The relationship between transpiration and chlorophyll synthesis in etiolated squash cotyledons

Etiolated cotyledons of squash (Cucurbita moschata Duch. var.melonaeformis Makino cv. Tokyo) treated with kinetin, KCl orfusicoccin in the dark increased both transpiration in the darkand chlorophyll synthesis after exposure to light. The minimumperiod of kinetin treatment to stimulate transpiration was similarto that to accelerate chlorophyll synthesis. On the other hand,treatment of cotyledons with vaseline or abscisic acid markedlyinhibited both transpiration and chlorophyll synthesis, suggestingthat cytokinin-induced stimulation of chlorophyll synthesiscould be mediated through the effect on the stomatal opening. (Received December 7, 1977; )     

Influence of Light Intensity on Sorbitol Metabolism, Growth, and Chlorophyll Content of Malus Tissue Cultures

Apple stem callus cultures of Malus pumila Mill., cultivars,McIntosh and Cortland, and of Malus robusta Rehd. No. 5 weregrown under light intensities of o, 850, 3350, and 7800 lx ona medium with sorbitol as the carbon source. Cortland callusgrew better in dark whereas growth of Robusta was not influencedby light. Light did not influence growth of McIntosh callusbetween o and 3350 lx, although slight stimulation in growthoccurred at 7800 lx. The chlorophyll content of the culturesincreased with light intensity, although that of Cortland wasvery low at all intensities. Tissue analysis of cultures grownin light at 7800 lx and in dark indicated that the growth ofthese callus tissues as influenced by light was directly relatedto their sorbitol and carbohydrate content. McIntosh and Cortlandshowed opposite trends in the levels of starch, sorbitol, sucrose,glucose, and total carbohydrate in light and dark. Similar tothe growth response of Robusta, the levels of these carbohydratesin this callus were apparently not influenced by light. Sucrose,accumulating in concentrations of between 45 and 62 per centof total carbohydrate, tended to be the predominant carbohydrateconstituent in both light- and dark-grown cultures.     

Effect of Light on Alkaloid Accumulation in Cell Cultures of Nicotiana Species

The effect of light on alkaloid accumulation in a range of cellcultures of tobacco was determined. Cell suspension culturesof Nicoriana rabacwn L. cv. Wisconsin-38 with differing degreesof photosynthetic activity, callus cultures of N. glauca Graham,root cultures of N. rustica L. and shoot cultures of N. tabacumwere used. The alkaloid content of green illuminated cultureswas greatly reduced compared with non-green cultures grown inthe dark, but decreased accumulation did not correlate withincreasing photosynthetic activity. The accumulation of allof the major alkaloids was affected, regardless of the speciesof tobacco used. Transfer of N. glauca callus from the darkinto the light caused a decrease in alkaloid accumulation, whilemoving cultures from the light into the dark resulted in anincrease in alkaloid content. In root cultures light causeda reduction in growth, which affected alkaloid synthesis. Inshoot cultures there were only traces of alkaloid detectable,regardless of whether or not cultures were illuminated. Lightappeared to cause a non-photosynthetic suppression of alkaloidaccumulation in visibly undifferentiated cultures, and thiseffect was modified in visibly differentiated cultures. Key words: Nicoriana spp, tobacco, alkaloid accumulation, cell culture     

