Chloroplast Starch Research Articles

Photosynthate partitioning into leaf starch as affected by daily photosynthetic period duration in six species

AbstractStarch accumulation rates increased up to five‐fold in the leaves of five out of six species examined when plants were shifted from a long (12 or 14‐h) to a short (7‐h) photosynthetic period. The five species that responded were corn (Zea mays L.), pangola (Digitaria decumbens Stent.), soybean (Glycine max L.) Merr.), spinach (Spinacia oleracea L.), and sugarbeet (Beet vulgaris L.) However, the rate of starch accumulation in alfalfa (Medicago sativa L.) was unaltered following a shift from a long to a short photosynthetic period. We conclude that photosynthate partitioning into chloroplast starch, at least in a wide variety of species, is a programmable process that can be manipulated by altering the length of the daily photosynthetic period.

THE EFFECT OF ORTHOPHOSPHATE CONCENTRATION AND EXOGENOUSLY SUPPLIED SUGARS ON THE DISTRIBUTION OF NEWLY FIXED CARBON IN SUGAR BEET LEAF DISCS

SUMMARYThe apportioning of newly fixed carbon in leaf discs of sugar beet (Beta vulgaris L.) is dependent on the incubation medium. Concentration of orthophosphate (Pi) influences the distribution of carbon between chloroplast starch and the cytoplasm, and the proportion of 14C incorporated into sucrose. In the presence of exogenous sucrose and glucose, starch synthesis is enhanced and gross changes in cytoplasmic metabolite pools occur. These observations are discussed in relation to sink/source interactions of whole plants. In the presence of mannose, which sequesters cytoplasmic Pi in leaves of sugar beet as mannose‐6‐phosphate, sucrose synthesis is severely inhibited and maltose accounts for up to 30 % of the fixed carbon. This is related to earlier reports of unusual accumulation of maltose.

Some Effects of Buthidazole on Corn (Zea mays) Photosynthesis, Respiration, Anthocyanin Formation, and Leaf Ultrastructure

The effect of the herbicide buthidazole {3-[5-(1,1-dimethylethyl)-1,3,4-thiadiazol-2-yl]-4-hydroxy-1-methyl-2-imidazolidinone} on photosynthesis, respiration, anthocyanin formation and leaf ultrastructure of corn (Zea mays L. ‘Pioneer 3780’) was studied following pre- or postemergence applications. Total photosynthesis and dark respiration were measured with an infrared CO2 analyzer in an open air flow system 12, 18, and 24 days after preemergence treatment with 0, 0.56, 1.12, and 2.24 kg/ha of buthidazole. The 0.56 and 1.12 kg/ha preemergence treatments had no effect on total corn photosynthesis even 24 days after treatment, whereas buthidazole at 2.24 kg/ha inhibited photosynthesis as early as 12 days. Total photosynthesis and dark respiration were also measured in whole plants, 30 cm tall, before herbicide application and 4, 24, 48, and 96 h after postemergence treatment with buthidazole at 0, 0.28, 0.56, 0.84, and 1.12 kg/ha. Following postemergence treatment, buthidazole inhibited total corn photosynthesis at any rate examined as early as 4 h after treatment. Neither pre- or postemergence buthidazole applications influenced respiration with the exception of a transitory increase caused by 0.56 kg/ha 12 days after preemergence treatment and by 0.84 and 1.12 kg/ha 4 h after postemergence treatment. Transmission electron micrographs revealed that buthidazole applied postemergence at 0.28 and 1.12 kg/ha reduced or prevented the accumulation of starch in bundle sheath chloroplasts as early as 24 h after treatment. Ultrastructural disruptions in some mesophyll chloroplasts of treated corn plants were also evident. Preemergence application of buthidazole at rates of 0.28, 0.42, 0.56, and 1.12 kg/ha inhibited anthocyanin formation indicating an alteration in corn metabolism.

