Differences in response to the toxin sirodesmin PL produced by Phoma lingam (Tode ex fr.) Desm. on protoplasts,cell aggregates and intact plants of resistant and susceptible Brassica accessions

Summary The selective property of sirodesmin PL, a toxin produced by Phoma lingam, was studied on protoplasts, cell aggregates, leaves and roots. Directly after isolation, protoplasts from all the different Brassica accessions were sensitive when treated with toxin in a concentration higher than 1 M. When more differentiated plant tissue. i.e. cell aggregates, leaves or roots, were investigated, insensitivity to the toxin was found in the plant material resistant to P. lingam, while the plant material susceptible to P. lingam was sensitive. The results reveal that a clear correlation between resistance to P. lingam and insensitivity to sirodesmin PL is present, and that the toxin can be used to distinguish resistant plant material from susceptible both in vivo and in vitro.     

Incorporation of hygromycin resistance in Brassica nigra and its transfer to B. napus through asymmetric protoplast fusion

Summary With the idea to develop a selection system for asymmetric somatic hybrids between oilseed rape (Brassica napus) and black mustard (B. nigra), the marker gene hygromycin resistance was introduced in this last species by protoplast transformation with the disarmed Agrobacterium tumefaciens strain C58 pGV 3850 HPT. The B. nigra lines used for transformation had been previously selected for resistance to two important rape pathogens (Phoma lingam, Plasmodiophora brassicae). Asymmetric somatic hybrids were obtained through fusion of X-ray irradiated (mitotically inactivated) B. nigra protoplasts from transformed lines as donor with intact protoplasts of B. napus, using the hygromycin resistance as selection marker for fusion products. The somatic hybrids hitherto obtained expressed both hygromycin phosphotransferase and nopaline synthase genes. Previous experience with other plant species had demonstrated that besides the T-DNA, other genes of the donor genome can be co-transferred. In this way, the produced hybrids constitute a valuable material for studying the possibility to transfer agronomically relevant characters — in our case, diseases resistances — through asymmetric protoplast fusion.     

Growth and metabolic flexibility in groundwater bacteria

Groundwater bacteria isolated from an oligotrophic-saturated soil showed a mixed strategy of economized metabolism and migration when grown in a continuous-flow column system simulating poor or nutrient-amended growth conditions. The cells were generally <0.5m in diameter in pure groundwater, but doubled in size when the concentration of dissolved organic carbon and phosphate in groundwater was increased 10-fold. The biomass, estimated from analysis of muramic acid (MuAc) in cell wall peptidoglucans, increased at the same time by a factor of 5 when the solid support in the columns was gravel and by a factor of 10 when it was glass beads. Bacteria in pure groundwater stored 10 times more of the energy-rich polysaccharide, poly--hydroxybutyric acid (PHB), than bacteria in enriched groundwater, and those cells that were attached to the gravel stored 10 times as much as cells in the interstitial pore water. Once phosphate was added to groundwater, stored PHB was metabolized. The proportion of free-living to attached bacteria was 2 to 10 times higher in enriched compared with pure groundwater indicating a mass transport of cells as the carrying capacity of their habitat rose.     

Selective illumination of single photoreceptors in the house fly retina: local membrane turnover and uptake of extracellular horseradish peroxidase (HRP) and Lucifer Yellow

Summary Single photoreceptor cells in the compound eye of the housefly Musca domestica were selectively illuminated and subsequently compared electron-microscopically with the unilluminated photoreceptors in the immediate surroundings. The rhabdomeres of the illuminated cells remain largely unaffected, but the cells show an increase in the number of coated pits, various types of vesicles, and degradative organelles; some of the latter organelles are described for the first time in fly photoreceptors. Coated pits are found not only at the bases of the microvilli, but also in other parts of the plasma membrane. Degradative organelles, endoplasmic reticulum (ER) and mitochondria aggregate in the perinuclear region. The rough ER and smooth ER are more elaborate, the number of Golgi stacks, free ribosomes and polysomes is increased, and the shape and distribution of heterochromatin within the nuclei are altered. Illuminated photoreceptors also interdigitate extensively with their neighbouring secondary pigment cells. These structural changes in illuminated fly photoreceptor cells indicate an increase in membrane turnover and cellular metabolism. When applied to the eye, Lucifer Yellow spreads into the extracellular space and is taken up only by the illuminated photoreceptor cells. These cells show the same structural modifications as above. Horseradish peroxidase applied in the same way is observed in pinocytotic vesicles and degradative organelles of the illuminated cells. Hence, the light-induced uptake of extracellular compounds takes place in vivo at least partially as a result of an increase in pinocytosis.     

