Site-selective 8-chloroadenosine 3',5'-cyclic monophosphate inhibits transformation and transforming growth factor alpha production in Ki-ras-transformed rat fibroblasts

A site-selective cAMP analog, 8-chloroadenosine 3',5'-cyclic monophosphate (8-Cl-cAMP), was demonstrated to be a potent inhibitor of both the monolayer and soft agar growth of normal rat kidney (NRK) fibroblasts that had been transformed with the v-Ki-ras oncogene or treated with transforming growth factor alpha (TGF alpha). The growth inhibition was dose dependent and reversible and was accompanied by reversion of the transformed phenotype, suppression of TGF alpha production, and a decrease in p21 ras protein levels. These effects of 8-Cl-cAMP were linked to the cAMP analog's selective modulation of the type I and type II cAMP-dependent protein kinase regulatory subunits, RI and RII, present in Ki-ras-transformed and TGF alpha-treated NRK cells.     

Herbicide effects on planktonic systems of different complexity

Bioassays of different complexity were compared with respect to their capability to predict the environmental impact of the herbicide atrazine in aquatic systems. Acute toxicity tests with Daphnia did not yield meaningful results. Sublethal tests with Daphnia (feeding inhibition, reduction of growth and reproduction) were more sensitive, but effective concentrations of atrazine were still rather high (2 mg/L). A relatively complicated artificial food chain system that incorporated direct and indirect effects on Daphnia yielded significant reduction of daphnid population growth at 0.1 mg/L. Enclosure experiments with natural communities were by far the most sensitive tools. Community responses could be measured at concentrations as low as 1 µg/L and 0.1 µg atrazine/L. At the lowest concentration, however, communities recovered after three weeks. We conclude that in complex systems indirect effects can be more important than direct effects, so that, contrary to the conditions in simple tests, non-target organisms may be the better indicators of herbicide stress to natural communities.     

Purification of acetyl-CoA: 17-O-deacetylvindoline 17-O-acetyltransferase from Catharanthus roseus leaves

The enzyme acetyl-CoA: 17-O-deacetylvindoline 17-O-acetyltransferase which terminates vindoline biosynthesis has been isolated from Catharanthus roseus leaves, further characterized and purified to homogeneity by three step column chromatography and subsequent preparative isoelectric focusing. Kinetic properties concerning the enzyme reaction are discussed. Five multiple forms of the acetyl-transferase could be observed, each consisting of two subunits. This enzyme is now the best characterized of the enzymes involved in vindoline biosynthesis.Abbreviations DTE dithiothreitol - EDTA ethylenediamine-tetraacetic acid - HEPES N-(2-hydroxyethyl)-piperazine-N-2-ethanesulfonic acid - IEF isoelectric focusing - KP_i potassium phosphate - M_r rel.molecular mass - PEG polyethylene glycol - SDS-PAGE sodium dodecylsulfate polyacrylamide gel electrophoresis - Tris 2-amino-2-(hydroxymethyl)-1,3-propandiol     

Study of plasmids isolated from saccharolytic Clostridia: Construction of hybrid plasmids and transfer toEscherichia coli andBacillus subtilis

Summary Small plasmids ofClostridium acetobutylicum and related strains were isolated and studied. Their restriction maps were established and different hybrid plasmids were constructed by ligation with plasmid pHV33.     

The quaternary structure of four crustacean two-hexameric hemocyanins: immunocorrelation,stoichiometry, reassembly and topology of individual subunits