A light-microscope study of dry and germinating spores of Funaria hygrometrica Hedw., in dark and light conditions

Abstract Observations on sections of Funaria hygrometrica spores confirm that dry spores store only lipids, mostly located in a central drop; the nucleus surrounded by cell organelles is in a peripheral position; a space is observed between the spore wall and cytoplasm. In light conditions, the distension phase is reached between 16 to 20 hrs and rhizoid develops very quickly (growth phase) between 24 to 48 hrs, whereas the protonema remains as a protuberance. After a week in the dark, only swelling phase is completed. After 4 hrs of hydration, a considerable amount of starch in chloroplasts, exine cracking, diameter increase and greening of chloroplasts is observed both in dark and light conditions.

Ultrastructural characteristics of Hydrilla leaf tissue

The general anatomy and leaf ultrastructure of Hydrilla verticillata (L.f.) Royle, a submersed vascular hydrophyte, are reported. The transversely sectioned leaf consists of only two contiguous epidermal layers, and the single midvein is composed of three to four concentric layers of cells. There are numerous intercellular spaces throughout the tissue. Electron microscopy confirmed the presence of sieve elements in the mid-vein area. P-protein, plastid inclusions and branched plasmodesmata leading into the companion cells are all considered indicative of phloem-type cells. Plastid inclusions are cuneate and lack associated starch granules, characteristics typical of monocots. The outer wall of the lower epidermis protrudes into the cell. Plant cells with wall ingrowths are termed ‘transfer cells’ and, in aquatics, function in the absorption of solutes from the external environment. In addition to the lower epidermis, spiny projections of the leaf margin could be included in this category. There is a thick fibrous layer which appears to be part of the outer wall of the upper epidermis. Secretory vesicles containing polysaccharide empty into this layer and into the wall. Cell nuclei are distinctly polymorphic. The granal arrangement, presence or absence of starch and peripheral reticulum in chloroplasts varies.

Ultrastructure and cytochemistry of Miocene angiosperm leaf tissues

Angiosperm leaf fossils (16.7-25 x 10(6) years by potassium/argon dating) referable to Zelkova were isolated from pyroclastic deposits in the Succor Creek Formation, Oregon. These fossils reveal a three-dimensional structure in mesophyll cell layers and vascular bundles, while transmission electron microscopy of fixed tissues reveals well-preserved chloroplasts with grana stacks and starch, as well as nuclei with condensed chromatin. Transmission electron microscopy indicates that cell walls retain a cellulosic microfibrillar architecture, while ultrastructural and cytochemical confirmation is presented for chloroplast starch and the presence of native cellulose. The preservation of cell walls and cytologic structures in fossil materials of this age indicates an unusually mild fossilization process attending the post-depositional maturation of associated sediments. The preservation observed appears to be associated with rapid physical dehydration coupled with endogenous fixation by organic acids (possibly tannins and/or chlorophyll derivatives).

Light-controlled sugar-starch interconversion in epidermal chloroplasts of light-grown cucumber hypocotyl sections and its relationship to cell elongation

Sugar-starch interconversion in epidermal chloroplasts of light-grown cucumber hypocotyl sections as a regulatory mechanism of the osmotic potential of the cell was studied in relation to cell elongation. The presence of chloroplasts in epidermal cells was confirmed by electron microscopy, and also the chloroplasts were shown to act as the site of sucrose-starch interconversion. Chloroplast starch formation was induced by light, which was more distinct in the presence of sucrose (50 mM). The starch formation was microscopically detectable even at 1 hr incubation in the light with sucrose. On the other hand, no starch formation was observed in the dark both in the presence and absence of sucrose. Red light was effective, but not blue light. A photosynthetic inhibitor, 3-(4-chlorophenyl)-1:1-dimethylurea, also inhibited starch formation. Thus, epidermal chloroplast starch formation was induced under conditions where cell elongation is small and the osmotic potential (Ψ 0) of the epidermal cell is high. The sugar quantity (free sugar and reducing sugar) as the osmotica of the cell was larger in the dark than in the light, whereas the quantity of starch was greater in the light than in the dark. It is assumed from these results that one of the regulatory mechanisms of the osmotic potential of the epidermal cells in sugar-starch interconversion which occurs in epidermal chloroplasts.

