Untersuchungen über stabile und während der frühen Quellungsstadien neu synthetisierte Poly(A)-RNA vonAgrostemma-Samen

Summary RNA isolated from dry embryos ofAgrostemma githago seeds contains poly(A)-sequences, but in very small amounts. In the early phase of imbibition, an intensive synthesis of poly(A)-containing RNA is brought about. The importance of this synthesis of poly(A)-RNA is discussed.

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Herrn Dr.E. Serfling (Gatersleben, DDR) danke ich für methodische Hinweise und für die Überlassung einiger Chemikalien.     

Poly(A) associated RNA from mitochondria and microsomes of rat brain

Summary Rat brain mitochondria contain a significant proportion of poly(A) associated RNA which is higher than that found in microsomes from the same source. When steady state poly(A) RNA of brain mitochondria was analyzed by microelectrophoresis, it displayed a characteristic separation pattern with a large amount of free poly(A).     

Compared expression of murine plasmacytoma associated virus in tumor cells and in mouse embryo cells]

A DNA complementary to the viral genome of C-type particles produced by a Mouse myeloma derived cell line (MF2 cell line) was synthesized. This cDNA was used as a probe to study the viral genome expression among the total RNA and the poly (A)-rich RNA extracted from the MF2 and Balb/c embryonic cells. As evidenced by molecular hybridization experiments, the presence of at least one endogenous Balb/c virus in the MF2 virus stocks is suggested. In the productive cells, the viral RNA sequences are expressed in the poly (A)-rich RNA fraction.     

Satellite RNA (RNA3) of tomato black ring virus is found with one of the 2 major RNAs (RNA2) in a new capsid nucleoprotein]

Tomato Black Ring Virus (TBRV) like other NEPOviruses posseses two nucleoproteins M and B and two major RNAs, RNA1 and RNA2 respectively distributed in B and M. A new nucleoprotein has just been discovered and comprises one molecule of RNA2 associated with one molecule of RNA3. RNA3 is a small RNA of molecular weight 500,000 d considered to be a satellite RNA. Its level appears to depend on the infection stage, local or systemic. RNA3 is able to modify the relative proportions of nucleoproteins M and B and their respective RNAs. The satellite RNA, might be part of the genome and represent a monocistronic mRNA for protein capsid synthesis. However it seems perhaps more tempting to correlate TBRV-RNA3 with satellite RNA5 of certain strains of Cucumber mosaic virus.     

Purfication and molecular weight of an RNA-dependant RNA polymerase from Brassicae oleracea var. Botrytis]

An RNA-dependent RNA polymerase has been completely purified from Cauliflower inflorescences. Analysis of the purified enzyme on SDS-polyacrylamide gels showed one polypeptide chain with a molecular weight of 140,000 dalton. The enzyme is monomeric in its native state. The in vitro activity was completely dependent on added RNA, the most efficient templates being poly (U), poly (U, C), poly (I) and viral RNA.     

Autoradiographic studies on RNA synthesis and transport in the salivary gland of Lygaeus sp. (Hemiptera-Lygaeidae)

Autoradiographic studies using 3H-uridine in the salivary gland of adult Lygaeus sp. were carried out. The gland cell nuclei, particularly the multiple nucleoli, are the sites of incorporation of the label exhibiting RNA synthesis. The labelled molecules (RNA) are transported to the cell cytoplasm and then into the gland lumen in which no turnover of the radioactivity is observed.     

Experimentelle Untersuchungen zum Wirkungsmechanismus der beiden entwicklungsphysiologisch aktiven Fraktionen des «Gehirnhormons» der Insekten (Aktivationsfaktor I und II) auf die Prothoracaldrüse

Summary 2 prothoracotropical factors (activation factor I and II) have been obtained by gel filtration techniques from brains and corpora cardiaca of the cockroachPeriplaneta americana. In contrast to activation factor II, activation factor I caused significant influence of RNA synthesis. The RNA pattern of prothoracic glands stimulated by activation factor I as demonstrated by polyacrylamide gel electrophoresis consists of different kinds of RNA. Short time incubation revealed effects on sRNA synthesis, while long time incubation demonstrated predominantly increase of ribosomal RNA synthesis. Measurements of the membrane potential of the prothoracic gland cells of the wax mothGalleria mellonella indicated an increase by activation factor II; activation factor I was without any visible effect. Our results demonstrate for the first time that the two activation factors induce different effects at cellular level.