Summary Two-hexameric (2×6) hemocyanins from the brachyuran crabsCancer pagurus andCallinectes sapidus, the freshwater crayfishAstacus leptodactylus and the lobsterHomarus americanus were isolated and dissociated into native subunits.The subunits of each hemocyanin were analyzed by electrophoresis and immunology. Three immunologically distinct subunit types, which were termed, and, could be identified in each case. They were isolated preparatively, and interspecifically correlated. Subunit is subdivided into several electrophoretically distinct isoforms which are immunologically closely related (Astacus) or identical (other species). InAstacus andCancer one of these isoforms was shown to dimerize and to act as inter-hexamer bridge. It represents a fourth subunit type termed. A fifth, diffuse component, which in PAGE migrated at the position of a dimer, was identified in the crossed immunoelectrophoretic patterns as denatured hemocyanin.A common feature of the four hemocyanins is the presence of 4 copies of and 8 copies of/ within the 2×6 particles. The: ratio is 4:4 in the two Astacidea and 6:2 in the two Brachyura. exists in 2 copies inAstacus andCancer which means that a single dimer- is present in a two-hexamer. This leaves 2 monomeric copies inAstacus and 4 inCancer.Every subunit from the four species except ofAstacus - was capable to form hexamers in reassembly experiments. If subunit combinations were tested, hetero-hexamers were formed preferentially. Two-hexamers were reconstituted only in the presence of all subunit types and the native subunit stoichiometry was required to obtain twohexamers in considerable yields. Factors limiting 2×6 reassembly are discussed.Authentic 2×6 molecules ofAstacus, Homarus andCancer hemocyanin were immunolabeled with subunit-specific antibody fragments (F_ab) or IgG molecules, and the resulting immuno complexes were studied in the electron microscope. A topological model of the quaternary structure of decapod 2×6 hemocyanins is derived, showing the position of each copy of the four subunit types. In this model, the inter-hexamer bridge- is surrounded by two and two subunits forming the central core of the dodecamer. Two additional and two additional subunits form the periphery together with one subunit occupying the peripheral short edges of each hexameric half structure. The model is discussed with respect to the current literature.Abbreviations PAGE polyacrylamide gel electrophoresis - SDS sodium dodecyl sulfate This paper is decicated to Professor Dr. Bernt LinzenPreliminary accounts of this work have been presented in the proceedings of a symposium at Tutzing 1985. Linzen B (ed) (1986) Invertebrate oxygen carriers. Springer, Berlin Heidelberg New York. This also includes: Stöcker et al. 1986; Markl et al. 1986) and in a review article (Markl 1986)     

Unusual evolutionary conservation of 5S rRNA pseudogenes inAspergillus nidulans: Similarity of the DNA sequence associated with the pseudogenes with the mouse immunoglobulin switch region

Summary AllAspergillus nidulans 5S rRNA pseudogenes known so far are the result of integration of an approx. 0.2-kbp-long DNA sequence into the 5S rRNA genes. This sequence, called block C, is present in at least five copies in theA. nidulans genome and seems to be associated either with 5S rRNA genes or pseudogenes. In contrast to the 78% sequence conservation of the C-block in pseudogenes, the truncated 5 halves of the pseudogenes are very highly conserved (96.9–100%). We postulate that the 5S rRNA pseudogenes are still a subject of concerted evolution. The C-block sequence shows similarity to the switch region of the mouse heavy chain immunoglobulin gene. A characteristic motif GGGTGAG is repeated several times in both sequences; the sequence conservation is 63%.     

Diminished oestrogen sensitivity and increased ovulation rate in adult female rats after prepubertal treatment with oestrogen or pimozide

Immature female rats were implanted with oestradiol benzoate or cholesterol in the medial preoptic area at different ages, and the inhibition of the ovariectomy-induced increase of LH secretion by s.c. injected oestradiol was investigated. Medial preoptic oestrogen implants reduced the inhibition of LH secretion in 4-week-old rats, but not in younger animals. Elevation of the circulating oestrogen concentration or suppression of the central nervous dopamine activity by daily injections of oestradiol and pimozide, respectively, from Day 26 to the day of vaginal opening, i.e. during the time when the mechanism of the oestrogen-induced desensitization of the negative oestrogen feedback matures, resulted in considerable diminution of the LH-inhibiting effect of oestradiol in ovariectomized adult females. In intact cyclic rats, both prepubertal treatments led to a significant increase of the average number of eggs per ovulation that was mainly caused by reduction of the number of animals with a low ovulation rate.     