Characterization of Starch Breakdown in the Intact Spinach Chloroplast

Starch degradation with a rate of 1 to 2 microgram-atom carbon per milligram chlorophyll per hour was monitored in the isolated intact spinach (Spinacia oleracea) chloroplast which had been preloaded with (14)C-starch photosynthetically from (14)CO(2). Starch breakdown was dependent upon inorganic phosphate and the (14)C-labeled intermediates formed were principally those of the Embden-Meyerhof pathway from glucose phosphate to glycerate 3-phosphate. In addition, isotope was found in ribose 5-phosphate and in maltose and glucose. The appearance of isotope in the intermediates of the Embden-Meyerhof pathway but not in the free sugars was dependent upon the inorganic phosphate concentration. Dithiothreitol shifted the flow of (14)C from triose-phosphate to glycerate 3-phosphate. Iodoacetic acid inhibited starch breakdown and caused an accumulation of triose-phosphate. This inhibition of starch breakdown was overcome by ATP. The inhibitory effect of ionophore A 23187 on starch breakdown was reversed by the addition of magnesium ions. The formation of maltose but not glucose was impaired by the ionophore. The inhibition of starch breakdown by glycerate 3-phosphate was overcome by inorganic phosphate. Fructose 1,6-bisphosphate and ribose 5-phosphate did not affect the rate of polysaccharide metabolism but increased the flow of isotope into maltose. Starch breakdown was unaffected by the uncoupler (trifluoromethoxyphenylhydrazone), electron transport inhibitors (rotenone, cyanide, salicylhydroxamic acid), or anaerobiosis. Hexokinase and the dehydrogenases of glucose 6-phosphate and gluconate 6-phosphate were detected in the chloroplast preparations. It was concluded (a) that chloroplastic starch was degraded principally by the Embden-Meyerhof pathway and by a pathway involving amylolytic cleavage; (b) ATP required in the Embden-Meyerhof pathway is generated by substrate phosphorylation in the oxidation of glyceraldehyde 3-phosphate to glycerate 3-phosphate; and (c) the oxidative pentose phosphate pathway is the probable source of ribose 5-phosphate.

Chloroplast Phosphofructokinase

Chloroplast phosphofructokinase from spinach (Spinacia oleracea L.) was purified approximately 40-fold by a combination of fractionations with ammonium sulfate and acetone followed by chromatography on DEAE-Sephadex A-50. Positive cooperative kinetics was observed for the interaction between the enzyme and the substrate fructose 6-phosphate. The optimum pH shifted from 7.7 toward 7.0 as the fructose 6-phosphate concentration was taken below 0.5 mm. The second substrate was MgATP(2-) (Michaelis constant 30 mum). Free ATP inhibited the enzyme. Chloroplast phosphofructokinase was sensitive to inhibition by low concentration of phosphoenolpyruvate and glycolate 2-phosphate (especially at higher pH); these compounds inhibited in a positively cooperative fashion. Inhibitions by glycerate 2-phosphate (and probably glycerate 3-phosphate), citrate, and inorganic phosphate were also recorded; however, inorganic phosphate effectively relieved the inhibitions by phosphoenolpyruvate and glycolate 2-phosphate. These regulatory properties are considered to complement those of ADP-glucose pyrophosphorylase and fructosebisphosphatase in the regulation of chloroplast starch metabolism.

Inhibition of spinach-leaf phosphofructokinase by 2-phosphoglycollate

A considerable amount of the CO? fixed during photosynthesis in green plants is utilised for the synthesis of starch in chloroplasts. The possibility that the subsequent degradation of this starch might involve formation of triose phosphates which can be easily exported to the cytoplasm [ 1 J was recently strengthened by the detection, in this laboratory, of phosphofructokinase activity in chloroplasts [2] . Experiments described in this communication have confirmed that phosphoglycollate, a proposed intermediate of photorespiration, inhibits the activity of spinach-leaf phosphofructokinase. Strong inhibition of the chloroplast enzyme may significantly hinder the degradation of chloroplast starch during daylight hours.