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Für technische Unterstützung danken wir FräuleinA. Zinsser und FrauR. Meissner.

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Durchgeführt mit Mitteln des Ministeriums für Wissenschaft und Technik und mit Unterstützung durch die Sächsische Akademie der Wissenschaften zu Leipzig.     

Brain factor from Galleria mellonella (Lepidoptera) stimulating silk gland activity.

Brain extracts from day 1-4 last instar larvae of Galleria mellonella (Lepidoptera) stimulate RNA synthesis in cultured silk glands from day 3 last instar larvae. When the fibroin-synthesizing posterior parts of silk glands were incubated for 3 h in vitro in the presence of brain extract (0.1 brain equivalent), [3H]-uridine incorporation into RNA was stimulated more than twofold. The stimulating effect of brain extract showed a dose response relationship. It is suggested that the heat-resistant and protease-sensitive brain factor is a peptide.     

Contrasting effects of RNA and protein synthesis blocking on natural and lectin-dependent cell-mediated cytotoxicity against adherent HEp-2 cells

In this study, earlier observations concerning the independence of both natural (NCMC) and lectin-dependent cell-mediated cytotoxicity (LDCC) from DNA synthesis have been confirmed. In addition, blocking of RNA synthesis by actinomycin D and of protein synthesis, reversibly by puromycin (PM) and irreversibly by emetine (EM) had different effects on NCMC and LDCC against 3H-thymidine-prelabeled HEp-2 target cells. Similarly to the Con A-induced proliferation of lymphocytes, LDCC activity was also inhibited by blocking of RNA and protein synthesis. NCMC to HEp-2 target cells was not affected by blocking of RNA synthesis, while both PM and EM strongly enhanced NCMC activity.     

Brain factor fromGalleria mellonella (Lepidoptera) stimulating silk gland activity

Brain extracts from day 1–4 last instar larvae ofGalleria mellonella (Lepidoptera) stimulate RNA synthesis in cultured silk glands from day 3 last instar larvae. When the fibroin-synthesizing posterior parts of silk glands were incubated for 3 h in vitro in the presence of brain extract (0.1 brain equivalent), [³H]-uridine incorporation into RNA was stimulated more than twofold. The stimulating effect of brain extract showed a dose response relationship. It is suggested that the heat-resistant and protease-sensitive brain factor is a peptide.     

Contrasting effects of RNA and protein synthesis blocking on natural and lectin-dependent cell-mediated cytotoxicity against adherent HEp-2 cells

Summary In this study, earlier observations^2,9 concerning the independence of both natural (NCMC) and lectin-dependent cell-mediated cytotoxicity (LDCC) from DNA synthesis have been confirmed. In addition, blocking of RNA synthesis by actinomycin D and of protein synthesis, reversibly by puromycin (PM) and irreversibly by emetine (EM) had different effects on NCMC and LDCC against³H-thymidine-prelabeled HEp-2 target cells. Similarly to the Con A-induced proliferation of lymphocytes, LDCC activity was also inhibited by blocking of RNA and protein synthesis. NCMC to HEp-2 target cells was not affected by blocking of RNA synthesis, while both PM and EM strongly enhanced NCMC activity.     

Poly-ADP-ribosylation in health and disease

Poly(ADP-ribosyl)ation is required by multicellular eukaryotes to ensure genomic integrity under conditions of mild to moderate genotoxic stress. However, severe stress following acute neuronal injury causes overactivation of poly(ADP-ribose) polymerase-1, which results in unregulated poly(ADP-ribose) (PAR) synthesis and widespread neuronal cell death. Once thought to be a necrotic cell death resulting from energy failure, PARP-1 activation is now known to induce the nuclear translocation of apoptosis-inducing factor, which results in caspase-independent cell death. Conversely, poly(ADP-ribose) glycohydrolase, once thought to contribute to neuronal injury, now appears to have a protective role as demonstrated by recent studies utilizing gene disruption technology. Thus, the emerging mechanism dictating the fate of neurons appears to involve the regulation of PAR levels in neurons. Therefore, therapies targeting poly(ADP-ribosyl)ation in the treatment of neurodegenerative conditions such as stroke and Parkinson's disease are required to inhibit PAR synthesis and/or facilitate its degradation.     

