RNA-Dependent DNA Polymerase Activity of RNA Tumor Viruses I. Directing Influence of DNA in the Reaction

The template requirements and deoxyribonucleic acid (DNA) products of the DNA polymerases isolated from Rauscher leukemia and avian myeloblastosis viruses have been examined. All DNA preparations or synthetic polydeoxynucleotides which are active as primers possess a duplex structure containing single-stranded regions with a 3'-hydroxyl terminus. Native DNA and fully single-stranded DNA are inactive; moreover, their activity is not enhanced by sonic oscillation or treatment with micrococcal nuclease, Neurospora nuclease, or low levels of deoxyribonuclease I. Poor DNA templates are activated by treatment with exonuclease III, large amounts of deoxyribonuclease I, or an endonuclease isolated from Rauscher viral preparations. In reactions primed with deoxyadenylate-deoxythymidylate copolymer, the product formed is covalently attached to primer strands, indicating that no new strands are initiated. DNA polymerase products formed with exonuclease III- or deoxyribonuclase I-treated DNA are duplex structures. Short single-stranded regions are completely filled in, whereas long single-stranded regions are only partly repaired. DNA preparations containing extensive single-stranded regions are poorly utilized as templates.     

Bestimmung von deoxynivalenol mittels LC - MS/MS (Tandem Massenspektrometrie)

A sensitive and selective method using high-performance liquid chromatography in combination with atmospheric pressure chemical ionization tandem mass spetrometry (LC-APCI-MS/MS) has been developed for the determination of Deoxynivalenol (DON) in trace levels. The extract was purified with a MultiSep? column followed by the Vicam? DON immunoaffinity column. Quantification is based on an external standard method using positive Multiple Reaction Monitoring (MRM). The limit of detection was 5 μg/kg with a signal to noise ratio of 3:1.     

Replenishment of fish populations in the enclosed lagoon of Taiaro Atoll: (Tuamotu Archipelago, French Polynesia) evidence from eggs and larvae

 Taiaro Atoll Lagoon is normally isolated from the ocean, but at least 125 marine fish species of 31 families are present there. We sampled fish larvae in Taiaro Lagoon and the nearby ocean in February 1994 with plankton net, neuston net and light trap to investigate which taxa were completing their life cycles in the lagoon. Concentrations of fish eggs and larvae were very high in the lagoon indicating intense spawning, but larvae of only 18 taxa of 10 families were present. Only six, a callionymid, gobiids, a hemiramphid, a microdesmid, and two pomacentrids, were present across a full range of pelagic sizes, and were clearly completing their pelagic stage in the lagoon. Four other taxa, an apogonid, two labrids and a scarid, were common, but the largest individuals were small (<5 mm) postflexion larvae. These may have been completing their pelagic stage in the lagoon. The remaining lagoonal larvae (eight taxa) were rare and at the preflexion stage, so we could only conclude that they hatched from eggs spawned in the lagoon. Nineteen taxa of 15 families found as adults in the lagoon were present outside the lagoon as larvae, but not inside, suggesting that they may not normally complete their life cycles in the lagoon. Horizontal distributions of larvae in the lagoon are apparently due to the interaction of larval vertical distribution behaviour with a wind-driven countercurrent system. Accepted: 16 October 1996     

Experimental considerations on the measurement of prostaglandins during long-time incubations of neonatal mouse calvaria

The influence of experimental conditions during long-time (72 h) incubations of neonatal mouse calvaria on the measurement of prostaglandins was investigated. Incubations of the cultured calvaria were carried out in the presence and absence of stimulating agents of bone resorption, such as thrombin and parathyroid hormone. It was found that during the first 24 h prostaglandin levels, estimated by gas chromatography negative ion chemical ionization mass spectrometry, did not correlate with calcium liberation, but were merely an artefact resulting from surgery by preparing the calvaria.     

Site-specific phosphorylation of avian retrovirus nucleocapsid protein pp12 regulates binding to viral RNA. Evidence for different protein conformations

Phosphorylation of serine 40 of the major nucleocapsid protein of avian retroviruses, pp12, regulates binding to viral RNA (Leis, J., Johnson, S., Collins, L. S., and Traugh, J. A. (1984) J. Biol. Chem. 259, 7726-7732). The phosphorylation state of the protein can be altered in vitro, resulting in the interconversion of the protein between a state of high affinity for single-stranded RNA and low affinity for single- or double-stranded RNA. The reversible phosphorylation of serine 40 is accompanied by a change in the conformation of the protein as demonstrated by quenching of intrinsic tryptophan fluorescence and chemical modification studies. Quenching of fluorescence of the sole tryptophan residue, Trp 80, by poly(U), KI, and CsCl indicates that the microenvironment of this residue is more positive in pp12 than in p12. Chemical modification studies indicate that the 3 lysine residues at positions 36, 37, and 39 of pp12 react with 2,4,6-trinitrobenzenesulfonic acid, while only 1 of these residues reacts in p12. The addition of single-stranded, but not double-stranded RNA, to pp12 protects 2 of the 3 lysine residues from chemical modification, suggesting that the two protected lysyl groups are required for binding to single-stranded viral RNA. In contrast to the phosphorylation of serine 40, phosphorylation of serine 43, catalyzed by protease-activated kinase II in vitro, does not induce changes in the protein conformation nor does it alter the RNA binding properties of the protein.