Analysis of the H+/sugar symport in yeast under conditions of depolarized plasma membrane

H⁺/sugar symport in the obligatory aerobic yeastRhodotorula glutinis was analyzed under conditions where the plasma membrane was selectively depolarized by the lipophilic cation tetraphenylphosphonium (TPP⁺). Control experiments showed that this treatment did not impair the transmembrane pH, the cell energy charge, and the function of plasma membrane H⁺-ATPase. The kinetic data were fitted to elementary functions derived from a model constructed on the basis of some simplifying premises for ordered (either C + H⁺ + S or C + S + H⁺) and random reaction mechanisms. In addition, the comparison of the kinetic parameters in fully energized and depolarized cells provided information about the free carrier charge. It was concluded that the binding sequence of formation of the ternary carrier/H⁺/substrate complex follows a random mechanism and that the carrier bears a negative charge.     

Effects of nucleotides and divalent cations on phospholipase activity in Saccharomyces cerevisiae

Divalent cations activate the lysophospholipase and transacylase reactions catalyzed by the same enzymes in the yeast Saccharomyces cerevisiae. The activation was observed at neutral pH, but not at the pH optimum of lysophospholipase/transacylase, near 3.5. Adenine nucleotides, especially AMP and ADP, are strong inhibitors of the same group of enzymes. Half maximal inhibition by AMP was found at a concentration of about 20 M. The inhibition by nucleotides in low concentrations is enhanced by divalent cations.     

Function of methanofuran,tetrahydromethanopterin, and coenzyme F420 in Archaeoglobus fulgidus

Archaeoglobus fulgidus is an extremely thermophilic archaebacterium that can grow at the expense of lactate oxidation with sulfate to CO₂ and H₂S. The organism contains coenzyme F₄₂₀, tetrahydromethanopterin, and methanofuran which are coenzymes previously thought to be unique for methanogenic bacteria. We report here that the bacterium contains methylenetetrahydromethanopterin: F₄₂₀ oxidoreductase (20 U/mg), methenyltetrahydromethanopterin cyclohydrolase (0.9 U/mg), formyltetrahydromethanopterin: methanofuran formyltransferase (4.4 U/mg), and formylmethanofuran: benzyl viologen oxidoreductase (35 mU/mg). Besides these enzymes carbon monoxide: methyl viologen oxidoreductase (5 U/mg), pyruvate: methyl viologen oxidoreductase (0.7 U/mg), and membranebound lactate: dimethylnaphthoquinone oxidoreductase (0.1 U/mg) were found. 2-Oxoglutarate dehydrogenase, which is a key enzyme of the citric acid cycle, was not detectable. From the enzyme outfit it is concluded that in A. fulgidus lactate is oxidized to CO₂ via a modified acetyl-CoA/carbon monoxide dehydrogenase pathway involving C₁-intermediates otherwise only used by methanogenic bacteria.Non-standard abbreviations APS adenosine 5-phosphosulfate - BV benzyl viologen - DCPIP 2,6-dichlorophenolindophenol - DMN 2,3-dimethyl-1,4-naphthoquinone - DTT DL-1,4-dithiothreitol - H₄F tetrahydrofolate - H₄MPT tetrahydromethanopterin - CH₂ H₄MPT, methylene-H₄MPT - CH H₄MPT, methenyl-H₄MPT - Mes morpholinoethane sulfonic acid - MFR methanofuran - Mops morpholinopropane sulfonic acid - MV methyl viologen - Tricine N-tris(hydroxymethyl)-methylglycine - U mol product formed per min