Paul Ziegler

Carbon dioxide assimilation and stomatal response of afroalpine giant rosette plants

Maximal rates of CO2 assimilation of 8–11 μmol m-2 s-1 at ambient CO2 concentration were measured for Dendrosenecio keniodendron, D. brassica, Lobelia telekii and L. keniensis during the day in the natural habitat of these plants at 4,200 m elevation on Mt. Kenya. Even at these maximal rates, the CO2 uptake of all species was found to correspond to the linear portion of the CO2 response curve, with a calculated stomatal limitation for CO2 diffusion of 42%. Photosynthesis was strongly reduced at temperatures above 15° C. In contrast to this sensitivity to high temperatures, frozen leaves regained full photosynthetic capacity immediately after thawing. Stomata responded to dry air, but not to low leaf water potentials which occurred in cold leaves and at high transpiration rates. During the day reduced rates of CO2 uptake were associated with reduced light interception due to the erect posture of the rosette leaves and with high temperatures. Stomata closed at vapour pressure deficits which were comparable in magnitude to those characteristic of many lowland habitats (40 mPa Pa-1).

Specific determination of starch hydrolyzing enzyme activities of leaves

Summary Endo- and exoamylase and debranching enzyme were isolated from wheat leaves and used to test the validity of assays proposed to be specific for determining discrete leaf amylolytic activities. Procedures using a blocked PNP-linked G7-maltooligosaccharide substrate and a mixture of PNP-linked GS- and G6maltooligosaccharides were specific for wheat endo- and exoamylase, respectively. An alternative procedure (named «βββ») for specifically assaying endoamylase with β-limit dextrin as the substrate was also developed and confirmed to be specific for wheat leaf endoamylase. These assay procedures were applied to crude leaf extracts from a variety of species. The procedure using PNPGS/G6 as substrate appears to be of widespread value as a sensitive and reliable assay of exoamylase activity in leaf extracts. Endoamylase activity in the extracts was in most casesvery low with BPNPG7 as substrate; the βββ procedure was in general more sensitive in detecting this type of activity. The ratio of the activity units obtained with the βββ procedure to those measured with BPNPG7 as substrate varied considerably with the species investigated. The suitability of using BPNPG7 to assay leaf endoamylase activity may depend on the relative ability of the corresponding enzyme to attack small oligosaccharide and larger branched substrates, respectively.

Lack of effect of the photosystem II-based herbicides diuron and atrazine on growth of photoheterotrophic Chenopodium rubrum cells at concentrations inhibiting photoautotrophic growth of these cells

Abstract The effects of the photosynthetic herbicides diuron and atrazine on the growth and photosynthesis of a photoautotrophic cell suspension culture of Chenopodium rubrum have been studied with regard to the possible basis of phytotoxicity of these herbicides. The cells can also grow photoheterotrophically in the presence of sucrose while maintaining a high chlorophyll (Chl) content and photosynthetic capacity. Atrazine and diuron strongly inhibit CO 2 -dependent O 2 evolution of both photoautotrophically and photoheterotrophically grown cells at submicromolar concentrations. Atrazine and diuron at submicromolar levels also inhibit photoautotrophic growth of the cells, but 10-fold higher concentrations of these herbicides have no effect upon photoheterotrophic growth while sucrose is still present in the culture medium. We conclude that any photooxidative damage alone caused by continued light absorption by an inhibited photosynthetic apparatus does not detectably inhibit cell growth. Only in the absence of an additional energy source are these herbicides inhibitory to growth. Such a cell culture could provide a means of screening potential photosynthetic herbicides for specificity of action. Thus, compounds inhibiting photoautotrophic growth but not photoheterotrophic growth of the same cells would be specific photosynthesis inhibitors.

Light‐induced degradation of storage starch in turions of Spirodela polyrhiza depends on nitrate

Light induces both the germination of turions of the duckweed Spirodela polyrhiza and the degradation of the reserve starch stored in the turions. The germination photoresponse requires nitrate, and we show here that nitrate is also needed for the light-induced degradation of the turion starch. Ammonium cannot substitute for nitrate in this regard, and nitrate thus acts specifically as signal to promote starch degradation in the turions. Irradiation with continuous red light leads to starch degradation via auto-phosphorylation of starch-associated glucan, water dikinase (GWD), phosphorylation of the turion starch and enhanced binding of alpha-amylase to starch granules. The present study shows that all of these processes require the presence of nitrate, and that nitrate exerts its effect on starch degradation at a point between the absorption of light by phytochrome and the auto-phosphorylation of the GWD. Nitrate acts to coordinate carbon and nitrogen metabolism in germinating turions: starch will only be broken down when sufficient nitrogen is present to ensure appropriate utilization of the released carbohydrate. These data constitute the first report of control over the initiation of reserve starch degradation by nitrate.

