Beobachtungen anLocustella luscinioides im Kremmer Luch

Ohne Zusammenfassung     

High Irradiance Blue Light Affects Cortical Microtubules in the Green Alga Mougeotia scalarisi

High irradiance blue light (HIBL), but not red light, diminishedanti-tubulin indirect immunofluorescence of cortical microtubulesin the green alga Mougeotia. Quantitation of MT-specific immunofluorescenceby digital image analysis revealed taxol to counteract the HIBL-eliciteddecrease in immunolabelling. Taxol caused no significant increaseof immunofluorescence in red light. ¹Dedicated to Professor Horst Senger on the occasion of his65th birthday     

Die Wirkung von Neomycin auf Zellteilung,Wachstum und Nucleinsäuren synchronisierter Chlorellen

Zusammenfassung Autotrophe und mixotrophe Synchronkulturen von Chlorella pyrenoidosa zeigen in Lösungen, die mehr als 0,5 mg/ml an Neomycin enthalten, am Ende eines Zellcyclus keine Zellteilung. Die Produktion von Trockensubstanz und Zellstickstoff ist dabei nur geringfügig verringert. Hierin lassen sich ebenfalls keine Unterschiede zwischen autotrophen und mixotrophen Anzuchten feststellen.Nach 24 Std Behandlung mit 1 mg/ml an Neomycin ist der DNA-Gehalt um 20–30%, der RNA-Gehalt um 16% niedriger als bei der Kontrolle. Durch Chromatographie kann keine Nucleinsäure-Fraktion festgestellt werden, die durch die Neomycin-Behandlung besonders betroffen wäre.Bei Auftrennung der Ribosomen aus behandelten Zellen ergibt sich ein gegenüber der Kontrolle deutlich erhöhter Gehalt an Polysomen der cytoplasmatischen Ribosomen.Der Wirkungsort des Neomycins auf die Teilungshemmung wird diskutiert.

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Effect of neomycin on cell division, growth and nucleic acids in synchronized Chlorella cells

Summary Synchronously grown Chlorella cells show no more cell division after autotrophic and mixotrophic culture in the presence of 0.5 mg/ml neomycin. At the same time, the production of dry matter and of cell nitrogen is nearly unaffected by the drug.After colorimetric determination, the DNA-content of the cells can be shown to be 20 to 30 per cent, the RNA-content to be 16 per cent lower than in the untreated control cells. The chromatography of the nucleic acid preparations from neomycin-treated and untreated cells on MAK-columns shows no nucleic acid fraction which is specially affected by the antibiotic treatment.The preparation of ribosomes from treated and untreated cells and their separation in sucrose density gradients results patterns which show an increase of polysomes after neomycin-treatment of the cells.The results are discussed with respect to the possible site of action of neomycin in the cell division process.

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Z. Z. Centre de Biochimie et de Biologie Moléculaire, C.N.R.S., F-13 Marseille, Frankreich.     

Einbau von Uridin und Orotsäure in plastidäre ribosomale RNA von Chlorella nach Antibiotica-Behandlung

Zusammenfassung Chlorella pyrenoidosa inkorporiert unter normalen Anzuchtbedingungen kurzfristig angebotenes Uridin fast ausschließlich in plastidäre ribosomale RNA. Es lassen sich rasch markierte Ribosomen und deren Untereinheiten von 70 S, 50 S und 30 S nachweisen. Diese Markierung wird durch Rifampin in geringen Konzentrationen bereits nach wenigen Minuten unterbunden. Auf das Zellwachstum hat Rifampin bei heterotropher Anzucht dagegen auch in höheren Konzentrationen keinen Einfluß. Chloramphenicol hemmt den kurzfristigen Uridin-Einbau in ribosomale Partikeln von 70 S, 50 S und 30 S, nur geringfügig dagegen denjenigen in ribosomale RNA. Auch die Wirkung des Chloramphenicols tritt rasch ein. Cycloheximid beeinflußt den Kurzzeit-Einbau von Uridin in ribosomale Partikeln und in RNA nicht, wenn die Inkubationszeit 60 min nicht überschreitet.Die Markierung der Nucleinsäuren von Chlorella mit 6-(14C)-Orotsäure zeigt vergleichbare Empfindlichkeiten gegen die drei Antibiotica wie der Einbau von 6-(14C)-Uridin und 5-(3H)-Uridin.

