Expression of a novel chitinase by the fungal endophyte in Poa ampla

Many wild and cultivated cool-season grass species are naturally infected with fungal endophytes of the genera Neotyphodium and Epichlo?. These associations generally are considered mutualistic with the plants benefiting from reduced herbivory and the fungi benefiting from nutrients supplied by the plants. The fungi secrete proteins that might have a role in the interspecies symbiosis. In the interaction between Poa ampla Merr. and the endophyte Neotyphodium sp., a fungal chitinase was detected in the apoplastic protein fraction. The chitinase was also the major protein secreted in culture. Sequence analysis of the chitinase revealed it has a low level of amino acid sequence identity to other fungal chitinases and one of the conserved active site residues is altered. DNA gel-blot analysis indicated the chitinase was encoded by a single gene. Expression of similar chitinases also was detected in endophyte-infected tall fescue (Festuca arundinacea Schreb.), perennial ryegrass (Lolium perenne L.) and Chewings fescue (Festuca rubra L. subsp. fallax [Thuill] Nyman). This is the first report of an endophyte chitinase expressed in the infected host grass. As a secreted hydrolytic enzyme, the chitinase might have roles in the nutrition, growth or defense of the endophyte.     

Antifungal activity of extracellular metabolites produced by Sporothrix flocculosa

The antifungal properties of extracellular compounds produced by the epiphytic fungus Sporothrix flocculosa were bioassayed against phytopathogenie fungi on the basis of inhibition of spore germination, and mycelial growth and induction of cellular leakage. Following incubation in stationary culture, S. flocculosa released antifungal metabolites into the culture medium which were extractable with méthylene chloride. When separated by thin layer chromatography, extracted metabolites yielded a compound(s) at R_f0.65 which inhibited development of Cladosporium cucumerinum and several other phytopathogenic fungi. Treatment of Botrytis cinerea and Fusarium oxysporum f.sp. radicis‐lycopersici (FORL) with the same compound(s) greatly reduced spore germination and biomass growth of both fungi. Additionally, both B. cinerea and FORL leaked electrolytes and proteins when grown in presence of the metabolites. Observations under electron microscopy revealed that FORL reacted to the presence of S. flocculosa metabolites by retraction of the plasmalemma and rapid disintegration of the cytoplasm. These reactions were similar to the ones induced by conidia of S. flocculosa when applied on powdery mildew fungi. These results provide strong evidence of the production of antifungal compounds in vivo and of their role in the antagonistic properties of S. flocculosa.     

Stoichiometry and compartmentation of NADH metabolism in Saccharomyces cerevisiae

In Saccharomyces cerevisiae, reduction of NAD(+) to NADH occurs in dissimilatory as well as in assimilatory reactions. This review discusses mechanisms for reoxidation of NADH in this yeast, with special emphasis on the metabolic compartmentation that occurs as a consequence of the impermeability of the mitochondrial inner membrane for NADH and NAD(+). At least five mechanisms of NADH reoxidation exist in S. cerevisiae. These are: (1) alcoholic fermentation; (2) glycerol production; (3) respiration of cytosolic NADH via external mitochondrial NADH dehydrogenases; (4) respiration of cytosolic NADH via the glycerol-3-phosphate shuttle; and (5) oxidation of intramitochondrial NADH via a mitochondrial 'internal' NADH dehydrogenase. Furthermore, in vivo evidence indicates that NADH redox equivalents can be shuttled across the mitochondrial inner membrane by an ethanol-acetaldehyde shuttle. Several other redox-shuttle mechanisms might occur in S. cerevisiae, including a malate-oxaloacetate shuttle, a malate-aspartate shuttle and a malate-pyruvate shuttle. Although key enzymes and transporters for these shuttles are present, there is as yet no consistent evidence for their in vivo activity. Activity of several other shuttles, including the malate-citrate and fatty acid shuttles, can be ruled out based on the absence of key enzymes or transporters. Quantitative physiological analysis of defined mutants has been important in identifying several parallel pathways for reoxidation of cytosolic and intramitochondrial NADH. The major challenge that lies ahead is to elucidate the physiological function of parallel pathways for NADH oxidation in wild-type cells, both under steady-state and transient-state conditions. This requires the development of techniques for accurate measurement of intracellular metabolite concentrations in separate metabolic compartments.     

Proctolin: a review with emphasis on insects

The distribution, physiological role, mode of action, and pharmacology of the pentapeptide neuroregulator proctolin are reviewed, with special emphasis on insects. Whereas proctolin is distributed extensively throughout arthropods, its presence in molluscs, annelids, or chordates is not well established. In the arthropods, proctolin acts as a neuromodulator and possibly as a neurohormone. It does not appear to function as a conventional neurotransmitter. Two model proctolinergic systems are highlighted: motor control of the visceral muscles of the locust oviduct and of the skeletal muscles of the locust ovipositor. In these preparations proctolin is a cotransmitter acting to enhance neuromuscular transmission and muscular contraction. The mode of action of proctolin is not well understood, although the second messengers cAMP, phosphatidyl inositol, and calcium have been implicated in various systems. Pharmacologically, the proctolin receptor has been examined with structure/activity studies, and the effects of a variety of amino acid substitutions and deletions of the pentapeptide are described. It is unfortunate that no specific antagonists of the proctolin receptor appear to be available and that no receptor-binding studies have been reported. The prospects are good for advances in our understanding of modulatory mechanisms, since proctolin appears to be emerging as the model for studies of this type.     

Changes in plasma steroid levels after single administration of hCG or LHRH agonist analogue in dog and rat

The present study was designed to investigate the effect of acute administration of gonadotropin on testicular steroid secretion in dog and rat. Animals received a subcutaneous injection of 25 IU/kg of hCG or 1.5 microgram/kg of [D-Trp6, des-Gly-NH2(10)]LHRH ethylamide (LHRH-A). Testosterone is the predominant steroid measured, in dog plasma, under basal conditions. After LHRH-A injection, testosterone levels are not significantly changed while dehydroepiandrosterone and androst-5-ene-3 beta,17 beta-diol (delta 5-steroids) levels are stimulated by almost 20-fold (P less than 0.01). When dogs were injected with hCG, we also observed a marked stimulation of dehydroepiandrosterone levels (20-fold; P less than 0.01) accompanied by a small increase of plasma testosterone concentration (2-fold, P less than 0.01). In rats injected with either hCG or the LHRH analogue, an increment of plasma testosterone (7-fold, P less than 0.01) is detected in the first hour while plasma dehydroepiandrosterone levels are slightly stimulated. Moreover, in rats injected with hCG, low plasma steroid levels are present between 4-12 h after injection due to testicular desensitization. This marked decrease is then followed by a second peak of steroid secretion 24 h later. Acute testicular steroidogenic responsiveness to hCG on the dog is, however, different: after stimulation, the levels of plasma dehydroepiandrosterone are maintained at a plateau and slowly decline after 24-48 h. Our data indicate that in dogs, stimulation of testicular steroidogenesis leads to an increase of plasma delta 5-steroid levels while the same stimuli cause, in the rat, a stimulation of delta 4-androgen, particularly testosterone.