Starch associated particles in D-glucosamine-treated leaf tissue

Light incubation of barley and wheat leaf segments with 20 mM D-glucosamine at pH 6.0 or pH 8.0 results in the formation of clusters of particles (150-225 A) closely associated with chloroplast starch grains which appear to break down at the periphery. Thes ultrastructural changes are apparent after 6 hours treatment. Only relatively minor changes can be detected in the dark. D-mannosamine has a similar though much less pronounced effect but no ultrastructural changes are observed with either D-galactosamine or N-acetyl D-glucosamine. When leaf segments are treated in the light with both D-glucosamine and tris (hydroxymethyl) aminomethane in equimolar concentrations, the different ultrastructural changes induced by each substance can be observed in the same chloroplast. The glucosamine-induced particles appear to contain protein.

Chloroplast structure and starch grain production as phylogenetic indicators in the lower Rhodophyceae

The position of starch grain production, the shape of the starch grains and the depth to which the pyrenoid is embedded in the chloroplast are used as indicators of evolution in the lower Rhodophyceae. A cell with cytoplasmic allantoid starch grains encasing the pyrenoid and no thylakoids between the chloroplast envelope and the pyrenoid is considered to be evolutionarily primitive. A cell with oval starch grains not associated with the pyrenoid and with a pyrenoid deeply embedded in the chloroplast is thought to be evolutionarily advanced. A polyphyletic origin of the Porphyridiales is discussed.

Role of Mitochondria in the Origin of Chloroplast Starch Grains

By phase microscopic observation of living palisade parenchyma cells in sections of Nicotiana excelsior leaves from plants previously placed in the dark for 72 hours, 30 to 45 minutes of light is found to induce mitochondria to remain stationary within the concavity of the chloroplasts and become round. Extending the illumination period to 60 to 90 minutes causes the stationary mitochondria in the concavity to change from a translucent to an opaque appearance, the change coinciding with the first appearance of starch as detected by blue staining of the grains with I(2)-KI. It is speculated that an interaction bearing some resemblance to the previously described interaction between mitochondria and the mobile phase of the chloroplasts may also operate in the starch grain phenomenon.

EFFECT OF 4'‐CHLOROGLUTARANILIC ACID ON GROWTH AND DEVELOPMENT OF SUNFLOWER SEEDLINGS

The potential growth‐regulating properties of 4'‐chloroglutaranilic acid (CGA) as well as oxygen and nitrogen isosteres were examined in whole‐plant bioassay systems utilizing sunflower seedling (Helianthus annuus L.). Test systems included both soil‐grown plants and hydroponic studies. Foliar applications of CGA produced growth inhibition which was detectable within 24 hr and which persisted for at least 30 days. Direct application to roots produced a growth inhibition which was 15% greater than foliar applications. Foliar or root application at 10‐5 m or greater concentrations produced gross changes in leaf morphology. Total plant‐height inhibition was principally associated with stem growth in an area located between primary leaves and the shoot meristem. CGA was found to be 32 % less active than IAA as a growth promoter, but was 76 % more active than IAA as a growth inhibitor; the most severely affected organ was the leaf and the least affected was the root system. Leaves from soil‐grown plants treated wth a 10‐3 m foliar application of CGA possessed several abnormalities. These included an increase in thickness and dry weight, a reduction in chloroplast starch vacuoles, extractable starch, soluble hexoses, and soluble proteins.

Suboptimal Temperature and Assimilate Accumulation in Leaves of ‘Pangola’ Digitgrass (Digitaria decumbens Stent.)1

This study was conducted to determine the effect of metabolic sink activity on mobilization at low temperature of chloroplast starch in a tropical grass. Entire plants or partially shaded plants of ‘Pangola’ digitgrass (Digitaria decumbens Stent.) accumulated photosynthate during a 12‐hr period of illumination of 30 klux at 30C. Plants were then either entirely or partially subjected to a suboptimal temperature of 10C for a dark period of 12 hr. Temperature treatments provided metabolic sinks at 30C in locations proximate to areas containing low temperature‐ immobilized assimilate. Distribution of reducing sugar, sucrose, and starch was determined. No evidence was found that the presence of active metabolic sinks negated the influence of suboptimal temperature in causing assimilate immobilization. Further, the action of suboptimal temperature in causing assimilate immobilization appeared to be located at the area of accumulation, the leaf or perhaps the chloroplas.