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Incorporation of uridine and of orotate into chloroplast ribosome RNA of Chlorella after treatment with antibiotics

Summary Normal grown cells of Chlorella pyrenoidosa incorporate uridine exclusively into chloroplast ribosomal RNA after short time labeling. With sucrose gradient separation, labeled ribosomal particles of 70 S, 50 S and 30 S can be shown. This labeling is prevented by rifampin in low concentrations after a few minutes. At the same concentration of the antibiotic and also with 10-fold higher concentration, no effect on heterotrophic cell growth is observed. This indicates clearly that mitochondria cannot be influenced by rifampin. Chloramphenicol also inhibits the formation of uridine labeled ribosomal particles of 70 S, 50 S and 30 S. In the presence of this antibiotic, some labeled ribosomal RNA is formed. Also the effect of chloramphenicol can be shown after short incubation periods. Cycloheximide treatment of the cells within 30 and 60 min and up to the 10-fold concentration of protein synthesis inhibition (Morris, 1967) results in no effect on labeling of ribosomal RNA and of ribosomal particles in Chlorella with uridine. Only after prolonged treatment of the cells with cycloheximide is some effect on uridine incorporation observed.The comparison of the incorporation patterns of 6-(14C)-orotate, (6-14C)-uridine and 5-(3H)-uridine into nucleic acids in the presence of rifampin, chloramphenicol and cycloheximide shows some similarities. After 60 min incubation with the precursors, the incorporation is reduced by all three antibiotics. In rifampin treated cells, orotate and both uridines are preferentially incorporated into DNA. With chloramphenicol, the relative incorporation of orotate and of uridine into the 5 S and the 16 S RNA is higher as compared with the 23 S RNA. Cycloheximide results in an increase in the relative incorporation of orotate as well of uridine into DNA. The similarities of the effects of the three antibotics indicate that the preferential incorporation of uridine into chloroplast ribosomes of Chlorella is not due to a compartmentation of the uridine-UMP-pathway.

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Abkürzungen BisMSB bis(O-Methylstyryl)-Benzol - PPO 2,5-Diphenyloxazol - MAK-Säule Säule aus methyliertem Albumin mit Kieselgur     

Loss of the orphan nuclear receptor NR2F6 enhances CD8+ T-cell memory via IFN-γ

Memory formation is a hallmark of T cell-mediated immunity, but how differentiation into either short-lived effector cells (SLECs, CD127⁻KLRG1⁺) or memory precursors cells (MPECs, CD127⁺KLRG1⁻) and subsequent regulation of long-term memory is adjusted is incompletely understood. Here, we show that loss of the nuclear orphan receptor NR2F6 in germ-line Nr2f6-deficient mice enhances antigen-specific CD8⁺ memory formation up to 70 days after bacterial infection with Listeria monocytogenes (LmOVA) and boosts inflammatory IFN-γ, TNFα, and IL-2 cytokine recall responses. Adoptive transfer experiments using Nr2f6^−/− OT-I T-cells showed that the augmented memory formation is CD8⁺ T-cell intrinsic. Although the relative difference between the Nr2f6^+/+ and Nr2f6^−/− OT-I memory compartment declines over time, Nr2f6-deficient OT-I memory T cells mount significantly enhanced IFN-γ responses upon reinfection with increased clonal expansion and improved host antigen-specific CD8⁺ T-cell responses. Following a secondary adoptive transfer into naïve congenic mice, Nr2f6-deficient OT-I memory T cells are superior in clearing LmOVA infection. Finally, we show that the commitment to enhanced memory within Nr2f6-deficient OT-I T cells is established in the early phases of the antibacterial immune response and is IFN-γ mediated. IFN-γ blocking normalized MPEC formation of Nr2f6-deficient OT-I T cells. Thus, deletion or pharmacological inhibition of NR2F6 in antigen-specific CD8⁺ T cells may have therapeutic potential for enhancing early IFN-γ production and consequently the functionality of memory CD8⁺ T cells in vivo.Subject terms: Interferons, Bacterial infection     

Investigation of the in vitro biotransformation and simultaneous enantioselective separation of thalidomide and its neutral metabolites by capillary electrophoresis

Reversed-phase liquid chromatography is a long established method for the analysis of drug metabolism. The current investigation demonstrates that micellar electrokinetic capillary chromatography can be an attractive alternative. Two methods were developed using sodium dodecyl sulfate and hexadecyltrimethylammonium bromide for the determination of possible hydroxylated metabolites of the former sedative drug thalidomide (Contergan) in order to study the in vitro metabolism of the drug by incubation with rat liver microsomes. The biotransformation was found to be stereoselective: S-(−)-thalidomide mainly formed 5-hydroxythalidomide, whereas R-(+)-thalidomide was preferentially transformed to two metabolites tentatively assigned to be diastereomers of 5′-hydroxythalidomide.Furthermore, the simultaneous enantioseparation of thalidomide and two of its possible hydroxylated metabolites was achieved using capillary electrophoresis with negatively charged carboxymethyl-β-cyclodextrin. The dependencies of the selectivity of the enantioseparation on the concentration of the chiral additive and the pH of the run buffer were investigated.