Circadian and circannual variation of the carrageenin inflammatory effect in rat

The circadian variation of edema produced by carrageenin (carr.) administration into plantar tissue was studied in rats kept under a 12 light - 12 dark regimen. Three doses were used (125, 250 and 500 micrograms per rat) injected at different time (02.00, 08,00, 14.00 and 20.00 h). With the high doses, the level of edema for the four hour period after carr. administration was similar whatever the hour of injection. In contrast, with the lower dose (125 micrograms) a circadian rhythm in the intensity of the edema produced was observed, showing a maximum of susceptibility during the light span. Repetitive experiments performed at different periods of the year validated this finding. Comparing mean mesors, analysis of this data showed two distinct levels of inflammation, with the lower level observed in autumn and winter indicating evidence for a circannual variability.     

Purification and properties of a plasminogen activator from cultured rat prostate adenocarcinoma cells

Zymographic analysis of the supernates from confluent cultures of a rat prostate adenocarcinoma cell line, PA-III, revealed the existence of two molecular forms of specific plasminogen activators, one of molecular weight of approximately 80 000 and another of approximate molecular weight of 45 000, in sodium dodecyl sulfate. The low molecular weight form has been purified 364-fold in 66% yield from the culture medium by a combination of gel filtration on Sephacryl S-200 and affinity chromatography on Sepharose 4B-benzamidine. The purified material possessed a specific activity of 192 000 urokinase CTA units mg-1. This enzyme displayed activity toward human Glu1-plasminogen, characterized by a Km of 1.7 +/- 0.2 microM and a Vmax of 0.53 +/- 0.1 pmol of plasmin min-1 unit-1. A synthetic chromogenic substrate, H-D-Ile-Pro-Arg-p-nitroanilide (S-2288), was found for the activator. The enzyme possessed a Km of 0.33 mM and a kcat of 55 s-1 for S-2288. The activator was found to be a serine protease, inhibited by diisopropyl fluorophosphate (iPr2PF). At a concentration of 1 mM iPr2PF, and 30 nM enzyme, the half-time of this inhibition was 3.8 min. The 45 000 molecular weight enzyme was found to be inhibited by rabbit antibodies to human urokinase, thus characterizing the activator as a member of the urokinase class. The 80 000 molecular weight enzyme was not neutralized by anti-human urokinase but was neutralized by rabbit anti-human melanoma activator, likely allowing it to be classified as the tissue activator type.     

Glucose transport in Entamoeba histolytica

That the uptake of glucose by the parasitic amoeba Entamoeba histolytica occurs by an equilibrative transport system is supported by the following observations. 1. The rate of glucose uptake is several orders of magnitude greater than the uptake by pinocytosis. 2. The uptake of glucose exhibits saturation kinetics, with K(m)=1.6mm and V(max.) ranging from 2 to 5mumol/min per ml of cells at 37 degrees C. 3. The glucose analogues 2-deoxyglucose, 3-O-methylglucose and d-xylose are transported by the glucose system although with much less affinity. Competitive inhibition was observed between pairs of substrates, with K(i) values for any sugar closely coincident with the corresponding K(m). 4. d-Xylose, a sugar not metabolized by the cells, equilibrated with 80% of the amoebal cell water. 5. Cells equilibrated with xylose exhibited countertransport of this sugar against its concentration gradient when another substrate was added to the medium. 6. Blocking of glycolysis by iodoacetate or F(-) has no immediate effect on transport. The presence of a glucose-transport system in E. histolytica contrasts with the situation found in the non-parasitic amoeba, where pinocytosis seems to be the only mechanism of solute uptake.     

Unusual properties of a cold-labile fraction of Atlantic cod (Gadus morhua) brain microtubules

A cold-labile fraction of microtubules with unusual properties was isolated from the brain of the Atlantic cod (Gadus morhua). The yield was low, approximately six times lower than that for bovine brain microtubules. This was mainly caused by the presence of a large amount of cold-stable microtubules, which were not broken down during the disassembly step in the temperature-dependent assembly-disassembly isolation procedure and were therefore lost. The isolated cold-labile cod microtubules contained usually only a low amount of microtubule-associated proteins (MAPs). Three high molecular mass proteins were found, of which one was recognized as MAP2. Cod MAP2 differed from mammalian brain MAP2; it was not heat stable and had a slightly higher molecular mass. In contrast to mammalian MAPs, MAP1 was not found in the cold-labile fraction of microtubules. A new heat-labile MAP of higher molecular mass (400 kilodaltons) was however present, as well as a heat-stable protein of slightly lower molecular mass than MAP2. These MAPs showed similar tubulin-binding characteristics as bovine brain MAPs, since they coassembled with taxol-assembled bovine brain microtubules consisting of pure bovine tubulin. In spite of the fact that Ca2+ bound equally to cod and porcine tubulins, it did not inhibit cod microtubule assembly even at high concentrations (greater than 1 mM). In contrast, rings, spirals, and macrotubules were formed. The results show that there are major differences between this fraction of cod microtubules and microtubules from mammalian brain.     

A comparative study of ciliated protozoa communities in activated-sludge plants

Abstract: Ciliated protozoa present in ten activated sludge plants at Madrid (Spain) were identified. The abundance of key groups of ciliates was determined at each plant; attached ciliates made up the most abundant and representative group. Multivariate statistical analysis was employed to study relationships between ciliates and both the physico-chemical and operational parameters of the plants. Partial correlation analysis revealed: (1) The indicator value of attached ciliates in assessing management and performance of the activated sludge process, (2) the relationship of swimming ciliates with short-aged sludges and lower quality effluents and (3) the direct association between swimming-crawling ciliates and bad settlement conditions of the sludge. Factor analysis showed the associations of the most frequent species of ciliates with the operational parameters of the plants, suggesting the indicator value of some of the species: Vorticella striata was related with poor quality of effluent; Aspidisca cicada with stable plant conditions, and Litonotus lamella with a deficiently settling sludge.     