Characterization of Senescence in Lemna gibba G3: A Determinate Growth System

Frond senescence in Lemna gibba G3 was characterized, and itscontrol by light, ABA and kinetin investigated. The plant exhibitsa determinate growth pattern with a frond producing a set numberof daughter fronds before undergoing senescence and death regardlessof whether or not it flowers. When a frond was cut in half,the distal half (half frond) which lacks any meristem underwentrapid senescence as compared with intact fronds. In both intactand half fronds, the onset of senescence was accelerated byABA and retarded by kinetin. Continuous white light acceleratedsenescence in both intact and half fronds over the dark controls.Under different photoperiodic light regime, the pace of daughterfrond production is accelerated in proportion to the lengthof light period. In half fronds, however, very short photoperiodiclight treatments (e.g. 1L: 23D or 3L: 21D) rather delayed senescenceover the dark controls. Two separate light control systems operatingin opposite directions in Lemana senescence appear to exist. ¹Present address: Department of Biology, Yonsei University,Seoul 120-749, Korea ²Present address: U.S. Department of Agriculture, Aero SpaceBuilding, Rm. 323, 901 D Street, S.W., Washington, D.C. 20251-2200, U.S.A. (Received July 13, 1989; Accepted May 8, 1990)     

Effect of Light and NaCI Salinity on the Growth of Callus Cultures of Ipomoea pes-caprae (L.) R. Brown and Ipomoea batatas (L.) Lam

Callus cultures of Ipomoea pes-caprae and I. batatas were establishedon MS medium containing 10^–5 M 2,4-D and 10^–8 Mbenzyladenine. Ipomoea pes-caprae calli exhibited green pigmentationand grew better in the light than in darkness. Callus tissuesof I. batatas showed a pale-yellow colouration and they grewat the same rate in light as in dark conditions. I. pes-capraeand I. batatas callus cultures were sensitive to the presenceof 60 mM NaCl in the culture medium, the growth of the formerbeing more sensitive in light than in darkness. The significanceof the responses of I. pes-caprae callus cultures in relationto the mechanism of salt tolerance is discussed. Ipomoea batatas, Ipomoea pes-caprae, sweet potato, railroad vine, callus cultures, salinity, light     

Influences of Light Quality, Growth Regulators and Inhibitors of Protein Synthesis on Respiration of Protoplasts from Meristematic Cells in Barley Seedlings

The influences of light of different wavelengths and plant growthregulators on the respiration of protoplasts isolated from tissue0 to 5 mm above the basal intercalary meristem of barley (Hordeumvulgare L. cv. Patty) leaves were studied. Respiration was measuredusing oxygen electrodes and a Cartesian-diver technique. Red,far-red and blue light all stimulated respiration in the protoplastsbut not in mitochondria isolated from them. Gibberellic acid stimulated respiration in protoplasts but abscisicacid had the opposite effect. Physiological concentrations ofindole-3-acetic acid and kinetin had no influence in eitherdirection. Combinations of gibberellic acid with light of anywavelength always increased respiration. Red or far-red light treatments in the presence of abscisicacid decreased dark respiration and only blue light significantlyreversed the inhibitory effect of abscisic acid. Cycloheximidemarkedly increased dark respiratory activity; chloramphenicolwas without effect. These results indicate that mitochondrialactivity in the leaf basal intercalary meristem was partiallycontrolled through phytochrome and a blue light receptor, andby gibberellic and abscisic acids. Changes in cytosolic proteinsynthesis were important for the initiation of enhanced mitochondrialactivity in meristems. Hordeum vulgare L., barley, abscisic acid, Cartesian-diver microrespirometry, gibberellic acid, meristematic respiration, protoplasts     

Control of Hypocotyl Hook Angle in Phaseolus mungo L. Role of Growth Substances

Effects of growth hormones on the hook angle and light responseof Phaseolus mungo L. hypocotyl hooks are described and theresults are discussed with reference to the functions of otherparts of the seedling in controlling the growth and shape ofthe hook. Apically applied IAA (indolyl acetic acid) prevented hook openingin decapitated seedlings in the dark and in all the red-irradiatedseedlings. [¹⁴C]IAA experiments showed that only a small quantityof IAA (2–6 ?g per hook) was required to produce theseeffects, and that transport of IAA through the hook was negligibleand unaffected by red irradiation. ABA (abscisic acid) had little effect on the hooks or theirlight response. Applied ethylene and IAA-induced ethylene slightly closed thehooks, but only slightly reduced light-induced opening. IAAreduced the effect of ethylene in the dark, but after irradiationthe hooks appeared more sensitive to the ethylene in the presenceof IAA, resulting in light-induced hook closure. Basally applied kinetin (6-furfurylaminopurine) prevented decapitatedhooks from opening in the dark, especially when GA₃ (gibberellicacid) was also present. Some combinations of kinetin and GA₃(with high kinetin concentrations) also prevented light-inducedopening, but combinations with lower kinetin concentrationsallowed almost as much opening as was found in intact hooks. It is proposed that the terminal parts act by regulating thesupply of cytokinins and gibberellins from the basal parts,and that IAA does not mediate this funotion in this species. The results are compared with those reported for other species.     