Cereal β-amylases : diversity of the β-amylase isozyme status within cereals

Summary β-Amylase (EC 3.2.1.2.) activity was determined in the kernels of 29 cultivars and inbred lines of seven cereals ( Triticum aestivum L., Triticum durum L., Hordeum vulgare L., Secale cereale L., Avena sativa L., Zea mays L., Sorghum vulgare L.) belonging to 4 tribes of the Gramineae . The ratio activity/mg kernels was found to be much higher for the Triticeae than for all other tribes. Attempts were made to identify - in one representative of each of these tribes - the «endosperm» and the «ubiquitous» β-amylase according to criteria previously established, namely comparison of antigenic specificities as well as in vivo and in vitro post-translational modifications (Daussant et al., 1991). All tribes investigated exhibit the «ubiquitous « type β-amylase. The «endosperm» isoenzyme, however, appears to be restricted to the Triticeae tribe. The results are discussed in terms of the significance of β-amylase for germination and with respect to current knowledge as to the genetics of cereal β-amylases.

The major β-amylase isoforms of wheat leaves correspond to one of two ubiquitously expressed β-amylase genes

Abstract Leaves of wheat ( Triticum aestivum L.), cv. Star) exhibit five distinguishable isoforms of a β-amylase (EC 3.2.1.2) considered to represent the tissue-‘ubiquitous’ type of exohydrolase common to all cereals. The object of this study was to determine whether the multiple leaf isoforms originate from different genes or reflect post-translational processing of an isoform first expressed in juvenile leaf tissue. Two different cDNAs encoding for β-amylase were isolated from leaves and each produced an active β-amylase protein upon heterologous expression in Escherichia coli . Transcripts of these two genes were detected in tissues of wheat leaves, roots, flowers and seeds. However, only one of the two heterologously expressed β-amylases appeared to correspond to the β-amylase isoforms detectable in non-endosperm wheat tissues. It exhibited specific sequence identities with, and electrophoretic mobility under non-denaturing conditions similar to, the initially expressed leaf β-amylase isoforms. As does the initially in vivo expressed leaf isoform, the heterologously expressed β-amylase was converted by a β-amylase-free wheat leaf extract into secondary isoforms which closely resemble β-amylase isoforms appearing in vivo upon the maturation of leaf tissue. The molecular masses and the N-terminal amino acid sequences of the heterologously expressed β-amylase, its secondary conversion products and the extractable leaf β-amylases indicate that at least the major components of wheat leaf β-amylase polymorphism reflect C-terminal proteolytic processing of a single β-amylase translation product.

Posttranslational origin of wheat leaf β-amylase polymorphism

Summary β-Amylase (EC 3.2.1.2.) from wheat leaves was resolved by anion exchange chromatography into five active species. These were designated as βI–βV in accordance with the order of their elution from the anion exchange column and their increasing R f -values upon non-denaturing polyacrylamide gel electrophoresis. The major proteins of the isolated activity species reacted with an immune serum raised against barley seed β-amylase and showed only very slight differences in molecular mass (all in the range of 57–59 kD). βI–βV thus appear to most characteristically represent differendy charged isomers of one protein. βV, which is the only β-amylase species found in very young leaf tissues, was convened to βII and βIII (which appear in vivo upon leaf tissue maturation) and βIV upon in vitro incubation at pH 6. Massive βV conversion was related to a specifically gelatin-degrading proteolytic activity in the protein extract. Both the conversion of βV to other isoforms and the gelatin-degrading activity were specifically inhibited by PMSF. It is concluded that the polymorphism exhibited by wheat leaf β-amylase largely results from limited proteolysis effecting significant charge modification of a singly expressed primary form of the exohydrolase.

Effect of the Photosynthetic Herbicides DCMU and Atrazine on the Growth of Photoautotrophic and Photoheterotrophic Cell Suspension Cultures of Chenopodium Rubrum

A structurally diverse range of herbicides, e. g., phenyl ureas, triazines and uracils, inhibit photosynthesis by apparently adhering to a plastoquinone binding site on the thylakoid membrane and thereby blocking photosynthetic electron transport (1). That atrazine neither binds to thylakoids nor interferes with light-dependent electron transport in triazine herbicide-resistant weeds (2) strongly indicates that this inhibition is responsible for the herbicidal action of these compounds. Some uncertainty still exists, however, as to the physiological basis of their action (3). Death of the plant may result from starvation due to obstructed photosynthesis, from the action of harmful radicals generated as a result of continued light absorption by a blocked photosynthetic electron transport system, or from a combination of both these alternatives. Inhibition of the photoheterotrophic growth of plant cell cultures by photosynthetic herbicides (4) supports the toxic radical hypothesis (5), but these results were obtained using concentrations of herbicides much higher than necessary to inhibit photosynthetic electron transport in vivo and may thus have revealed non-specific effects of the herbicides.