Effect of Tillering and Cool Nights on Photosynthesis and Chloroplast Starch in Pangola1

Clonal propagviles of Digitaria decumbens Stent. were given night‐temperature treatments of 30, 10, 8, and 6 C, with day temperatures of 30 C. Chloroplasts of actively tillering plants accumulated little starch and NCE (net carbon dioxide exchange) rates showed no effect of cool nights. However, chloroplasts of nontillering plants accumulated starch during the day. When night temperatures were cool translocation was apparently reduced and the starch remained within the chloroplasts. The greater chloroplast starch content of nontillering plants after cool nights was negatively correlated (1% level) with NCE rates. Plants developing new tillers and therefore possessing large assimilate sinks had faster NCE rates than nontillering plants.

Effects of temperature on accumulation of starch or lipid in chloroplasts of grapevine

The amount of starch accumulated in chloroplasts of grapevine leaves fell, with increasing temperature, from 23% of dry weight at 18/13° C (day/night) to 1% at 35/30° C, whereas total lipids in the leaf rose from 6 to 16% of dry weight. Preparations viewed by electron microscopy had large starch granules in the chloroplasts at 18/13° C, and large lipid droplets at 35/30° C. At 35/30° C chlorophyll content of chloroplasts was high and grana prominent, suggesting that the results were due to selective effects of temperature on metabolic pathways.

The ultrastructure of chromoplast development in red tomatoes

Plastid morphogenesis in ripening red tomato fruit tissues has been studied by light and electron microscopy. Color changes of the fruit flesh result from the metamorphosis of chloroplasts 3–6 μ in diameter into chromoplasts 15–48 μ in length and 1–2 μ in diameter, which accumulate lycopene as the major pigment component. In most of the red-fruited varieties examined, the chloroplast starch reserves and granal thylakoids disappear prior to or during the initiation of one or more pigmented crystalline plates. Retention of grana in many nearly mature chromoplasts of the Green Flesh mutant is apparently correlated with the continuation of chlorophyll synthesis in the ripening fruit. The crystalline plates are interpreted as flattened membranous sacs (thylakoids) that are usually initiated in close association with membranes apparently persisting from the chloroplast progenitor. Increased longitudinal growth of a given plate is thought to cause overall plastid elongation; transverse growth is accompanied by involution of the lateral edges of the membranous sac to produce a tubular, crystalline, myelinic configuration, which causes little increase in plastid diameter. Simultaneous growth of several plates within a single chromoplast may result in the inclusion of one tubular crystal within another, thus forming a “compound crystal.” The small tablet- or needle-shaped chromoplast crystals found in certain specimens were not typical of the specimens used in this study and therefore are treated only superficially. Even in the very late stages of chromoplast differentiation, the crystals and remaining stroma are surrounded by an intact plastid envelope, which can be consistently demonstrated at the ultrastructural level.

Biosynthesis of starch in chloroplasts.

The enzymic synthesis of ADP-glucose and UDP-glucose by chloroplastic pyrophosphorylase of bean and rice leaves has been demonstrated by paper chromatographic techniques. In both tissues, the activity of UDP-glucose-pyrophosphorylase was much higher than ADP-glucose-pyrophosphorylase. Glycerate-3-phosphate, phosphoenolpyruvate and fructose-1,6-diphosphate did not stimulate ADP-glucose formation by a pyrophosphorylation reaction. The major metabolic pathway for UDP-glucose utilization appears to be the synthesis of either sucrose or sucrose-P. On the other hand, a specific precursor role of ADP-glucose for synthesizing chloroplast starch by the ADP-glucose-starch transglucosylase reaction is supported by the coupled enzyme system of ADP-glucose-pyrophosphorylase and transglucosylase, isolated from chloroplasts. None of the glycolytic intermediates stimulated the glucose transfer in the enzyme sequence of reaction system employed.