Photosynthesis and starch metabolism of chloroplasts during prolonged illumination

Summary The starch metabolism of the chloroplasts in the leaves of Stellaria media was studied by means of electron microscopy. During the night the starch grains diminished in size but did not disappear entirely. In the light they grew due to photosynthesis. After prolonged illumination of the plant the grains almost filled up the chloroplasts. However, after an illumination of 26–27 hr a sudded disintegration took place. This was apparently caused by the increased activity of -amylase observed in an earlier investigation to occur at this time in the chloroplasts. After the disintegration the starch grains of the chloroplasts showed irregular changes.The rate of photosynthesis and respiration was measured by an infra-red gas analyser. During prolonged illumination Stellaria media showed a rather intensive and constant rate of assimilation. The role of starch disintegration and -amylase synthesis in making possible this constant assimilation has been discussed.     

The Effect of 6-Benzylaminopurine on the Starch Metabolism of Stellaria media

The effect of 6-benzylaminopurine (BAP) was studied by measuring the amylase activity of leaf extracts and the starch content of chloroplasts in Stellaria media-plants cultivated for five days in solutions containing different quantities of BAP. Small amounts of BAP decreased amylase activity in weak-growing plants. In strong-growing ones the effect was variable. However, in all experiments very high amount of this growth regulator caused a great increase in amylase activity which was accompanied by a relatively high concentration of starch in the chloroplasts. The induction of amylase activity was thus not high enough to diminish the starch content. This is supposed to be due to the lack of a sink organ since the growth of roots and shoots was greatly inhibited in concentrated BAP-solutions (10^-5M or more). In these cases the plants usually died during the first week. In some cases small amounts of BAP increased the growth of S. media, and high concentrations inhibited it causing characteristic morphological changes, such as creeping growth pattern, small, thick leaves and the turning up of the leaf margins. Benzylaminopurine also caused increased resistance to aging, especially in low concentrations (10^–7–10^–6M).     

The ecological distribution of C4 and C3 grasses in the Hawaiian Islands

Summary Isoenzyme analyses were carried out on the hypotetraploid Stellaria media and the diploid Stellaria pallida occurring on abandoned sandy fields in the coastal dunes of the Netherlands. Amylase isoenzymes in germinating seeds of Stellaria media showed a striking intrapopulational variability, which is in accordance with the germination behaviour of the different subpopulations. In contrast to these findings no intrapopulational variation nor developmental change in isoenzyme pattern could be detected on basis of 14 other analysed leaf enzymes. For five enzymes there was a difference in isoenzyme pattern between the species Stellaria media and Stellaria pallida. Of these five enzymes two showed an interpopulational variation in Stellaria media. The fact that there was almost no variability of the isoenzyme pattern in Stellaria pallida could be explained by the permanent autogamous (cleistogamous) state of the species. Effects of polyploidy on the isoenzyme pattern and its activity could not be demonstrated. Consequences of the extreme difference between the monomorphic pattern of the leaf enzymes as opposed to the polymorphism of the amylase are discussed in view of the difference in germination ecology and lifecycle strategy of Stellaria media subpopulations.     

Studies on the seed transmission of cucumber mosaic virus in chickweed (Stellaria media) in relation to the ecology of the virus

Cucumber mosaic virus (CMV) was transmitted in the seed of infected Stellaria media plants. The rate of seed transmission varied both in manually infected plants (3–21%) and in plants grown from infected seed (21–40%). In naturally infected plants the rates of transmission found were 4–29%. Seeds recovered from field soil carried 4–5% infection and in infected seed placed in the soil the virus persisted for at least 5 months. Seed transmission of CMV also occurred in infected Lamium purpureum (4%), Cerastium holosteoides (2%) and Spergula arvensis (2%) but it could not be demonstrated in six other more common weed species in five botanical families. Seed transmission in Stellaria media occurred with a British (W) and an American (Y) strain of CMV. The virus was shown to occur in S. media pollen. The importance of CMV-infected S. media seed in the soil in relation to the epidemiology of the virus is discussed.     

