Factors affecting production of COS and CS2 in Leucaena and Mimosa species

Carbon disulfide (CS₂) and carbonyl sulfide (COS) are colorless, foul-smelling, volatile sulfur compounds with biocidal properties. Some plants produce CS₂ or COS or both. When used as an intercrop or forecrop, these plants may have agronomic potential in protecting other plants. Most of the factors which affect production of these plant-generated organic sulfides are unknown. We determined the effects of sulfate concentration, plant age, nitrogen fixation, drought stress, root injury (through cutting), and undisturbed growth on COS production in Leucaena retusa or Leucaena leucocephala and the effect of some of these factors on CS₂ production in Mimosa pudica. In addition, we determined if organic sulfides were produced in all Leucaena species. When L. retusa and M. pudica seedlings were grown in a plant nutrient medium with different sulfate concentrations (50 to 450 mg SL^-1), COS or CS₂ from crushed roots generally increased with increasing sulfate concentration. COS production was highest (74 ng mg^-1 dry root) for young L. retusa seedlings and declined to low amounts (<5 ng mg^-1 dry root) for older seedlings. Nitrogen fixation reduced the amounts of COS or CS₂ produced in L. leucocephala and M. pudica. Under conditions of undisturbed growth, root cutting, or drought stress, no COS production was detected in 4-to 8-weeks-old L. retusa plants. COS or CS₂ or both was obtained from crushed roots or shoots of all 13 known Leucaena species.     

The effect of waxy mutations on the granule-bound starch synthases of barley and maize endosperms

The effects of waxy mutations on starch-granule-bound starch synthases (EC 2.4.1.18) in the developing endosperm of barley (Hordeum vulgare L.) and maize (Zea mays L.) have been investigated. Three granule-bound starch synthases in barley endosperm were identified by use of antibodies to known starch synthases, by reconstitution and assay of individual proteins from sodium dodecyl sulphate-polyacrylamide gels of granule-bound proteins, and by partial purification of proteins released by enzymic digestion of starch. These are proteins of 60, 77 and 90 kDa. Use of antibodies to known starch synthases and partial purification of proteins released by enzymic digestion of starch indicated that there may be at least four granule-bound starch synthases in maize endosperm: proteins of 59, 74, 77 and 83 kDa. Mutations at the waxy loci of both species affected only the 60- (barley) and 59-(maize) kDa isoforms. No evidence was found that other putative isoforms are altered in abundance or activity by the mutations. The contribution of our results to understanding of the starch synthase activity of intact starch granules and the mechanism of amylose synthesis is discussed.We are very grateful to Dr. Roger Ellis (SCRI, Dundee, Scotland) for the gift of barley seeds, and to Drs Roger Ellis, Alan Schulman and Cathie Martin for helpful advice and comments during the course of this work.     

Purification and characterization of fructan: fructan fructosyl transferase from chicory (Cichorium intybus L.) roots

