa Duch. cv Ozark Beauty) fruit development was studied by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. An 81,000-dalton polypeptide appeared between 5 and 10 days after pollination. Polypeptides with molecular weights of 76,000 and 37,000 daltons were formed after 10 days. The control exerted by auxin in the stage-specific formation of polypeptides was investigated by stopping fruit growth after removing the achenes and reinitiating fruit growth by the application of a synthetic auxin, α-naphthaleneacetic acid (NAA). When the achenes were removed from the 5- and 10-day-old fruits, the fruits failed to grow, the 81,000 dalton polypeptide was not formed between 5 and 10 days, and the 76,000- and 37,000-dalton polypeptides were not formed between 10 and 20 days. Application of NAA to fruits deprived of auxin by removal of achenes resulted in the resumption of growth and also in the appearance of these polypeptides. Removal of achenes of the 5- or 10-day-old fruits and growing them without auxin resulted in the formation of 52,000- and 57,000-dalton polypeptides. These two polypeptides were not formed when NAA was applied to fruits after removal of achenes. Supply of NAA to auxin-deprived fruits 5 days after removal of achenes resulted in resumption of growth and also in the disappearance of these two polypeptides, pointing out their possible relation to the inhibition of fruit growth.     

Demonstration of auxin binding to strawberry fruit membranes

Presence of specific auxin-binding sites in strawberry fruit (Fragaria ananassa Duch. cv. Ozark Beauty) membranes has been demonstrated. These 1-naphthaleneacetic acid (NAA)-binding sites in the 80,000g to 120,000g fraction of the strawberry fruit membrane were pronase sensitive with an estimated equilibrium dissociation constant for NAA of 1.1 × 10⁻⁶ molar. The minimum concentration of NAA required to stimulate strawberry fruit growth was at least one order of magnitude higher than the minimum concentration of NAA required to stimulate corn coleoptile elongation. This was consistent with the higher equilibrium dissociation constant (lower affinity) for auxin binding to strawberry fruit membranes than to corn coleoptiles. Twelve auxin analogs, ranging from very strong to weak auxins, were tested for abilities to stimulate in situ strawberry fruit growth and to bind (displace or compete with NAA) to strawberry fruit membranes. The observed positive correlation (r = 0.74) between the in vitro binding to the 80,000 to 120,000 membrane fraction and the in situ biological activity of these analogs indicated that the NAA-binding sites in strawberry fruit membranes may represent physiologically relevant auxin receptors.     

Hormonal changes during non-climacteric ripening in strawberry

In contrast to climacteric fruits, where ethylene is known to be pivotal, the regulation of ripening in non-climacteric fruits is not well understood. In the non-climacteric strawberry (Fragaria anannassa), auxin and abscisic acid (ABA) are thought to be important, but the roles of other hormones suggested to be involved in fruit development and ripening are not clear. Here changes in the levels of indole-3-acetic acid (IAA), ABA, GA(1), and castasterone from anthesis to fully ripened fruit are reported. The levels of IAA and GA(1) rise early in fruit development before dropping to low levels prior to colour accumulation. Castasterone levels are highest at anthesis and drop to very low levels well before ripening commences, suggesting that brassinosteroids do not play an important role in ripening in strawberry. ABA levels are low at anthesis and gradually rise through development and ripening. The synthetic auxin, 1-naphthaleneacetic acid (NAA), can delay ripening, but the application of GA(3), the gibberellin biosythesis inhibitor paclobutrazol, and ABA had no significant effect. IAA and ABA levels are higher in the developing achenes than in the receptacle tissue and may be important for receptacle enlargement and ripening, and seed maturation, respectively. Contrary to a recent report, the biologically active GA(4) was not detected. The pattern of changes in the levels of the hormones are different from those reported in another well studied non-climateric fruit, grape, suggesting that a single consistent pattern of hormone changes does not occur in this group of fruit during ripening.     