Kainic acid on the rostral ventrolateral medulla inhibits phrenic output and CO2 sensitivity

We used the neurotoxin, kainic acid, which is known to stimulate neuronal cell bodies as opposed to axons of passage by binding to specific amino acid receptors to determine whether cells with such receptors have access to the ventrolateral medullary surface and are involved in central ventilatory chemosensitivity. Pledgets with 4.7 mM kainic acid were placed bilaterally on the rostral, intermediate, or caudal ventilatory chemosensitive areas for 1-2 min in chloralose-urethan-anesthetized, paralyzed, vagotomized, glomectomized, and servo-ventilated cats. Application of kainic acid on the caudal or intermediate areas produced no consistent significant effects on eucapnic phrenic output or on the slope or maximum value of the phrenic nerve response to increased end-tidal PCO2. Rostral area kainic acid produced immediate augmentation and then diminution of blood pressure and phrenic output. Apnea developed in six of nine cats by 40 min. In all five cats in which it could be tested, the slope of the CO2 response was clearly decreased. Of [3H]kainic acid applied to the rostral area, 88.4% was shown to be within 2 mm of the ventral surface. Comparison of surface application sites of this and other studies suggests that an area overlapping the border of the original rostral and intermediate areas allows access to neurons involved in the chemoreception process, which may also provide tonic facilitatory input to cardiorespiratory systems.     

Die Ölblumensymbiosen – Parallelismus and andere Aspekte ihrer Entwicklung in Raum and Zeit1, 2

The oil-bee/oil-flower relationships: parallelism and other aspects of their evolution in space and time A survey is given of our present knowledge and existing hypotheses concerning the biogeography, history, and phylogeny of plant taxa yielding fatty oil as a floral reward, and of the bee genera involved in their pollination. Four syngenetic complexes of the symbiosis arose convergently: The neotropical, the paleotropical, the holarctic, and the capensic complex. On the basis of the mutual structural adaptations of bees and flowers it is concluded that, in addition, parallelism within related groups as a result of a common tendency to develop the respective organs, has played an important role in the evolution of the oil-based floral interrelationships.     

Studies on haemosiderin and ferritin from iron-loaded rat liver

Summary Haemosiderin has been isolated from siderosomes and ferritin from the cytosol of livers of rats iron-loaded by intraperitoneal injections of iron-dextran. Siderosomal haermosiderin, like ferritin, was shown by electron diffraction to contain iron mainly in the form of small particles of ferrihydrite (5Fe₂O₃ · 9H₂O), with average particle diameter of 5.36±1.31 nm (SD), less than that of ferritin iron-cores (6.14±1.18 nm). Mössbauer spectra of both iron-storage complexes are also similar, except that the blocking temperature,T _B, for haemosiderin (23 K) is lower than that of ferritin (35 K). These values are consistent with their differences in particle volumes assuming identical magnetic anisotropy constants. Measurements of P/Fe ratios by electron probe microanalysis showed the presence of phosphorus in rat liver haemosiderin, but much of it was lost on extensive dialysis. The presence of peptides reacting with anti-ferritin antisera and the similarities in the structures of their iron components are consistent with the view that rat liver haemosiderin arises by degradation of ferritin polypeptides, but its peptide pattern is different from that found in human-thalassaemia haemosiderin. The blocking temperature, 35 K, for rat liver ferritin is near to that reported, 40 K, for human-thalassaemia spleen ferritin. However, the haemosiderin isolated from this tissue, in contrast to that from rat liver, had aT _B higher than that of ferritin. The iron availability of haemosiderins from rat liver and human-thalassaemic spleen to a hydroxypyridinone chelator also differed. That from rat liver was equal to or greater, and that from human spleen was markedly less, than the iron availability from either of the associated ferritins, which were equivalent. The differences in properties of the two types of haemosiderin may reflect their origins from primary or secondary iron overload and differences in the duration of the overload.