The antiglucocorticoid, cortexolone, fails to promote in vitro activation of cytoplasmic glucocorticoid receptors from the human leukemic cell line CEM-C7

Cortexolone functions as an antiglucocorticoid in the human leukemic cell line CEM-C7, since it blocks the growth inhibition and cell lysis mediated by the potent agonist triamcinolone acetonide (TA). At high concentrations (10(-5) M) cortexolone alone is inactive. The ability of cortexolone to block the TA-mediated biological effects is reflected in its ability (1000-fold molar excess) to effectively block the binding of [3H]TA to the cytoplasmic unactivated form of the receptors eluted from DEAE-cellulose at approx. 180 mM potassium phosphate (KP). Likewise a 1000-fold molar excess of TA inhibits the specific binding of [3H]cortexolone to the unactivated receptors and to a peak which elutes at low salt concentration (35 mM KP) but does not appear to represent activated [3H]cortexolone-receptor complexes. Thermal activation/transformation (25 degrees C for 30 min +/- 10 mM ATP) of the [3H]TA-receptor complexes significantly enhances the subsequent DNA-cellulose binding capacity of these complexes and also results in their elution from DEAE-cellulose at the low salt (50 mM KP) activated position. In contrast, exposure of the cytoplasmic [3H]cortexolone-receptor complexes to identical in vitro activating (transforming) conditions fails to enhance subsequent DNA-cellulose binding capacity or to result in the appropriate shift in DEAE-cellulose elution profile. This inability of [3H]cortexolone to facilitate activation/transformation of receptors was also verified using cytosol prepared from the glucocorticoid-resistant 'activation-labile' mutant, 3R7. Taken collectively the data suggest that cortexolone, unlike an agonist such as TA, fails to promote in vitro activation/transformation, a conformational change which also occurs in vivo under physiological conditions and is a prerequisite for nuclear binding.     

Uptake and metabolism of taurine in the green alga Chlorella fusca

Taurine entered the alga Chlorella fusca Shihira et Krauss strain 21l-8b via a pH and energy-dependent system ("permease"). Transport followed triphasic kinetics from 10⁻⁶ to 10⁻² M with K_m values for taurine of 5.4 × 10⁻⁵, 4.1 × l0⁻⁴ and l.5 × 10⁻³ M. This uptake system was specific for sulfonic acids and showed no affinity for α- and β -amino acids or Na⁺; thus the permease of C. fusca is different from all known taurine transport systems with respect to structural specificity and lack of Na⁺ -dependence. Uptake was not observed in sulfate-grown algae but developed as a response to sulfate limitation within 2 h. Sulfate addition caused a rapid decline in taurine transport capacity. Labeled taurine was rapidly metabolized in C. fusca to sulfate and ethanolamine, suggesting oxidative hydrolysis as the mechanism of C-S bond cleavage. Further incorporation of these catabolic products in C - and S -metabolism was demonstrated. Taurine catabolism was also detected in other green algae and some cyanobacteria.     

Different Rates of Synthesis and Degradation of Two Chloroplastic Ammonium-Inducible NADP-Specific Glutamate Dehydrogenase Isoenzymes during Induction and Deinduction in Chlorella sorokiniana Cells

The kinetics of accumulation (per milliliter of culture) of the α- and β- subunits, associated with chloroplast-localized ammonium inducible nicotinamide adenine dinucleotide phosphate-specific glutamate dehydrogenase (NADP-GDH) isoenzymes, were measured during a 3 hour induction of synchronized daughter cells of Chlorella sorokiniana in 29 millimolar ammonium medium under photoautotrophic conditions. The β-subunit holoenzyme(s) accumulated in a linear manner for 3 hours without an apparent induction lag. A 40 minute induction lag preceded the accumulation of the α-subunit holoenzyme(s). After 120 minutes, the α-subunit ceased accumulating and thereafter remained at a constant level (i.e. steady state between synthesis and degradation). From pulsechase experiments, using ³⁵SO₄ and immunochemical procedures, the rate of synthesis of the α-subunit was shown to be greater than the β-subunit during the first 80 minutes of induction. The α- and β-subunits had different rates of degradation during the induction period (t_½ = 50 versus 150 minutes, respectively) and during the deinduction period (t_½ = 5 versus 13.5 minutes) after removal of ammonium from the culture. During deinduction, total NADP-GDH activity decreased with a half-time of 9 minutes. Cycloheximide completely inhibited the synthesis and degradation of both subunits. A model for regulation of expression of the NADP-GDH gene was proposed.     

Optimum prediction of three-way crosses from single crosses in forage maize (Zea mays L.)

