Gibberellins in suspensor,embryo and integument from very young seeds of Phaseolus coccineus L.

Endogenous gibberellins (GAs) were extracted from suspensor, embryo and integument of very young seeds of Phaseolus coccineus L. and detected by combined gas chromatography-mass spectrometry (GC-MS). Results show the presence of one C₂₀-GA, GA₄₄ and five C₁₉-GAs in the suspensor: GA₁, GA₄, GA₅, GA₆ and GA₈, and four C₁₉-GAs in the integument: GA₁, GA₅, GA₆ and GA₈. Only traces of GA₁ and GA₅ were identified in the embryo. A compound structurally related to GAs was identified as tetrahydroxy-Kauranoic acid in suspensor, integument and, only in trace amounts, in the embryo.     

Toxins of Pseudomonas syringae pv. syringae affect H+-transport across the plasma membrane of maize

The activity of some phytotoxic metabolites of Pseudomonas syringae pv. syringae Van Hall strains B359 and B301 on in vivo and in vitro systems of H⁺-transport across the plasma membrane of maize (Zea mays L., hybrid Paolo) was investigated. In particular syringomycin, the first lipodepsinonapeptide isolated from Pss and already studied in plants and yeasts for its effects on several physiological systems, was compared with the recently described lipodepsipeptides with 22 or 25 amino acid residues, so called syringopeptins. The in vivo activity of the phytotoxins was tested on fusicoccin-stimulated H^?-extrusion from cuttings of maize roots, which was inhibited by both types of toxins, with syringomycin more efficient than the syringopeptins. In vitro the H⁺-ATPase activity of predominantly right-side-out plasma membrane vesicles purified by two-phase partitioning was stimulated by 10 μM syringomycin and inhibited by higher levels, in agreement with the results of others with preparations of dicotyledons. Also the inhibition of the phosphohydrolytic activity of inside-out vesicles of mung bean plasma membrane was confirmed for maize. In both types of vesicles the syringopeptirts were better inhibitors than syringomycin. The pH gradient formed on addition of ATP to predominantly (25% latency) inside-out vesicles was immediately and completely collapsed by syringomycin and syringopeptins; H⁺-pumping was prevented if the toxins were added before ATP. The inhibition was concentration dependent, but at very low concentrations the effect was inverted. The results of the present investigation, carried out with maize preparations, confirm and extend the evidence so far obtained with dicotyledons in favour of the plasma membrane as an important site of interaction of syringomycin with the plant cell. They also indicate that, except for some details, the effects of syringopeptins at the level of the plasma membrane are the same as those of syringomycin.     

Polygalacturonase-inhibiting protein accumulates in Phaseolus vulgaris L. in response to wounding, elicitors and fungal infection

Polygalacturonase-inhibiting protein (PGIP) is a cell wall-associated protein that specifically binds to and inhibits the activity of fungal endopolygalacturonases. The Phaseolus vulgaris gene encoding PGIP has been cloned and characterized. Using a fragment of the cloned pgip gene as a probe in Northern blot experiments, it is demonstrated that the pgip mRNA accumulates in suspension-cultured bean cells following addition of elicitor-active oligogalacturonides or fungal glucan to the medium. Rabbit polyclonal antibodies specific for PGIP were generated against a synthetic peptide designed from the N-terminal region of PGIP; the antigenicity of the peptide was enhanced by coupling to KLH. Using the antibodies and the cloned pgip gene fragment as probes in Western and Northern blot experiments, respectively, it is shown that the levels of PGIP and its mRNA are increased in P. vulgaris hypocotyls in response to wounding or treatment with salicylic acid. Using gold-labeled goat-anti-rabbit secondary antibodies in EM studies, it has also been demonstrated that, in bean hypocotyls infected with Colletotrichum lindemuthianum, the level of PGIP preferentially increases in those cells immediately surrounding the infection site. The data support the hypothesis that synthesis of PGIP constitutes an active defense mechanism of plants that is elicited by signal molecules known to induce plant defense genes.     

Effect of propylthiouracil-induced hypothyroidism on cerebral cortex of young and aged rats: Lipid composition of synaptosomes,muscarinic receptor sites,and acetylcholinesterase activity

The effect of hypothyroidism on the lipid composition of synaptosomes, density and affinity of muscarinic receptor sites, and acetylcholinesterase activity in the cerebral cortex of young and aged rats was investigated. The animals were made hypothyroid by adding 0.05% propyl-2-thiouracil to their drinking water for four weeks. This pathological state induced an increase in the relative percentage of sphingomyelin in young rats. In aged rats hypothyroidism induced a decrease of sphingomyelin and glycerophosphocholine and an increase of cholesterol. The effect of hypothyroid state on cerebral cortex resulted in an increase of acethylcholinesterase activity both in young and aged rats and was also reflected in an increase of density of M1-AChRs but only in the former.     

Determination of ammonium and L-Glutamine in hybridoma cell cultures by sequential flow injection analysis

A flow injection anlytical system based on a gas diffusion membrane module for ammonia and an ammonium flow-through potentiometric detector has been set up for measurement of L-glutamine and ammonium ions in hybridoma cell cultures. The main feature of the system is that the same basic analytical concept and equipment is used in both measurements, the only difference being for the determination of L-glutamine, in which the sample flows through an immobilized glutaminase cartridge. The conditions to enable the performance of both analysis consecutively, avoiding potential interferences by unwanted deamination of other compounds in the samples, have been determined. Finally, the proposed system has been compared with reference analytical methods for batch hybridoma cell culture experiments.     

Low temperature optical spectroscopy of cobalt-substituted hemocyanin from Carcinus maenas

In this work we report the optical absorption spectra of three cobalt-substituted derivatives of hemocyanin (He) from Carcinus maenas, in the temperature range 300–20 K. The derivatives studied are the mononuclear (Co²⁺)-He with a single cobalt ion in the Cu_A site, the binuclear (Co²⁺)₂-He and the binuclear mixed metal (Co²⁺-Cu¹⁺)-He. At low temperature three main bands are clearly resolved; the temperature dependence of their zeroth, first and second moments sheds light on the stereodynamic properties in the surroundings of the chromophore. Within the limits of the reported analysis, in the binuclear derivatives the motions coupled to the chromophore appear to be essentially harmonic in the whole temperature range investigated; moreover the data are consistent with the presence of an exogenous ligand strongly bound to the two metal ions. For the mononuclear derivative an essentially harmonic behavior is evident only up to 200 K where the data are consistent with the presence of an exogenous ligand much less strongly bound, while at higher temperatures the behavior of the spectra indicates the onset of very large anharmonic contributions to motions, that plausibly involve the above exogenous ligand and, quite likely, the entire active site.Abbreviations He Hemocyanin - M₀ zeroth moment - M₁ first moment - M₂ second moment - (Co^2–)₂-He binuclear bicobalt hemocyanin derivative - (Co²⁺)-He mononuclear monocobalt hemocyanin derivative - (Co²⁺-Cu¹⁺)-He binuclear mixed metals hemocyanin derivative - LFT ligand field theory - CT charge transfer - EPR electronic paramagnetic resonance - XANES X-ray absorption near edge structure Correspondence to: L. Cordone