Enhancement of cadmium effects on growth and nutrient composition of birch (Betula pendula) by buthionine sulphoximine (BSO)

Binding of Cd to non-specific metal-binding peptides (phytochelatins)in birch roots has been suggested as an explanation for toleranceto Cd toxicity in birch (Betula pendula). In the present study,the tolerance of birch roots to Cd was further investigatedby using buthionine sulphoximine (BSO) as an inhibitor of phytochelatinsynthesis. Birch seedlings, grown in nutrient solution at pH4.2, were exposed to 0 or 2 µM CdCl₂ combined with 0 or0.1 mM BSO for 6 d. Plant growth (fresh weight increase andshoot to root dry weight ratio) and the nutrient compositionin fine roots, whole roots and shoots were determined. The effectsof Cd on growth confirms the results of earlier studies on birch,suggesting a reduced shoot growth, but preserved or stimulatedroot growth. When Cd and BSO were combined, overall plant growthwas severely reduced. BSO was also shown to aggravate Cd-inducedreductions of root and shoot concentrations of K, Ca and Mgbut to impede the accumulation of Cd. The results suggest that phytochelatins participate in protectingthe root against Cd interferences with growth, possibly by restrictingCd-induced changes in the nutrient composition of the plant. Key words: Betula pendula, buthionine sulphoximine, cadmium, phytochelatins, roots, tolerance     

The ultrastructure of the cuticle of Nematoda

 The ultrastructure of the body cuticle in species of six of seven representative genera of Stilbonematinae (Eubostrichus, Catanema, Laxus, Robbea, Leptonemella, and Stilbonema) was investigated using SEM and TEM techniques. Additionally, one species of Spirinia (Spiriniinae) and one of Desmodora (Desmodorinae) were studied for outgroup comparison. The body cuticle of all investigated stilbonematids shows a consistent pattern composed of specific elements in a characteristic arrangement to each other. This pattern does not occur in Stilbonematinae alone, but also in Desmodorinae and Spiriniinae. Furthermore, a comparison within the Desmodorida reveals that this cuticular pattern apparently is present in the cuticle of representatives of Monoposthiidae, Epsilonematidae, and Draconematidae. The present results lead to the following conclusions: (1) the cuticle of Stilbonematinae contains no autapomorphic characters for this taxon, (2) there is a common cuticular pattern within the Desmodorida, and (3) this pattern is an autapomorphic character for the order Desmodorida. Accepted: 4 February 1996     

Microbial transformation of azacarbazoles X: Regioselective hydroxylation of 5,11-dimethyl-5H-indolo [2,3-b]quinoline, a novel DNA topoisomerase II inhibitor, by Rhizopus arrhizus

Summary Microbial transformation of cytotoxic 5,11-dimethyl-5H-indolo[2,3-b]quinoline (a compound displaying antitumor activity and affecting the activity of calf thymus DNA topoisomerase II) was performed by the Rhizopus arrhizus strain and yielded a 9-hydroxy derivative. The metabolite obtained displayed a stronger cytotoxity against KB cells than the parent compound (ID₅₀=0.001 mol/mL), and stimulated also the formation of calf thymus topoisomerase II mediated pSP65 DNA cleavage in vitro at the concentration of 3 M. Being analogous to 9-hydroxyellipticine (which is an antitumor alkaloid), this novel indolo[2,3-b] quinoline derivative can be regarded as a novel potential antitumor agent.     

Tn5 insertion mutants of Pseudomonas sp. 267 defective in siderophore production and their effect on clover (Trifolium pratense) nodulated with Rhizobium leguminosarum bv. trifolii

Pseudomonas sp. strain 267 isolated from soil promoted growth of different plants under field conditions and enhanced symbiotic nitrogen fixation in clover under gnotobiotic conditions. This strain produced pyoverdine-like compound under low-iron conditions and secreted vitamins of the B group. The role of fluorescent siderophore production in the beneficial effect of strain 267 on nodulated clover plants was investigated. Several non-fluorescent (Pvd^-) Tn5 insertion mutants of Pseudomonas sp. strain 267 were isolated and characterized. The presence of Tn5 insertions was confirmed by Southern analysis of EcoRI digested genomic DNA of each derivative strain. The siderophore-negative mutants were compared to the parental strain with respect to their growth promotion of nodulated clover infected with Rhizobium leguminosarum bv. trifolii 24.1. We found that all isolated Pvd^- mutants stimulated growth of nodulated clover plants in a similar manner to the parental strain. No consistent differences were observed between strain 267 and Pvd^- derivatives strains with respect to their plant growth promotion activity under gnotobiotic conditions.Dr Deryto died in august 1994     

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.     

Keratinolytic fungi in sewage sludge

Sewage sludge from the Upper Silesia Region of Poland were surveyed for keratinolytic fungi. Out of 100 Petri dishes examined, 89 were positive for these micro-organisms. Altogether, 185 fungal appearances belonging to 10 species were observed. Trichophyton terrestre with its teleomorph Arthroderma quadrifidum, T. ajelloi with A. uncinatum, Microsporum gypseum with Arthroderma sp., and Chrysosporium keratinophilum with Aphanoascus keratinophilus prevailed in the sludges. The sewage treatment technologies together with the sludge structure, humidity and pH were found to be critical factors determining the occurrence of keratinolytic fungi in the sludge environment. The qualitative and quantitative composition of keratinolytic fungi could be a useful tool in evaluation of sludge treatment processes.     

