Evidence for precursor forms of the low isoelectric point alpha-amylase isozymes secreted by barley aleurone cells

Gibberellin-treated barley (Hordeum vulgare L.) aleurone cell protoplasts have been shown previously to contain two α-amylase isozymes which are not secreted (JV Jacobsen, JA Zwar, PM Chandler 1985 Planta 13: 430-438). This report shows that these intracellular forms are immunochemically related to the low isoelectric point but not the high isoelectric point group of α-amylase isozymes and that they arise by new synthesis like the secreted forms. Pulse-chase studies show that the intracellular isozymes are precursors to the secreted isozymes. Conversion of the intra- to the extracellular forms involves decreases in isoelectric points with no change in size detectable by SDS-PAGE. The precursor isozymes were also detected in aleurone layer homogenates but they were unstable. They could be stabilized by various treatments including heating the homogenate to 70°C for 10 minutes indicating that the instability was enzymically mediated. Using purified radioactive precursor isozymes, it was shown that instability did not involve inactivation but the conversion to secreted forms. The nature of the covalent modification associated with conversion was not determined but available data indicate that it does not involve glycosylation.     

Monensin inhibits the secretion of α-amylase but not polysaccharide slime from seedling tissues ofZea mays

Summary The effect of monensin on polysaccharide slime secretion by root tips of corn (Zea mays) was studied. Various treatment times and ionophore concentrations were tested: none resulted in inhibition of slime secretion. Because monensin changes the pH of the medium, its effect was also monitored in strongly buffered media and at different pH's. Even in such media, monensin did not inhibit slime secretion. We also measured the effect of the drug after a pulse with [³H]fucose or a pulse followed by a chase. The amount of labeled slimed secreted was not altered by the ionophore. However, 10M monensin affected the development of root tips and drastically reduced their growth. We showed that monensin inhibits the secretion of -amylase by the scutellum of the same plantlet. The importance of the nature of the secretory compound in relation to monensin inhibition of its secretion is discussed.Abbrevations Hepes N-2-hydroxyethylpiperazine-N-2-ethane-sul-fonic acid - Mes 2-(N-morpholino)ethane-sulfonic acid     

Histochemical and immunocytochemical study of the migration of neurons from the rat olfactory placode

Immunocytochemical and histochemical methods have been used to describe the neuronal population migrating from the rat olfactory placode and to analyze the spatio-temporal evolution of this neuronal migration during development. Several neuronal markers, such as binding to the lectin Ulex europaeus (UEA I) and the presence of neuron-specific enolase (NSE), olfactory marker protein (OMP), and luteinizing hormone-releasing hormone (LHRH), have been tested in order to determine whether migrating neurons originate from both the medial and the lateral parts of the placode and whether they all express LHRH. Our data show that a large population of differentiated migrating neurons can be identified with an antibody against NSE from the 14th day of gestation and with UEA I one day later. Migrating neurons are closely associated with both the vomeronasal axon fascicles emerging from the medial pit and the olfactory axons originating from the lateral pit. However, the neuron migration from the lateral pit appears to be more discrete than that from the medial pit. No LHRH immunoreactivity has been detected among neurons migrating from the lateral pit. Some neurons accompanying the olfactory axon fascicles exhibit a high level of maturation as shown by their OMP-positivity. Numerous neurons positive for both NSE and UEA I have also been observed within the presumptive olfactory nerve layer in early embryonic stages.     

Interactions of HIV-1 and HIV-2 envelope glycoproteins with sulphated polysaccharides and mannose-6-phosphate

Envelope glycoproteins of human immunodeficiency viruses (HIV-1and HIV-2) can interact with high-mannose glycans and with themannosyl or N-acetylglucosaminyl core of complex-type oligosaccharidicstructures. HIV-1 glycoproteins also specifically bind sulphatedpolysaccharides such as dextran sulphate (DS) and heparin. Here,we show that the latter property is shared by HIV-2 recombinantgp140 (rgpl40) precursor glycoprotein. Binding of rgpl40 andof corresponding rgp160 of HIV-1 to heparin- and DS-substituted(sulphated dextran beads; SDB) affinity matrices was inhibitedby the soluble specific ligand and also by fetuin, asialofetuinor the anionic simple carbohydrate derivative manncsse-6-phosphate(M6P). Interaction of HIV-1 rgpl20 subunit with the two affinitymatrices was also inhibited by M6P, but only rgpl20 bindingto heparin-agarose, and not that to SDB, was affected by fetuinand asialofetuin. These results suggest that HIV-1 and HIV-2envelope glycoproteins presumably display different sulphatedpolysaccharide and carbohydrate recognition sites. Some of thesemay be common or in close proximity: with respect to rgpl60,for example, the sites may be common on the gp41 moiety and/orin a region of gp120 which would be more accessible when expressedon rgpl60 than on processed gpl20, while they may be distincton the cleaved gpl20 subunit. Finally, because M6P is a markerof lysosomal enzymes, we verified that HIV-1 and HIV-2 envelopeglycoproteins could specifically bind in a M6P-inhibitable mannerto a representative lysosomal enzyme, bovine liver ß-glucuronidasecoupled to agarose, suggesting that they may possibly interferewith lysosomal enzyme sorting in HIV-infected cells. env glycoproteins HIV lectin mannose-6-phosphate sulphated polysaccharides     

Some biochemical properties of an acyclic oligonucleotide analogue. A plausible ancestor of the DNA?