Accumulation and Disintegration of Starch in the Chloroplasts of Stellaria media Grown under Constant Illumination

The persent paper contains a study un the starch content of the chloroplasts of the leaves of Stellaria media, performed by hsitochemical methods. Three leaves were collected at short intervals, fixed and cut in 10 μ sections, and stained according to the PAS-procedure. The starch content of single chloroplasts was measured by a microscope photometer. The chemical composition of the stainable material was demonstrated by enzyme specific experiments. At the end of a 12 hours' dark period the chloroplasts contained only traces of starch. Light caused the starch accumulation to begin and after a time chloroplasts seemed to be filled with starch. Subsequently, however, a sudden decrease in the starch content of the chloroplasts took place, in spite of constant illumination. A rather high level of starch content was restored in a few hours. The author puts forward some ideas about the nature of the factors causing the transitory disintegration of starch in illuminated leaves. The induction or the activation of certain enzymes seems to be the most probable explanalion.     

Effects of Poa trivialis, Stellaria media and Rumex obtusifolius on the growth of Lolium perenne in the glasshouse

In glasshouse experiments shoot dry weight, leaf area and tiller number of Lolium perenne, cv. S. 24, were compared when grown in monoculture and in competition with Poa trivialis, Stellaria media or Rumex obtusifolius at a range of densities and for different time intervals. The results showed that S. media and R. obtusifolius were more competitive than P. trivialis. Competition effects with S. media and R. obtusifolius appeared between the 4 and 6 wk stage, coinciding with the production of the first S. 24 tillers. Spraying with 2,4-D (0·84 kg acid equivalent/ha) for control of R. obtusifolius did not decrease grass yield whereas mecoprop (2·80 kg a.e./ha) for control of S. media damaged young L. perenne. It appeared that the optimum time for mecoprop treatment was between 4 and 6 wk after sowing when tillering had commenced. At this stage weed competition was only beginning and crop tolerance to the herbicide was adequate.     

Further studies on seed transmission in the ecology of some aphid-transmitted viruses

A small proportion (1–4%) of the seeds of Stellaria media extracted from fallow soil from three widely separated areas contained cucumber mosaic virus (CMV). S. media seeds buried for 21 months produced 5 % infected seedlings. S. media plants from Britain, N. America and Australia were least severely affected by the CMV strain obtained from their country of origin and showed more severe reactions when infected with two alien strains. Several weed species were experimentally infected with lettuce mosaic, turnip mosaic and cauliflower mosaic viruses but, although virus was detected in the seeds of some species, it was not transmitted to any of their seedlings.     

Physiological and ultrastructural changes in Stellaria media following treatment with fluroxypyr

The physiological and ultrastructural changes induced by fluroxypyr (4-amino-3, 5-dichloro–6–fluoro–2–pyridyloxyacetic acid) are investigated in a susceptible weed species, Stellaria media. The sequence of symptoms was an initial petiole curvature followed by stem elongation and thickening prior to leaf senescence, stem and petiole necrosis and plant death. Light and electron microscopy revealed leaf tissue containing cells with disrupted and swollen chloroplasts and a disrupted tonoplast. Within the stem tissue there was extensive meristematic differentiation and adventitious root development prior to stem tissue necrosis. Increased levels of reducing sugars and amino acids in treated foliar tissue indicated reserve mobilisation during the initial stages of symptom development. Fluroxypyr induced oat coleoptile elongation with an optimum concentration at 5 × 10^-5 m compared to 10^-4 m for auxin (IAA). A S. media shoot explant system revealed similar elongation which was maximal during the first 24 h of treatment. Uptake and incorporation of ¹⁴C-leucine was stimulated by 96 h after treatment. It is concluded that fluroxypyr activity against S. media is characterised by an initial cell elongation and reserve mobilisation, followed by extensive cell proliferation, increased RNA and protein synthesis, and ultimately plant death.     

Vorkommen von Tobacco Rattle-Virus in Roggen (Secale cereale) in der Bundesrepublik Deutschland

Natural occurrence of tobacco rattle virus in rye (Secale cereale) in the Federal Republic of Germany Near Bergen/Dumme in northern Germany, in an area known for the occurrence of tobacco rattle virus (TRV) on potatoes, 5–10 % of rye plants of one experimental plot were severely stunted and showed distinct chlorotic mottling of leaves. In these plants TRV was identified on the basis of particle morphology and serology (Immunoelectron microscopy and ELISA). TRV was also detected in Capsella bursa-pastoris and Stellaria media from an area adjacent to the rye plot but not in nearby commercial rye fields.     