Requirement of sodium chloride for the action of gibberellic acid in stimulating hypocotyl elongation of a halophyte, Salicornia herbacea L.

NaCl stimulated hypocotyl elongation of the halophyte Salicorniaherbacea L. grown either in light or dark. Its optimal concentrationwas around 0.1–0.2 M and its promoting effect was muchmore prominent in the dark. Gibberellic acid at 10^–5 Mstimulated hypocotyl elongation in light but not in the dark.Indole-3-acetic acid and kinetin were ineffective in promotinghypocotyl elongation. In light, gibberellic acid and NaCl synergisticallyenhanced hypocotyl elongation when both were given simultaneously.The action of NaCl could be replaced by KCl, but not by mannitol.Osmotic pressure of the epidermis of the Salicornia hypocotylincreased in response to gibberellic acid and/or NaCl treatment.Na⁺ content in the hypocotyl increased with NaCl application.Gibberellic acid and NaCl when given alone increased the extensibilityof the hypocotyl cell wall. Synergistic interaction in increasingthe extensibility was observed between gibberellic acid andNaCl. Stress-relaxation analysis of mechanical properties ofthe hypocotyl wall revealed that gibberellic acid and NaCl actedsynergistically in decreasing minimum relaxation time. Basedon these results, a possible mechanism by which gibberellicacid and NaCl regulate hypocotyl elongation of Salicornia herbaceaL., a typical halophilic plant, is discussed. ¹ Present address: Laboratory of Biology, Tezukayama College,Gakuen Minami, Nara 631, Japan. (Received June 13, 1978; )     

EFFECTS OF CHLORAMPHENICOL AND KINETIN ON UPTAKE AND INCORPORATION OF AMINO ACIDS BY TOBACCO LEAF DISKS

The effects of chloramphenicol and kinetin on uptake and incorporationof ³⁵S-methionine and some ₁₄C-amino acids have been investigatedin leaf-disks of Nicotiana rustica in light and dark. Chloramphenicolin a concentration of 1 mg per ml inhibits the uptake of aminoacids from 30 to 60 per cent compared with the water control.The incorporation of amino acids into bulk protein is stronglyinhibited in light (40 to 70 per cent), but only to a smalldegree in dark (10 to 20 per cent), as revealed also by ¹⁴CO₂-photosynthesisof the disks and following treatment with chloramphenicol indark. The stimulating effect of kinetin on uptake and incorporationof amino acids is dependent upon its concentration (10^–5to 10^–6 M ; but 10^–4 M solution inhibits stronglyboth uptake and incorporation). The stimulation seems to influencemore incorporation than uptake processes. Possible interactionsof chloramphenicol and kinetin in the protein metabolism oftobacco leaves have been discussed. (Received April 27, 1964; )     