Inter-varietal differences in xylem exudate composition and growth under contrasting forms of N supply in cucumber

Variations in the inorganic and organic composition of xylem exudate, growth and N content under contrasting forms of N supply in three cucumber cultivars (Hyclos, Medusa and Victory) were studied in glasshouse conditions. The plants were grown hydroponically with two NO₃ ^-:NH₄ ⁺ ratios (100:0 and 60:40).The xylem sap of Medusa grown with both N sources displayed an increase of organic N and carboxylate concentrations and a decrease of cations, inorganic anions and carbohydrates compared with that of those grown with NO₃ ^- alone, showing a higher growth and N content in tissues and thus better utilization of N supplied as NO₃ ^- and NH₄ ⁺. Mixed N nutrition in Hyclos caused the greatest amounts of NO₃ ^- and NH₄ ⁺ in xylem sap, lower root weight and N levels in the leaves, while its root was unable to generate an adequate supply of organic N compounds. Despite the levels of cations, inorganic and organic anions were reduced by the NH₄ ⁺ supplied to Victory, the ionic balance in the xylem sap, growth and N content remained similar to that of those supplied with NO₃ ^- alone. Finally, the cucumber cultivars studied here, responded differently to the form of N supplied, it may partly be due to their ability of assimilating N in the roots and partly to the form in which the N is translocated to the shoot.     

Identification of two different glyceraldehyde-3-phosphate dehydrogenases (phosphorylating) in the photosynthetic protist Cyanophora paradoxa

Two different glyceraldehyde-3-phosphate (G3P) dehydrogenase (phosphorylating) activities, namely NAD- and NADP-dependent, have been found in cell extracts of the cyanelle-bearing photosynthetic protist Cyanophora paradoxa. Whereas the two G3P dehydrogenase activities were detected with similar specific activity levels (0.1 to 0.2 U/mg of protein) in extracts of the photosynthetic organelles (cyanelles), only the NAD-dependent activity was found in the cytosol. Thus, a differential intracellular localization occurred. The perfect overlapping of the two G3P dehydrogenase activity peaks of the cyanelle in both hydrophobic interaction chromatography and subsequent FPLC (fast protein liquid chromatography) gel filtration indicated that the two activities were due in fact to a single NAD(P)-dependent G3P dehydrogenase (EC 1.2.1.-) with a molecular mass of 148,000. SDS-PAGE of active fractions from FPLC gel filtration showed that the intensity of the major protein band (molecular mass, 38,000) of the enzyme preparation clearly paralleled the activity elution profile, thus suggesting a tetrameric structure for the cyanelle dehydrogenase. On the other hand, FPLC gel filtration analysis of the cytoplasmic fraction revealed a NAD-dependent G3P dehydrogenase with a native molecular mass of 142,000, being equivalent to the classical glycolytic enzyme (EC 1.2.1.12) present in the cytosol of all the organisms so far studied. The significance of these results is discussed taking into account that the cyanobacteria, photosynthetic prokaryotes which share many structural and biochemical features with cyanelles and are considered as their ancestors, have a similar NAD(P)-dependent G3P dehydrogenase.Abbreviation FPLC Fast protein liquid chromatography     

Immunolocalization of the Plasma Membrane H+ -ATPase in Minor Veins of Vicia faba in Relation to Phloem Loading

The phytotoxin fusicoccin (FC), after binding to a plasma membrane-localized receptor, causes higher plant cells to excrete protons. Ligand-binding analysis has been used to show that the plasma membrane of mung bean (Vigna radiata L.) hypocotyls contains both high-affinity (HA) and low-affinity (LA) binding sites for FC. The effect of tissue maturation on these sites was determined on isolated membrane vesicles from the meristematic region (hook) and the elongation zone and from mature hypocotyl tissues. In the meristematic region the HA:LA ratio was 1:20. As hypocotyl tissues matured, the site density of HA increased and there was no change in LA density, so that the HA:LA ratio increased to 1:2 in maturet issues. FC-induced proton excretion correlates with the HA density, not the LA density. When sections isolated from each region were incubated with FC prior to isolation of membranes, there was an apparent conversion of LA to HA sites during the first 90 min in all regions. During the next 1 to 3 h there was a further 2.5- to 3- fold increase in binding sites in all regions, accompanied by a slight decline in dissociation constant. The increase in binding sites, but not the apparent conversion of LA to HA, was partly blocked by cycloheximide. These data suggest that FC alters FC-binding protein activity in two ways: first, by causing an increase in affinity for FC of preexisting LA receptors, and second by inducing the synthesis of additional FC receptors. This apparent up-regulation of a phytotoxin receptor by its ligand in plants has not previously been reported.     

Salt tolerance and methionine biosynthesis in Saccharomyces cerevisiae involve a putative phosphatase gene.

The progressive salinization of irrigated land poses a threat to the future of agriculture in arid regions. The identification of crucial metabolic steps in salt tolerance is important for the understanding of stress physiology and may provide the tools for its genetic engineering. In the yeast Saccharomyces cerevisiae we have isolated a gene, HAL2, which upon increase in gene dosage improves growth under NaCl and LiCl stresses. The HAL2 protein is homologous to inositol phosphatases, enzymes known to be inhibited by lithium salts. Complementation analysis demonstrated that HAL2 is identical to MET22, a gene involved in methionine biosynthesis. Accordingly, methionine supplementation improves the tolerance of yeast to NaCl and LiCl. These results demonstrate an unsuspected interplay between methionine biosynthesis and salt tolerance.