A starch-accumulating mutant of Arabidopsis thaliana deficient in a chloroplastic starch-hydrolysing enzyme

The aim of this work was to identify enzymes that participate in the degradation of transitory starch in Arabidopsis . A mutant line was isolated by screening leaves at the end of the night for the presence of starch. The mutant had a higher starch content than the wild-type throughout the diurnal cycle. This accumulation was due to a reduction in starch breakdown, leading to an imbalance between the rates of synthesis and degradation. No reduction in the activity of endo-amylase (α-amylase), β-amylase, starch phosphorylase, maltase, pullulanase or D-enzyme could be detected in crude extracts of leaves of the mutant. However, native PAGE in gels containing amylopectin revealed that a starch-hydrolysing activity, putatively identified as an endo-amylase and present in wild-type chloroplasts, was absent or appreciably reduced in the mutant. This is the first time that a specific enzyme required for starch degradation has been identified in leaves.     

Rendiconti delle Sedute e Brevi Comunicazioni

Riassunto

Nell'ambito della specie collettiva Stellaria media (L.) Cir. l'A. pone in evidenza l'esistenza di razze diploidi e di razze tetraploidi.     

The relationship between the rate of entry of ioxynil and effects on photosynthesis in normal and drought-stressed Stellaria media

The entry of ¹⁴C-labelled ioxynil octanoate into leaves of Stellaria media has been measured for plants grown in dry or moist soil. Of the total herbicide applied, 1–3% entered the leaf by 24 h and 2–5% by 72 h after treatment. Entry into moist-grown plants proceeded at about twice the rate of that into drought-stressed plants. Despite the limited rate of entry, the inhibitory action of ioxynil octanoate on photosynthetic carbon dioxide exchange was rapid, inhibition within 24 h reaching 70–90% in moist-grown plants, and 30–70% in dry-grown plants. Plants grown under moisture stress contained greater concentrations of the pigments chlorophyll a, carotene and lutein (a xanthophyll) than did moist-grown plants, and ioxynil-induced breakdown of these pigments was more rapid in moist-grown plants. It is suggested that these factors contribute to the greater tolerance of drought-stressed S. media to applications of ioxynil octanoate. The importance of continuous measurements of herbicide action is discussed in relation to the value and interpretation of ¹⁴C uptake data.     

Study of the Morphological and Functional State of Higher Plant Tissues by Optical Coherence Microscopy and Optical Coherence Tomography

Comparative analysis of two optical methods—optical coherence tomography (OCT) and optical coherence microscopy (OCM)—was made for vital visualization of plant tissues in tomato (Lycopersicon esculentum Mill), spiderwort (Tradescantia pallida (Rose) D. Hunt), orach (Atriplex sp.), and leaves and seeds of medium starwort (Stellaria media L.). The obtained OCT- and OCM-images allowed the morphological and functional state of plant tissues to be assessed in vivo. A higher spatial resolution of the OCM method, as compared to OCT method, allowed plant morphological structures to be identified with greater confidence. The morphological and functional state of tissues can be monitored with a time resolution of 1–4 s in intact plants, without removing them from the habitat.__________Translated from Fiziologiya Rastenii, Vol. 52, No. 4, 2005, pp. 628–634.Original Russian Text Copyright © 2005 by Kutis, Sapozhnikova, Kuranov, Kamenskii.     

Anatomy, chloroplast structure and compartmentation of enzymes relative to photosynthetic mechanisms in leaves and cotyledons of species in the tribe Salsoleae (Chenopodiaceae)