As acyclic oligonucleotides have been suggested as a primitive model of DNA or RNA in prebiotic times, we compared some biochemical properties of these analogues to that of natural ones. Firstly, an acyclic analogue of deoxyribonucleoside triphosphates was tested as a potential substrate of enzymes intervening in nucleic acids synthesis. GlyTTP, a dTTP analogue with a missing 2-methylene group is notaccepted as a substrate by either DNA polymerase or deoxynucleotidyl terminal transferase (TdT). Secondly, themodified dodecathymidylate (GlyT)₁₂, the racemic acyclic sugar analogue of (dT)₁₂, proved to be anefficient primer for DNA polymerase and TdT, though the associative properties of (GlyT)₁₂ are very weak as shown by UV spectroscopy in phosphate buffer without magnesium chloride. But (GlyT)₁₂ has the advantage to be 500-times more stable against hydrolysis by snake venom phosphodiesterase than the corresponding oligothymidylate.     

In vitro study of the proteolytic activity of rumen anaerobic fungi

Abstract To better define the antigenic structure of the outer cell membranes of Legionellae, a panel of 6 monoclonal antibodies was raised against partially purified outer membranes of Legionella pneumophila serogroup 1, Corby strain. This study describes the purification and characterization of one of these monoclonal antibodies reacting with a 135-kDa protein, which was shown to be common to all 14 serogroups of Legionella pneumophila . It shows no cross-reactivity with 20 other Legionella species, or 9 other Gram-negative species tested by SDS-PAGE and Western blotting procedures. The epitope would appear to be predominantly surface exposed and, from preliminary detergent extraction studies, not peptidoglycan-associated.     

Interactions between the toxicity of the heavy metals cadmium,copper, zinc in combinations and the detoxifying role of calcium in the brown algaCystoseira barbata

The toxicity of three heavy metals, Cd, Cu and Zn, and the detoxifying role of Ca have been studied for the brown algaCystoseira barbata formaaurentia after a 4-week laboratory culture. The experimental design was based upon a complete factorial design 2^k, which seems to be the first time it has been used in algal physiology. It was demonstrated that these three elements, applied jointly, act on weight-growth, chlorophyll a, c and carotenoid synthesis and Cd, Cu and Zn uptake. Cd and Zn act in synergy or in antagony, depending on their exogenous concentrations, on chlorophyll a and on carotenoid synthesis. Zn is antagonistic towards Cd and Cu on weight-growth in the combination Cd-Cu-Zn. From different element combinations, the protective role of Ca appears evident on weight-growth (Cd-Zn-Ca and Cu-Ca), chlorophyll a (Cd-Cu-Ca and Cu-Zn-Ca), chlorophyll c (Cd-Ca), carotenoid synthesis (Cd-Cu-Ca and Cu-Zn-Ca), Cd and Cu uptake (Cd-Cu-Ca) and Zn uptake (Cu-Zn-Ca). This role is confirmed by cytological investigations. This is apparently the first report concerning a Ca interaction with toxicity of heavy metals applied in combinations. However, the mechanisms of tolerance remain unknown.     

Systemic Responses in Arabidopsis thaliana Infected and Challenged with Pseudomonas syringae pv syringae

Attack of plants by necrotizing pathogens leads to acquired resistance to the same or other pathogens in tissues adjacent to or remotely located from the site of initial attack. We have used Arabidopsis thaliana inoculated with the incompatible pathogen Pseudomonas syringae pv syringae on the lower leaves to test the induction of systemic reactions. When plants were challenged with Pseudomonas syringae pv syringae in the upper leaves, bacterial titers remained stable in those preinfected on the lower leaves. However, there was a distinct decrease in symptoms that correlated with a local and systemic increase in salicylic acid (SA) and in chitinase activity. Peroxidase activity only increased at the site of infection. No changes in catalase activity were observed, either at the local or at the systemic level. No inhibition of catalase could be detected in tissue in which the endogenous levels of SA were elevated either naturally (after infection) or artificially (after feeding SA to the roots). The activity of catalase in homogenates of A. thaliana leaves could not be inhibited in vitro by SA. SA accumulation was induced by H2O2 in leaves, suggesting a link between H2O2 from the oxidative burst commonly observed during the hypersensitive reaction and the induction of a putative signaling molecule leading to system acquired resistance.