Fructan: fructan fructosyl transferase (FFT, EC 2.4.1.100) was purified from chicory (Cichorium intybus L. var. foliosum cv. Flash) roots by a combination of ammonium sulfate precipitation, concanavalin A affinity chromatography, and anion- and cation-exchange chromatography. This protocol produced a 60-fold purification and a specific activity of 14.5 mol·(mg protein) ^–1·min^–1. The mass of the enzyme was 69 kDa as estimated by gel filtration. On sodium dodecyl sulfatepolyacrylamide gel electrophoresis and mass spectrometry, 52-kDa and 17-kDa fragments were found, suggesting that the enzyme was a heterodimer. Optimal activity was found between pH 5.5 and 6.5. The enzyme used 1-kestose, 1,1-nystose, oligofructan and commercial chicory root inulin (degree of polymerization 10) as donors and acceptors. Sucrose was the best acceptor but could not be used as a donor. However, at higher concentrations sucrose acted as a competitive inhibitor for donors of FFT. 1-Kestose was the most efficient and 1,1-nystose the least efficient donor. The purified enzyme exhibited -fructosidase activity, specially at higher temperatures and lower substrate concentrations. The synthesis of fructans from 1-kestose decreased at higher temperatures (5–50°C). Therefore enzyme assays were performed at 0°C. The same fructan oligosaccharides, with a distribution similar to that observed in vivo, were obtained upon incubation of the enzyme with sucrose and commercial chicory root inulin.Abbreviations Con A concanavalin A - DP degree of polymerization - FFT fructan: fructan fructosyl transferase - Fru fructose - Glc glucose - Kes 1-kestose - MALDI-TOF MS matrix-assisted laser desorption ionisation time of flight mass spectrometry - Nys 1,1-nystose - pI isoelectric point - SST sucrose: sucrose fructosyl transferase - Suc sucrose The authors would like to thank E. Nackaerts for valuable assistance. W. Van den Ende is also grateful to the National Fund for Scientific Research (NFSR Belgium) for giving a grant for research assistants. P. Verhaert is a research associate of the NFSR. This work was also supported by grant OT/91/18 from the Research Fund K.U. Leuven.     

Glutathione synthesis in maize genotypes with different sensitivities to chilling

The effect of chilling on enzymes, substrates and products of sulfate reduction, gultathione synthesis and metabolism was studied in shoots and roots of maize (Zea mays L.) genotypes with different chilling sensitivity. At full expansion of the second leaf, chilling at 12 °C inhibited dry weight increase in shoots and roots compared to controls at 25 °C and induced an increase in adenosine 5-phosphosulfate sulfotransferase and -glutamylcysteine synthetase (EC 6.3.2.2) activity in the second leaf of all genotypes tested. Glutathione synthetase (EC 6.3.2.3) activity was about one order of magnitude higher than -glutamylcysteine synthetase activity, but remained unchanged during chilling except for one genotype. During chilling, cysteine and glutathione content of second leaves increased to significantly higher levels in the two most chilling-tolerant genotypes. Comparing the most tolerant and most sensitive genotype showed that chilling induced a greater incorporation of³⁵S from [³⁵S]sulfate into cysteine and glutathione in the chilling-tolerant than in the sensitive genotype. Chilling decreased the amount of³⁵S-label incorporated into proteins in shoots of both genotypes, but had no effect on this incorporation in the roots. Glutathione reductase (EC 1.6.4.2) and nitrate reductase (EC 1.6.6.1) activity were constitutively higher in the chilling-tolerant genotypes, but showed no changes in most examined genotypes during 3 d at 12 °C. Our results indicate that in maize glutathione is involved in protection against chilling damage.Abbreviations APSSTase adenosine 5-phosphosulfate sulfotransferase - EC -glutamylcysteine - GR glutathione reductase - OSH glutathione - NR nitrate reductase We thank M. Suter for preparing [³⁵S]adenosine 5-phosphosulfate, Dr. A. Fleming (both our Institute) for correcting the English and M. Soldati (Eschlikon, Switzerland) for his help with the plant material. This work was supported by COST 814 Crop development for the wet and cool regions of Europe.     

Resetting biological oscillators—A stochastic approach

We present a stochastic approach to phase-resetting of an ensemble of oscillators. In order to describe stimulation-induced dynamical phenomena we develop a stochastic model which consists of an ensemble of phase oscillators interacting via random forces. Every single oscillator is submitted to a phase stimulus. The ensemble's dynamics is determined by a Fokker-Planck equation. The stationary states are calculated explicitly, whereas the transients are analysed numerically. If the stimulus of a given (non-vanishing) intensity is administered at a critical initial cluster phase for a critical duration T _crit the ensemble's synchronized oscillation is annihilated. A transition from type 1 resetting to type 0 resetting occurs when the stimulation duration exceeds T _crit. Stimulation causes a shift of the mean frequency of every single oscillator. This frequency shift is explicitly calculated by deriving the mean first passage time. The model shows that there is a subcritical intensity which is connected with an enhanced vulnerability to stimulation. The desynchronized states, the so-called black holes, are typically associated with a double peak in the ensemble's phase distribution. This is important for analysing experimental data because simple peak-detection algorithms are not able to extract the underlying dynamics.Our results are discussed from the experimentator's point of view so that the insights derived from our model can improve data analysis and design of stimulation experiments.     