The Effect of Applied Growth Substances on Development of the Strawberry Fruit: I. INDUCTION OF PARTHENOCARPY

A series of experiments is described in which various growthregulators were applied in lanolin emulsions to the flowersof two pistillate varieties of strawberry, in an attempt topromote parthenocarpic development of their receptacles. Parthenocarpic development was obtained in a large majorityof the treated flowers over a range of concentrations, varyingfrom 500 p.p.m. to up to 6,000 p.p.m., after treatment withthe following growth regulators; 4-(indole-3-) butyric acid,2-naphthoxyacetic acid, and 1-naphthaleneacetic acid. Gibberellic acid promoted growth of the ‘neck’ regionin Freya and increased the response of this variety to 4-(indole-3-)butyric acid. It also reduced the period of development of thefruit from anthesis till ripening. An attempt to promote parthenocarpic receptacle developmentwith 4-(indole-3-) butyric acid after the removal of all carpelsbefore pollination was unsuccessful. It was concluded that theactive growth substances probably stimulated growth initiallyin the tissues of the unpollinated carpels, which in turn promotedthe development of the receptacle.     

The Effect of Applied Growth Substances on Development of the Strawberry Fruit.: II. Interactions of Auxins and Gibberellins

Growth-rates of receptacles developing after pollination werecompared with those of a similar set developing parthenocarpicallyafter treatment with 2-naphthoxyacetic acid (2-NoA). The lattergrew slightly more rapidly during the first 10 days, and enteredthe final ripening phase at the same time as the pollinatedreceptacles, but in the intervening period the growth-rate ofparthenocarpic receptacles was much reduced.Applications of2-NoA and gibberellic acid separately and in combination werecompared at anthesis, and the effects of supplementary treatments10 days later were investigated. Either chemical was capableof stimulating initial parthenocarpic development and maturationof the ripe fruit, but neither one alone maintained rapid growththroughout the second phase of fruit development. Combinationsof 2-NoA and gibberellic acid at high levels, and 2-NoA anda mixture of gibberellins A₄ and A₇ at lower levels, appliedeither together at anthesis, or sequentially during developmenteffectively maintained growth rates throughout the whole developmentalperiod.Rates of fruit maturation varied according to the chemicalstimulus inducing parthenocarpy. Treatment with gibberellinsresulted in early maturity and relatively small fruits. Treatmentwith 2-NoA led to a longer period of fruit development and relativelylarger fruits at maturity in spite of a low rate of growth throughoutthe second growth phase.     

Molecular cloning and sequencing of a cDNA for an auxin-repressed mRNA: correlation between fruit growth and repression of the auxin-regulated gene

A complementary DNA (cDNA) library has been constructed in gt10 from poly(A)⁺ mRNA isolated from auxin-deprived strawberry receptacles. By differential plaque filter hybridization, a cDNA (SAR5) to an auxin-repressed mRNA has been isolated. The expression of the auxin-repressed gene is studied at various stages of normal fruit development and in fruits of variant strawberry genotype using SAR5 as a probe. Northern analyses of RNA isolated from pollinated and unpollinated fruits of various developmental stages revealed that mRNA corresponding to the SAR5 clone is repressed during normal fruit development, and the level of SAR5 mRNA is regulated by endogenous auxin. Furthermore, results with both normal and variant genotype strawberry fruit indicate that there is a positive correlation between growth of strawberry fruit and repression of mRNA corresponding to the SAR5 clone. The SAR5 cDNA has been sequenced and is 723 nucleotides in length. The deduced protein has 111 amino acid residues with a molecular mass of 12.5 kDa. The putative polypeptide starts at nucleotide position 20 and ends at 352. The molecular weight of the predicted polypeptide is in agreement with the molecular weight of the in vitro translated polypeptide of hybrid selected mRNA. A comparison of the nucleotide and deduced amino acid sequence of SAR5 with nucleotide and protein sequences in data banks has not revealed any homology to known proteins.     