Peroxidase and Senescence

Kinetin and a, á-dipyridyl prevented the rapid decreaseof chlorophyll content in detached oat leaves senescing in thedark. In the light, detachment caused a 27–40% rise in peroxidaseactivity and kinetin enhanced the enzyme in the segments byabout 80%. Darkness prevented any detachment-induced rise ofthe activity and decreased the stimulating action of kinetinand mechanical injury. The effect of dipyridyl on peroxidaseactivity in the dark was similar to that of kinetin. Kinetin enhanced the same distinctive isoperoxidases under lightand dark conditions. Neither horseradish peroxidase nor that extracted from oat leavesshowed any ability to hydroxylate free proline in vitro. A systemwhich supposedly led to peroxidase-catalysed proline hydroxylationyielded small amounts of hydroxyproline in the absence of theenzyme. Staining with Fast Blue BB salt in the presence of IAA as asubstrate after electrophoresis indicated that all detectedoat isoperoxidases had an IAA oxidase activity visually parallelingtheir peroxidase activity. Crude extracts contained IAA oxidaseinhibitors that could be partially or fully removed by dialysis. The possible significance of the rise in peroxidase activityduring senescence is discussed.     

Studies on the Growth in Culture of Plant Cells: VII. EFFECTS OF KINETIN ON THE CARBOHYDRATE AND NITROGEN METABOLISM OF ACER PSEUDOPLATANUS, L CELLS GROWN IN SUSPENSION CULTURE

Aspects of the carbohydrate and nitrogen metabolism of Acerpseudcplatanus, L cell suspension cultures grown on a syntheticmedium containing 2 per cent glucose and 1.0 mg/l 2,4-dichlorophenoxyaceticacid and kinetin either at 0.25 mg/l (low kinetin) or at 2.5mg/l (high kinetin) are described. High kinetin inhibits growthas measured by increase in cell number, packed-cell volume,and cell dry weight. Although not inhibitory to glucose utdization,high kinetin inhibits the O₂ uptake of the cells. Such cellscontain only a trace amount of fructose and their rate of O₂uptake can be raised to that of the low kinetin cells by a periodof fructose feeding. The O₂ uptake of both kinds of cell issensitive to malonate but the stimulation of O₂ uptake inducedby bis(hexafiuoroacetonyl)-acetone (‘1799’) at 0.2mM is much less with the high-kinetin than the low-kinetin cells.The enzymes phosphoglucoseiseomerase and glucose-6-phosphatedehydrogenase are much less active in the high-kinetin cells.Mitochondria isolated from both kinds of cells show good respiratorycontrol although slightly lower values for Q_O2(N), ADP/O ratioand control ratio are recorded with mitochondria from the highkinetin cells. Kinetin at 2.5 mg/l slightly reduces the ADP/Oratio of isolated mitochondria but at 4.0 mg/l their responseto ADP is completely suppressed. Extracellular hemicelluloseformed in presence of high kinetin has a reduced content ofgalactose and xylose and an increased content of glucose. Theseobservations indicate that the inhibition of respiration byhigh kinetin is mainly due to suppression of glucose conversionto other sugars rather than to inhibition of glycolysis or terminalrespiration. High kinetin decreases the rate of protein but not of amino-acidsynthesis. Suppression of the synthesis of particular proteinsmay be an important factor responsible for the reduced cellyield of the cultures in presence of high kinetin. The significanceof these observations to our understanding of the critical metaboliceffects of cytokinina is discussed. Acer pseudoplatanus cells release amino acids into their culturemedium early in the period of batch culture and largely reabsorbthem as incubation proceeds.     

Chloroplast Differentiation in Tomato Root Galls Induced by the Root Knot Nematode Meloidogyne incognita

Primary roots of tomato, Lycopersicon esculentum cv. Marglobe, were cultured aseptically on agar containing a standard nutrient formulation with or without kinetin. When secondary roots developed, cultures were inoculated with the root-knot nematode, Meloidogyne incognita. Following inoculation, the cultures were divided into two groups which were incubated either in total darkness or in 16-h light-8-h dark cycles. At 24 h, 1, 2, 3, and 4 wk after incubation, roots from all cultures were processed for transmission electron microscopy. Fine structural observation of the parenchyma tissue in galls from the inoculated cultures indicated that starch containing plastids or amyloplasts, which are usually present and remain undifferentiated in these root cells, developed into chloroplasts. These chloroplasts contained a membrane system indistinguishable from those found in leaves of intact plants. Although plastid development was not affected when uninoculated cultures were incubated in the light, differentiation of the amyloplast was induced when roots were cultured on the medium containing kinetin. These results suggest that plastid differentiation in the inoculated tissue may be influenced by an accumulation of kinetin in the gall, which is induced by the nematode and serves as the nutrient sink for its feeding.     