The roles of poly(ADP-ribose)-metabolizing enzymes in alkylation-induced cell death

Poly(ADP-ribose) (PAR) has been identified as a DNA damage-inducible cell death signal upstream of apoptosis-inducing factor (AIF). PAR causes the translocation of AIF from mitochondria to the nucleus and triggers cell death. In living cells, PAR molecules are subject to dynamic changes pending on internal and external stress factors. Using RNA interference (RNAi), we determined the roles of poly(ADP-ribose) polymerases-1 and -2 (PARP-1, PARP-2) and poly(ADP-ribose) glycohydrolase (PARG), the key enzymes configuring PAR molecules, in cell death induced by an alkylating agent. We found that PARP-1, but not PARP-2 and PARG, contributed to alkylation-induced cell death. Likewise, AIF translocation was only affected by PARP-1. PARP-1 seems to play a major role configuring PAR as a death signal involving AIF translocation regardless of the death pathway involved. Received 7 November 2007; received after revision 19 December 2007; accepted 21 December 2007 O. Cohausz, C. Blenn: These two authors contributed equally to this work.     

Autoradiographic localization of RNA synthesis in vitro during oogenesis in Parascaris equorum]

Technical elaboration of in vitro incubation of Parascaris equorum gonads with 3H-Uridine has permitted, for the first time, the study of RNA synthesis during oogenesis along the whole gonadic tube. In germ cells, oocytes in diakinesis (oviduct) and in division of maturation (uterus) show no label. On the contrary oogonia and growing oocytes in ovary are labelled. RNA synthesis is always detected in all parietal cells but is more active in oviduct and uterus where the gonadic wall is particularly developed.     

Die Veränderung des Stärke-, Protein- und RNS-Gehaltes vonLemna minor L. unter dem Einfluss von Kinetin (6-Furfurylaminopurin)

Summary Kinetin (6-furfurylaminopurine) is rapidly absorbed byLemna minor L. 3 · 10^–8 M/ml medium cause an immediate but temporary stimulation of RNA and protein synthesis during the first hour of the treatment. The following excessive accumulation of starch is considered to be more or less a direct consequence of a disturbed RNA and protein metabolism.     

In vitro translation of human placental pregnancy-specific beta1-glycoprotein (SP1)

Summary Synthesis of SP₁-glycoprotein by the human placenta was directly demonstrated, by in vitro translation of RNA extracted from full term and from early placentas in a cell-free wheat germ system followed by specific immunoprecipitation of the radioactively labeled nascent peptides. De novo synthesized SP₁-glycoprotein in both RNA preparations accounted for 1–1.3% of total protein synthesis.Acknowledgment. The authors thank Drs A. Nirapatpongporn, V. Sirivasin and Professor H.F. Lodish for kindly providing placental tissues and wheat germ respectively. This work was supported by The Rockefeller Foundation (RF-8031).W.H. was supported by a Graduate Studies Fellowship, Mahidol University.     

High salt effect on RNA synthesis in Krebs ascites tumor cells

The incubation of Krebs ascites tumor cells in medium with a high salt concentration resulted in a partial inhibition of nuclear RNA synthesis. The residual RNA polymerase activity in such nuclei was only slightly inhibited by low concentrations (50 nM) of alpha-amanitin. This finding suggested an inhibition of RNA polymerase II activity under conditions of medium hypertonicity. Indeed, RNA polymerase II, isolated from the nuclei of cells exposed to hypertonicity, revealed only half of the control activity. On the other hand, RNA polymerase I was not affected by hypertonicity. Moreover, chromatin fractions isolated from cells incubated in hypertonic or isotonic medium were equally template-active in RNA synthesis.