Certain members of the family Chenopodiaceae are the dominant species of the deserts of Central Asia; many of them are succulent halophytes which exhibit C4-type CO2 fixation of the NAD- or NADP-ME (malic enzyme) subgroup. In four C4 species of the tribe Salsoleae, the Salsoloid-type Kranz anatomy in leaves or stems was studied in relation to the diversity in anatomy which was found in cotyledons. Halocharis gossypina, has C4 NAD-ME Salsoloid-type photosynthesis in leaves and C3 photosynthesis in dorsoventral non-Kranz cotyledons; Salsola laricina has C4 NAD-ME Salsoloid-type leaves and C4 NAD-ME Atriplicoid-type cotyledons; Haloxylon persicum, has C4 NADP-ME Salsoloid-type green stems and C3 isopalisade non-Kranz cotyledons; and S. richteri has C4 NADP-ME Salsoloid-type leaves and cotyledons. Immunolocalization studies on Rubisco showed strong labelling in bundle sheath cells of leaves and cotyledons of organs having Kranz anatomy. The C4 pathway enzyme phosphoenolpyruvate carboxylase was localized in mesophyll cells, while the malic enzymes were localized in bundle sheath cells of Kranz-type tissue. Immunolocalization by electron microscopy showed NAD-ME is in mitochondria while NADP-ME is in chloroplasts of bundle sheath cells in the respective C4 types. In some C4 organs, it was apparent that subepidermal cells and water storage cells also contain some chloroplasts which have Rubisco, store starch, and thus perform C3 photosynthesis. In non-Kranz cotyledons of Halocharis gossypina and Haloxylon persicum, Rubisco was found in chloroplasts of both palisade and spongy mesophyll cells with the heaviest labelling in the layers of palisade cells, whereas C4 pathway proteins were low or undetectable. The pattern of starch accumulation correlated with the localization of Rubisco, being highest in the bundle sheath cells and lowest in the mesophyll cells of organs having Kranz anatomy. In NAD-ME-type Kranz organs (leaves and cotyledons of S. laricina and leaves of H. gossypina the granal index (length of appressed membranes as a percentage of total length of all membranes) of bundle sheath chloroplasts is 1.5 to 2.5 times higher than that of mesophyll chloroplasts. In contrast, in the NADP-ME-type Kranz organs (S. richteri leaves and cotyledons and H. persicum stems) the granal index of mesophyll chloroplasts is 1.5 to 2.2 times that of the bundle sheath chloroplasts. The mechanism of photosynthesis in these species is discussed in relation to structural differences.     

Changes in the plastid ultrastructure duringSedum rotundifolium leaf development

The ultrastructure of plastids was investigated in succulent leaves ofSedum rotundifolium to examine their changes during development. Leaves were categorized as etiolated, immature, young, and mature, based on their developmental stage and size. Of particular interest were the features of the tubular inclusion bodies (TIBs) and starch grains. These, along with vacuole size, showed remarkable changes over time. Etioplasts of unexposed leaves had prolamellar bodies, abundant starch grains, large TIB, few plastoglobuli, and thylakoid systems. Membranes of the thylakoids were still continuous with those of the prolamellar body. The plastids were often influenced by the presence and profile of inclusion bodies and starch grains throughout the early stages. Morphology was highly variable in the etioplasts but consistently hemispherical or ovoid in mature chloroplasts. TIB was most abundant in the etiolated leaves, but disappeared completely with development. Starch grains also became significantly reduced in size. Both young and mature mesophyll cells exhibited a normal chloroplast ultra-structure and huge central vacuoles, with an extremely thin peripheral cytoplasm. Grana were extensive and comprised a large portion of the chloroplasts. Traces of peripheral reticulum were also discovered in the chloroplasts of expanded leaves. The implications of these ultrastructural changes in the tubular inclusions and starch grains are discussed with relevance to Crassulacean acid metabolism (CAM).     

Structural organization of chloroplast of tomato plants <Emphasis Type="Italic">Solanum lycopersicum</Emphasis> transformed by Fe-containing superoxide dismutase

In this work we analyzed the results of the transformation of tomato plants Solanum lycopersicum by gene FeSOD1 from Arabidopsis thaliana, equipped with the signal sequence for targeting into chloroplasts. PCR analysis showed that the gene was integrated into the genome of several tomato plants that underwent transformation followed by selection in the kanamycin-containing medium. Two lines, provisionally denoted as nos. 6 and 8, were selected from the independent transformants. Line 6 was characterized by a reduced growth rate and altered leaves and line 8, by normal growth and leaves typical for control plants. Both lines showed a significant increase in SOD activity. In line 8 the increase in SOD activity was accompanied by an increase of ascorbate peroxidase activity, and in line 6 this effect was not present. Electron microscopic analysis of parenchymal and guard cells of both lines was performed, with an emphasis on the ultrastructural organization of chloroplasts. It is shown that the chloroplasts of the two transgenic lines differ in the number and size of starch grains and deposited plastoglobules as well as in the organization of lamellae and grana. Taken together, the results indicate that the expression of the introduced gene FeSOD1 has a significant effect on metabolic processes in the plastids. The findings are discussed in relation to the hypothesis about the importance of low concentrations of ROS for the integration of structure and function of chloroplasts.     