Cytokinin Effects on Leaf Architecture in Phaseolus vulgaris L.

Treatment of expanding primary leaves of bean plants (Phaseolnsvulgaris L. cv. Limburgse vroege) with benzyladenine (BA) orkinetin at 0.5 mM for five consecutive days resulted in thickerleaves showing a significant decrease in intercellular air spacevolume. Compared with control plants, exposed mesophyll cellsurface area was lower per unit tissue volume, but unchangedwhen expressed per unit leaf surface area. Stomata of treatedplants were not fully closed in the dark and they did not openas wide as controls in the middle of the light period, suggestingthat the treatment resulted in impaired stomatal action. Allthe effects mentioned were more pronounced after treatment withBA, compared to kinetin. In spite of their magnitude, the observedchanges in leaf structure and function did not seem to havean important effect on total leaf diffusion resistance to carbondioxide during the course of the light period. Key words: Cytokinins, Leaf architecture     

Morphogenesis in Yucca schidigera Roezl. Root Organ Cultures

Root development in suspension cultures of Yucca schidigerawas light-mediated. The green cultures consisted of roots, smalltissue aggregates and suspension cells. Roots possessed an apicalmeristem with a root cap, meristematic region and region ofdifferentiating tissues. Phloem, xylem vessels and tracheidsoccurred in discrete polyarch vascular bundles. Xylary wallthickening was reticulate, and endodermis and pericycle werepresent. Roots of intact Y. schidigera plants had a similardistribution of vascular tissues. Dark-grown cultures were cream-colouredand contained only lobed tissue aggregates and suspension cells. Yucca schidigera Roezl., tissue cultures, morphogenesis, root organ, light/dark     

The Effect of Salinity on Light and Dark CO2-Fixation of Salt-Adapted and Unadapted Cell Cultures of Atriplex and Tomato

The effect of salinity on light and dark CO₂,-fixation was determinedin cells of A triplex portulacoides and tomato (Lycopersiconesculenturn Mill.) grown in culture. CO₂,-fixation of tomatocells was also determined in cultures adapted to mannitol andpolyethylene glycol (PEG). Salinity up to 400 mM NaCI in thecase of A triplex and up to 50 mM in the case of tomato enhancedthe rate of light-induced CO₂,-flxation in unadapted cells.Higher salt concentrations led to a marked decline in CO₂-flxationin both species. In salt-adapted A triplex cells no declinein the rate of light CO²,-flxation was seen even at 500 mM NaCl.Dark CO₂,-fixation was approximately 40% and 80% of the lightfixation in control cell cultures of A triplex and tomato, respectively.No enhancement in dark CO₂,-flxation was seen as salinity wasincreased, but a decline was found at similar salt concentrationsthat decreased fixation in the light. Mannitol-and PEG-adaptedtomato cells fixed CO₂, at somewhat lower rates than the controlcells in the light but not in the dark. Key words: Salinity, CO₂-fixation, cell cultures, Atriplex, tomato     

An Investigation of Factors which Influence the Production of Abnormal Terpenoids by Callus Cultures of Andrographis paniculata Nees

Callus and suspension cultures of Andrographis paniculata havebeen shown to produce three new sesquiterpene lactones whichhave been named paniculides. A, B, and C and not the andrographolides(diterpenes) produced by the intact plant. Cultures derived from leaves, stems, hypocotyls, roots, andembryos accumulated paniculides, but no andrographolides weredetected. Conversely andrographolides, but no paniculides weredetected in extracts from leaves, stems and roots of intactseedlings. Modifications of the medium including the use of different syntheticauxins and the replacement of coconut milk with kinetin andcasein hydrolysate did not influence paniculide production bythe hypocotyl cultures. In addition an established root isolatecultured for 9 months on a simple synthetic medium without auxin,meso-inositol or coconut milk still synthesized significantquantities of paniculides. Diurnal fluctuations of temperature and light did not influencethe pattern of compounds produced although cultures grown incomplete darkness produced the individual paniculides in differentproportions. Possible explanations for these findings are discussed.     