DPB1 * BR: an Mhc class II DPB1 allele (DPB1 * 6601) of negroid origin

The nucleotide sequence data reported in this paper have been submitted to the EMBL nucleotide sequence database and have been assigned the accession number X96986. The nameDPB1 ^* 6601 was officially assigned by the WHO Nomenclature Committee in May 1996. This follows the agreed policy that, subject to the conditions stated in the most recent Nomenclature Report (Bodmer et al. 1995), names will be assigned to new sequences as they are identified. Lists of such sequences will be published in the following WHO Nomenclature Report     

A new gene coding for an antigen recognized by autologous cytolytic T lymphocytes on a human renal carcinoma

Previous reports have described antigens that are recognized on human melanoma cells by autologous cytolytic T lymphocytes (CTL). The genes coding for a number of these antigens have been identified. Here we report the cloning of a gene that codes for an antigen recognized by autologous CTL on a human renal carcinoma cell line. This antigen is presented byHLA-B7 and is encoded by a new gene that we have namedRAGE1. No expression ofRAGE1 was found in normal tissues other than retina. RAGE1 expression was found in only one of 57 renal cell carcinoma samples, and also in some sarcomas, infiltrating bladder carcinomas, and melanomas. This represents the first identification of an antigen recognized by autologous CTL on a renal tumor.     

Developmental and genotypic differences in the response of pea stem segments to auxin

The objective of this investigation was to examine the response to exogenous auxin (indole-3-acetic acid; IAA)of stem segments at two developmental stages. The standard auxin response of excised stem segments and intact plants consists of an initial growth response and a prolonged growth response. We found that this biphasic response does not occur in internodes at very early stages. Stem segments of light grown pea of various genotypes were cut when the fourth internode was at 6–13% of full expansion (early-expansion) or at 18–25% of full expansion (mid-expansion). Length measurements of excised segments were made after 48 hours of incubation on buffer with or without auxin. An angular position transducer linked to a computerized data collection system provided high-resolution measurement of growth of stacks of segments incubated in buffer over 20 hours. Early-expansion segments of all genotypes deviated from the standard auxin response, while mid-expansion segments responded in a manner consistent with previous reports. Early-expansion segments of tall, light-grown plants were unique in showing an auxin-induced inhibition of growth. The auxin-induced inhibition correlated with high endogenous auxin content, as determined by HPLC and GC/MS, across genotypes and between early-expansion and mid-expansion segments of tall plants. Measurement of ethylene evolved from stem segments in response to auxin, and treatment of segments with the ethylene action inhibitor, norbornadiene, showed the inhibition to be mediated in part by heightened ethylene sensitivity. Growth of early-expansion segments of dwarf and severe dwarf plants was stimulated by exogenous auxin, but the growth rate increase was delayed compared to that in mid-expansion segments. This is the first time that such a growth response, termed the delayed growth response has been emonstrated. It is concluded that developmental stage and endogenous hormone content affect tissue response to exogenous auxin.     

Drought signal transduction in plants

Water deficit is one of the most common environmental limitations of crop productivity by affecting growth through alterations in metabolism and gene expression. The mechanisms involved in drought perception and signal transduction pathways are poorly understood. The participation of the plant hormone abscisic acid (ABA) has been well established. ABA levels increase when there are changes in the environment that result in cellular dehydration. Different approaches have been taken to understanding the molecular responses to desiccation and how ABA regulates gene expression. Recent efforts have identified particular topics of importance in the dissection of the signal transduction pathway which are summarized as follows: physiological approaches: identification of signalling molecules. Genetic approaches: the use of mutants, and Molecular approaches: promoter analysis.