Superhelical torsion controls DNA interstrand cross-linking by antitumor cis- diamminedichloroplatinum(II).

Negatively supercoiled, relaxed and linearized forms of pSP73 DNA were modified in cell-free medium by cis-diamminedichloroplatinum(II) (cisplatin). The frequency of interstrand cross-links (ICLs) formed in these DNAs has been determined by: (i) immunochemical analysis; (ii) an assay employing NaCN as a probe of DNA ICLs of cisplatin; (iii) gel electrophoresis under denaturing conditions. At low levels of the modification of DNA (<1 Pt atom fixed per 500 bp) the number of ICLs formed by cisplatin was radically enhanced in supercoiled in comparison with linearized or relaxed DNA. At these low levels of modification, the frequency of ICLs in supercoiled DNA was enhanced with increasing level of negative supercoiling or with decreasing level of modification. In addition, the replication mapping of DNA ICLs of cisplatin was consistent with these lesions being preferentially formed in negatively supercoiled DNA between guanine residues in both the 5'-d(GC)-3' and the 5'-d(CG)-3' sites. Among the DNA adducts of cisplatin the ICL has the markedly greatest capability to unwind the double helix. We suggest that the formation of ICLs of cisplatin is thermodynamically more favored in negatively supercoiled DNA owing mainly to the relaxation of supercoils.     

Isolation and characterization of extragenic mutations affecting the expression of the uroporphyrinogen decarboxylase gene (HEM12) in Sacharomyces cerevisiae

Uroporphyrinogen decarboxylase (Uro-d; EC 4.1.1.37), the fifth enzyme in the heme biosynthetic pathway, which catalyzes the sequential decarboxylation of uroporphyrinogen to coproporphyrinogen, is encoded by the HEM12 gene in Saccharomyces cerevisiae. The HEM12 gene is transcribed into a major short mRNA and a minor longer one, approximately 1.35 and 1.55 kb, respectively, in size, and that differ in the 5′ untranslated region. “Uroporphyric” mutants, which have no mutations in the HEM12 gene but accumulate uroporphyrinogen, a phenotype chracteristic of partial Uro-d deficiency, were investigated. Genetic analysis showed that the mutant phenotype depends on the combined action of two unlinked mutations, udt1 and either ipa1, ipa2, or ipa3. ipa1 is tightly linked to HEM12 The mutation udt1 apparently acts specifically on the HEM12 gene, and causes a six to tenfold decrease in the levels of the short HEM12 mRNA, in the β-galactosidase activity of a HEM12-lacZ fusion, in immunodetectable protein and enzyme activity. But heme synthesis is normal and porphyrin accumulation was modest. The mutations ipa1, ipa2, and ipa3 had no phenotype on their own, but they caused an increase in porphyrin accumulation in a udt1 background. This multiplicity of genetic factors leading to uroporphyric yeast cells closely resembles the situation in human porphyria cutanea tarda.     

Fluorescence study ofEscherichia coli cyclic AMP receptor protein

Time-resolved, steady-state fluorescence and fluorescence-detected circular dichroism (FDCD) have been used to resolve the fluorescence contributions of the two tryptophan residues, Trp-13 and Trp-85, in the cyclic AMP receptor protein (CRP). The iodide and acrylamide quenching data show that in CRP one tryptophan residue, Trp-85, is buried within the protein matrix and the other, Trp-13, is moderately exposed on the surface of the protein. Fluorescence-quenching-resolved spectra show that Trp-13 has emission at about 350 nm and contributes 76–83% to the total fluorescence emission. The Trp-85, unquenchable by iodide and acrylamide, has the fluorescence emission at about 337 nm. The time-resolved fluorescence measurements show that Trp-13 has a longer fluorescence decay time. The Trp-85 exhibits a shorter fluorescence decay time. In the CRP-cAMP complex the Trp-85, previously buried in the apoprotein becomes totally exposed to the iodide and acrylamide quenchers. The FDCD spectra indicate that in the CRP-cAMP complex Trp-85 remains in the same environment as in the protein alone. It has been proposed that the binding of cAMP to CRP is accompanied by a hinge reorientation of two protein domains. This allows for penetration of the quencher molecules into the Trp-85 residue previously buried in the protein matrix.     

Lead as an inductor of some morphological and functional changes in synaptosomes from rat brain

Summary 1. The effect of lead (in vivo) on the uptake of GABA, dopamine, and histidine as a precursor of histamine in synaptosomes obtained from chronically lead-treated rats was studied.2. Lead decreased the uptake of GABA, increased the uptake of dopamine, and did not change the uptake of histidine. These effects were independent of calcium concentration.3. Lead administration to the rat changed the morphology of the synaptosomes, as manifested in the decreased number of synaptic vesicles and disturbed mitochondrial structure.4. The results suggest the existence of several mechanisms of lead toxicity on uptake, related to individual neurotransmitters, which are not necessarily connected with a Pb²⁺/Ca²⁺ interaction.