Gametogenesis of Chlamydomonas in the dark

When cells of each mating type of Chlamydomonas sp. (Toyonakastrain) were transferred to a nitrogen-free medium and culturedat low temperature (10?C) for 10 to 12 hr, they showed matingactivity even in the dark. The highest activity observed inthe dark, at about 10?C, was similar to that in the light at20?C. Induction in the dark was suppressed by oxygen deficiency.It was also inhibited by addition of 1 to lOmin concentrationsof nutritional nitrogen compounds such as ammonium, nitrateor urea, and of cycloheximide (2 to 20 µg/ml), but notby addition of chloramphenicol, dihydrostreptomycin or spectinomycin.On the other hand, it was slightly promoted by addition of theophylline,aminophylline, cAMP, DB-cAMP, kinetin and abscisic acid. ¹Present address: Department of Animal Virology, The ResearchInstitute for Microbial Diseases, Osaka University, Suita, Osaka565, Japan. (Received June 3, 1976; )     

Germination of seeds in Jussiaea suffruticosa

Seeds of Jussiaea suffruticosa reach high germination percentagesonly when exposed to long periods of continuous illumination.The light reaction may be repeatedly reversed by short exposuresto red and far red light, thus being mediated by the phytochromesystem. Seeds also germinate at high percentages if exposedto various cycles of 1 hr light and 24 hr of darkness at 20°C.If the temperature in the periods of darkness is raised up to30°C or lowered to 10°C the promotive effect of lightis inhibited. High temperatures (35°C) during imbibitionhave a promotive effect, whereas a pure O₂ atmosphere decreasesthe response to light. KNO₃ and kinetin enhance the responseto light but do not provoke germination in the dark. Only ifseed coats are punctured or removed does germination in thedark occur. (Received January 14, 1969; )     

Physiological Studies of the Sulpholipids of Diatoms

The sulpholipids of three species of freshwater and marine diatomNitzschia palae Kutz, Navicula muralis Lewin and Navicula incertaGrün, have been investigated under various culture conditions.The plant sulpholipid, sulphoquinovosyl diglyceride, was predominantlysynthesized in the light rather than in the dark while the unknownsulpholipids, designated as U₁ and U₂, were produced more inthe dark than in the light. It was found that cells starvedof carbon or sulphate utilized their sulpholipid reserve assources of these materials. Generally, cultures incubated inthe light and bubbled with air (with or without CO₂) showeda high level of incorporation of ³⁶S into sulpholipids. In culturesbubbled with oxygen-free nitrogen the incorporation of tracerwas very small. The photosynthetic and respiratory inhibitors,DCMU and DNP appreciably reduced the amount of tracer incorporatedinto the sulpholipids.     

Gametogenesis in Chlamydomonas III. Daily fluctuation of sex-competence

The daily fluctuation of sex-competence in Chlamydomonas cellsgrown in synchronous vegetative cultures was investigated. 1)The sex-competence of cells in synchronous vegetative culturesfluctuated during the light-dark cycle; being high in the latedark phase to early light phase, low in the early dark phase,and highest at the end of the dark phase (D-12; the time ofrelease of daughter cells). 2) There was also a daily fluctuationin the sex-competence of cells in gametogenic cultures duringincubation in the dark except at a low temperature (7?C). Thisdaily fluctuation in gametogenic cultures in the dark occurredwithout cell multiplication. 3) The pregamete (and/or the latentgamete) discussed in the preceding paper (1) might have an inactiveform of sex-competence. (Received March 7, 1973; )