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     

Effects of CO2 Enrichment at Different Irradiances on Growth and Yield of Wheat: I. EFFECTS OF CULTIVAR AND OF DURATION OF CO2 ENRICHMENT

Wheat, Triticwn aestivum L., the winter cultivars Hobbit andCappelle-Desprez, and the spring cultivars Sicco and KJeiber,were grown in normal air or air enriched with CO₂ either outdoorsin a glass-roofed cage or in controlled environment rooms. Inneither the winter nor the spring wheat was growth increaseddue to enrichment with CO₂ before anthesis. Enrichment of thetwo winter wheat cultivars increased shoot dry weight significantlyat 15 d after anthesis but produced no significant increasein grain yield. With the spring cultivars there was a significantincrease in shoot dry weight by 18 d after anthesis and thegrain yield was also larger due to an increase in grain size.Shoot weight increased because the stems were larger, and therewas a diversion of assimilate from grain growth to late tillerproduction. Root tissue comprised less than 20% of the totaldry matter at anthesis (for all cultivars); effects of CO₂ enrichmenton root growth appeared to be less important than effects onshoot and ear growth. Growth and yield responses to CO₂ enrichmentwere observed (for the spring cultivars) at irradiances of both250 and 635 µE m^–2 s^–1, but the effects weregreater at the lower irradiance. Key words: CO₂ enrichment, Wheat, Cultivar     

NaCl-and Gibberellic Acid-induced Changes in the Content of Auxin and the Activities of Cellulase and Pectin Lyase During Leaf Growth in Rice (Oryza sativa)

The interactive effect of NaCl salinity and gibberellic acidin the activities of cellulase and pectin lyase, and on thecontent of auxin and chlorophyll, has been determined duringleaf growth (fifth from base) in rice. The linear growth, chlorophyllcontent, activity of cellulase, and the auxin level of leaveswere markedly decreased when plants were exposed to salt stress(12 dS m^–1). However pectin lyase activity did not registerany significant alteration in the leaves of salt-stressed plantscompared with the control. Treatment of plants with gibberellicacid (GA₃) (10 ppm) increased the leaf growth and chlorophyllcontent with a concomitant rise in the activity of cellulaseunder stressed as well as non-stressed conditions. A markedincrease in the content of auxin was discernible in the leavesof salt-stressed plants treated with GA₃ compared with non-treatedsalinized ones. An appreciable increment in the activity ofpectin lyase in response to GA₃ administration was detectedonly in the leaves of non-stressed plants. These results indicatethat enhancement of cellulase activity and the augmentationof endogenous auxin content may be involved in the stimulationof rice leaf growth by GA₃ under saline conditions. Oryza sativa, rice, leaf growth, NaCl salinity, gibberellic acid, cellulase, pectin lyase, auxin     

Cell Wall Metabolism in Developing Strawberry Fruits

Cell wall metabolism was studied in strawberry receptacles (Fragariaananassa, Duchesne) of known age in relation to petal fall (PF).Polysaccharide and protein composition, incorporation of [¹⁴C]glucoseand [¹⁴C]proline by excised tissue, and the fate of ¹⁴CO₂ fixedby young, attached fruits were followed in relation to celldivision, cell expansion, fine structure, and ethylene synthesis. Cell division continued for about 7 d after PF although vacuolationof cells was already beginning at PF and the subsequent cellexpansion was logarithmic. There was an associated logarithmicincrease in sugar content per cell and a decreasing rate ofethylene production per unit fresh weight. During cell expansion radioactivity from [¹⁴C]glucose was incorporatedinto fractions identified as starch and soluble polyuronideand into glucose and galactose residues in the cell wall. Radioactivityfrom [¹⁴C]proline was also incorporated into the cell wall,but only 10 per cent of this activity was found in hydroxyproline.Correspondingly wall protein contained a low proportion of hydroxyprolineresidues. The proportion of radioactivity from ¹⁴CO₂ fixed byfruitlets remained constant in most sugar residues in the cellwall. The proportion of radioactivity in galactose fell, indicatingturnover of these residues. Between 21 and 28 d after PF receptacles became red and softenedbut there was no change in the rate of ethylene production.Cell expansion continued for at least 28 d. Tubular proliferationof the tonoplast and hydration of middle lamella and wall matrixmaterial had begun 7–14 d after PF but became extremeduring ripening. Associated with the hydration of the wall,over 70 per cent of the polyuronide in the wall became freelysoluble, and arabinose and galactose residues lost from thewall appeared in soluble fractions. There was no increase intotal polysaccharide during ripening and incorporation of [¹⁴C]glucoseinto polysaccharides ceased, although protein increased andincorporation of [¹⁴C]proline into wall protein continued.     