Effects of Phytoplasma Infection on Growth and Photosynthesis in Leaves of Field Grown apple (Malus Pumila Mill. cv. Golden Delicious)

The contents of chlorophyll (Chl), leaf biomass, and soluble proteins were markedly decreased in phytoplasma infected apple leaves. Similar results were also observed for ribulose-1,5-bisphosphate carboxylase, ¹⁴CO₂ fixation, and nitrate reductase activity. In contrast, the contents of sugars, starch, amino acids, and total saccharides were significantly increased in phytoplasma infected leaves. In isolated chloroplasts, phytoplasma infection caused marked inhibition of whole photosynthetic electron chain and photosystem 2 (PS2) activity. The artificial exogenous electron donor, diphenyl carbazide, significantly restored the loss of PS2 activity in infected leaves. Similar results were obtained when F_v/F_m was evaluated by in vivo Chl a fluorescence kinetic measurements.     

Starch and apparent rate of photosynthesis of leaves of Lolium perenne grown at different temperatures

Summary Leaves from Lolium perenne plants grown at day/night temperatures of 15°/10° had slower rates of apparent photosynthesis at 15° than leaves from plants grown at 25°/20°. Electron microscopy showed a higher starch concentration in chloroplasts from low- compared with the higher temperature-grown plants. However, all differences in apparent photosynthesis were negatively associated with differences in size of mesophyll cells. It is suggested that the presence of starch per se had no effect on photosynthesis rate and that temperature during growth influences subsequent rates because leaves from the higher temperature have smaller cells and chloroplasts than those from the lower one.     

THE ROLE OF ALLELOPATHIC SUBSTANCES IN THE “SMOTHER CROP” BARLEY

The high degree of success of barley as a “smother crop” generally has been attributed to physical competition for nutrients and water. However, it was found that even in the absence of such competition, barley still inhibits germination and growth. This occurred both in mixed cultures receiving adequate nutrients and water and in germination tests. Aqueous leachates of seeds and roots of barley caused similar inhibition of germination and growth, thereby indicating an inhibitory allelopathic substance. A specificity of reaction was found, with the greatest inhibition occurring with Stellaria media (L.) Cyr., less with Capsella bursa-pastoris (L.) Medic. and tobacco (Nicotiana tabacum L.), and no significant effect with wheat (Triticum aestivum L.). A concentration effect and possible periodic production of the inhibitor were indicated. Living plants and aqueous leachates of living roots were more inhibitory than dead ones, thereby supporting the hypothesis of an active metabolic secretion of the allelopathic substance. Preliminary attempts to identify the active inhibitory components demonstrated the presence of alkaloids, with a much greater concentration of substance in the living than in the dead root leachates. The alkaloid, gramine, known to occur in barley, was found to have an inhibitory effect on the growth of Stellaria media and it is suspected as an active component of the root leachates. These results suggest that factors other than the previously assumed physical competition are involved in the mechanism of the “smother crop” barley.     

Regreening of senescent Nicotiana leaves. II. Redifferentiation of plastids

Single senescent leaves attached to decapitated shoots of Nicotiana rustica L. regreened, especially when treated with cytokinin. Regreening caused an increase in leaf thickness, due to cell expansion. Senescent leaf plastids (gerontoplasts) were smaller than green chloroplasts, with degenerated membrane systems and stroma, and larger plastoglobuli. At advanced senescence, micrographs showed disintegrating gerontoplasts, reduced numbers of plastids were counted, and regreening became variable. The redevelopment of grana and stroma in regreening plastids was accelerated by cytokinin. All plastids in regreening leaves were identifiable as redifferentiating gerontoplasts because of their content of plastoglobuli and starch. Immunogold labelling showed significant association of POR with etioplasts in cotyledons, but with mature plastids in regreening leaves. No proplastids or dividing chloroplasts were observed in regreening leaves. Plastids numbers declined during senescence and did not increase again during regreening. It is concluded that the chloroplasts of regreening leaves arose by redifferentiation of gerontoplasts.Keywords: Chloroplasts, cytokinin, Nicotiana, senescence, regreening.