Summary Three-way cross means were predicted with formulae involving linear functions of general (GCA) and specific combining ability (SCA) effects estimated from single-cross factorials between genetically divergent populations. Data from an experiment with 66 single-cross and 66 three-way cross forage maize (Zea mays L.) hybrids was used for comparing the prediction formulae. The genotypic correlation (r) between observed and predicted three-way crosses increased with increasing , the weighting factor of SCA effects, for plant height and ear dry matter (DM) content. It displayed slightly convex curves for total and stover DM yield, ear percentage, and metabolizable energy content of stover. For Jenkins' method B, r was considerably less than 1.0 for all traits, indicating the presence of epistasis. The square root of heritability (h) of the predicted means decreased with increasing , the reduction being small with a greater number of test environments. Using the product r·h as a criterion of efficiency, none of the prediction methods was consistently superior and the differences among them were rather small (< 7.5%) for all traits, irrespective of the number of test environments. We recommend evaluating the GCA of a greater number of lines from each parent population in testcrosses with a small number of elite lines from the opposite population. All possible three-way or double crosses between both sets of lines should be predicted by Jenkins's method C. This procedure allows one to select with a higher intensity among the predicted hybrids and thus should increase the genetic gain.Extended version of a paper (Geiger et al. 1986) read at the sixth meeting of the EUCARPIA Section Biometrics in Plant Breeding held at Birmingham, UK, July 28–August 1, 1986     

Involvement of the 2'-5' A pathway in the augmentation of natural killer activity

Pretreatment of human large granular lymphocytes (LGL) or unseparated peripheral blood mononuclear cells with interferon (IFN) resulted in a significant augmentation of natural killer (NK) activity. This increase was paralleled by an increase in the 2'-5'A synthetase activity. In order to investigate the possibility that IFN might be inducing augmentation of NK cells via the 2'-5'A pathway, we tested the effects of nonphosphorylated core material [(A2'p)2A] and of the triphosphorylated form of the 2'-5'A [ppp(A2'p)2A]. The core material had no detectable effect on NK activity. In contrast, when experiments were performed with the triphosphorylated form of 2'-5'A, NK activity was stimulated. In order to achieve activation, permeabilization of LGL with calcium chloride was necessary and, under these conditions, a dose-dependent augmentation of NK activity was seen. However, the calcium treatment had considerable toxic effects on basal levels of NK activity. Collectively, these results suggest that IFN may be inducing augmentation of NK activity via the 2'-5'A pathway. Further studies will be necessary to determine the effects of IFN and/or 2'-5'A on subsequent activation steps in the process leading to cytotoxicity by NK cells.     

Sulfur metabolism in Beggiatoa alba

The metabolism of sulfide, sulfur, and acetate by Beggiatoa alba was investigated under oxic and anoxic conditions. B. alba oxidized acetate to carbon dioxide with the stoichiometric reduction of oxygen to water. In vivo acetate oxidation was suppressed by sulfide and by several classic respiratory inhibitors, including dibromothymoquinone, an inhibitor specific for ubiquinones. B. alba also carried out an oxygen-dependent conversion of sulfide to sulfur, a reaction that was inhibited by several electron transport inhibitors but not by dibromothymoquinone, indicating that the electrons released from sulfide oxidation were shuttled to oxygen without the involvement of ubiquinones. Intracellular sulfur stored by B. alba was not oxidized to sulfate or converted to an external soluble form under aerobic conditions. On the other hand, sulfur stored by filaments of Thiothrix nivea was oxidized to extracellular soluble oxidation products, including sulfate. Sulfur stored by filaments of B. alba, however, was reduced to sulfide under short-term anoxic conditions. This anaerobic reduction of sulfur was linked to the endogenous oxidation of stored carbon and to hydrogen oxidation.     

Interaction of small dermatan sulfate proteoglycan from fibroblasts with fibronectin

Immunogold labeling was used to localize the core protein of small dermatan sulfate proteoglycan (DS-PG) on the surface of cultured human fibroblasts. At 4 degrees C, DS-PG core protein was uniformly distributed over the cell surface. At 37 degrees C, gold particles either became rearranged in form of clusters or remained associated with fibrils. Double-label immunocytochemistry indicated the co-distribution of DS-PG core protein and fibronectin in the fibrils. In an enzyme-linked immunosorbent assay, binding of DS-PG from fibroblast secretions and of its core protein to fibronectin occurred at pH 7.4 and at physiological ionic strength. Larger amounts of core protein than of intact proteoglycan could be bound. Fibronectin peptides containing either the heparin-binding domain near the COOH-terminal end or the heparin-binding NH2 terminus were the only fragments interacting with DS-PG and core protein. Competition and replacement experiments with heparin and dermatan sulfate suggested the existence of adjacent binding sites for heparin and DS-PG core protein. It is hypothesized that heparan sulfate proteoglycans and DS-PG may competitively interact with fibronectin.