In Vivo Fixation of CO2 by Attached Pods of Brassica campestris L.

In vivo net CO₂ exchange characteristics of attached Brassicapods were studied during the entire period of their growth anddevelopment after anthesis. ¹⁴CO₂ was fed both from the externalatmosphere and internally through the pod cavity, and the anatomyof the pod-wall was examined microscopically. Stomata were observedin the outer epidermal layer of the pod wall. Net in vivo CO₂fixation by the pods was observed throughout the period of theirdevelopment and was maximum on day 42 after anthesis (DAA).Compared to the internal feeding experiments, ¹⁴CO₂ fixationfrom the external environment was very high. Apparent translocationof fixed carbon from the pod wall to seeds was rapid. Pod photosynthesiscontributed substantially to seed growth. pods, Brassica campestris L, CO₂ fixation, stomata     

Biochemical Evidence that the Higher Plant Photosystem II Core Complex is Organized as a Dimer

The organization of the pigmented multiprotein core complexof higher plant PS II has been examined. Oxygen-evolving PSII particles or thylakoid membranes of wild-type and Chi b-lessbarley were extracted with various glycosidic surfactants andelectrophoretically fractionated. The predominant multiproteincore complex II (CC II) fractions had sizes on gel electrophoresisof M_r=230,000 and M_r= 140,000 and were photochemically active.Both fractions had identical absorption spectra, contained thebeta-carotene-chl a-proteins (Cp47 and Cp43), the PS II reactioncenter subunits (Dl and D2), and the two cytochrome b₅₅₉ subunitsin unit stoichiometry. The M_r=230,000 fraction could evolveoxygen in the light and contained an M_r=33,O0O oxygen evolutionenhancer (OEE 33) polypeptide, whereas the M_r= 140,000 fractionlacked OEE 33 and could not evolve oxygen. The apparent sizesof the two fractions were also estimated by gel filtration asM_r=490,000 and M_r=220,000, respectively; the estimates by gelfiltration more accurately reflect their predicted sizes. Furtheranalyses by nondenaturing gel electrophoresis indicated thatCp47, Cp43 and the three OEE gene products probably occur ashomodimers in situ. Our data suggest that phosphorylation ofCC II subunits occurs when they are located in the oligomericform. We propose that the native state of the PS II core complexin higher plants is dimeric, and that this state, which waspreviously observed only in thermophilic cyanobacteria, is probablythe form present in all oxygenic organisms. (Received August 9, 1991; Accepted September 26, 1991)     

In Vivo Fixation of CO2 by Attached Pods of Brassica campestris L.

In vivo net CO₂ exchange characteristics of attached Brassicapods were studied during the entire period of their growth anddevelopment after anthesis. ¹⁴CO₂ was fed both from the externalatmosphere and internally through the pod cavity, and the anatomyof the pod-wall was examined microscopically. Stomata were observedin the outer epidermal layer of the pod wall. Net in vivo CO₂fixation by the pods was observed throughout the period of theirdevelopment and was maximum on day 42 after anthesis (DAA).Compared to the internal feeding experiments, ¹⁴CO₂ fixationfrom the external environment was very high. Apparent translocationof fixed carbon from the pod wall to seeds was rapid. Pod photosynthesiscontributed substantially to seed growth. pods, Brassica campestris L., CO₂ fixation, stomata