The intrinsic synthesis of juvenile hormone-III diol by locust corpora allata in vitro.

Juvenile hormone (JH)-III 10-11-diol is intrinsically synthesized and released from the corpora allata (CA) of adult locust females in vitro, together with JH-III. JH-III synthesis is preferentially stimulated and diol production only slightly enhanced, by cerebral locust allatotropin. The identification of JH-III diol is based on: similar ratio of incorporation of 14C/3H from radiolabelled [2-14C]acetate and [methyl-3H]methionine, to that of JH-III; similar chromatographic properties to those of synthetic diol on an RP-18 column eluted with acetonitrile, and similar chromatographic properties of acetylated derivatives; mass spectrometric analysis of derivatives and fragmentation products. Exogenous radiolabelled JH-III is not degraded during incubation with locust CA in vitro, corroborating the endogenous production of JH-III diol. Allatal diol formation may be an additional mechanism for the control of JH-III levels in locusts, preceding release into the hemolymph.

Inhibition of Manduca sexta corpora allata in vitro by a cerebral allatostatic neuropeptide.

When an in vitro assay system and radioimmunoassays specific for juvenile hormones (JH) I and III were used to probe the effect of day 4 last instar larval brains on JH synthesis by day 0 last instar larval corpora allata (CA) of the tobacco hornworm, Manduca sexta, a selective inhibition of JH I synthesis by the CA was observed. The nature of this inhibition suggested the presence of an allatostatin specific for the synthesis of JH I. Its occurrence in the day 4 brain was demonstrated by the ability of a crude brain extract to inhibit the CA in a dose-dependent manner. The allatostatic factor (ASF) appears to be a protein, based on its heat lability and pronase sensitivity, and it has apparent molecular weights of 6.8 and 13 kDa. Inhibition of JH I synthesis occurs within 1 min of exposure of the CA to the factor and is reversible by 6 h after this exposure. Thus it appears that a cerebral neuropeptide specifically inhibiting JH I synthesis by the CA is present in Manduca on day 4 of the last larval instar, a time when the hemolymph titer of JH must drop to ensure the occurrence of pupal commitment.

Juvenile hormone synthesis in vitro by larval and pupal corpora allata of Manduca sexta.

The synthesis of juvenile hormone I (JH I) and juvenile hormone III (JH III) in vitro by isolated corpora allata (CA) and brain-corpora cardiaca-corpora allata complexes (BR-CC-CA) from fourth and fifth instar larvae and pupae of Manduca sexta was assessed by JH I and JH III radioimmunoassays of incubation medium. Fluctuations in synthesis occurred at specific stages during larval-pupal development. These fluctuations temporally approximated those of previously determined hemolymph titers of JH I and III for this developmental period, except for the synthesis of JH III during the prepupal period, when the JH III hemolymph titer was negligible but JH III synthesis in vitro increased to the highest rate observed. The overall agreement between CA activity and the JH hemolymph titers suggests that the in vitro system supports physiological gland activity and that changes in biosynthesis contribute significantly to changes in the titers. A comparison of JH I and JH III synthesis by isolated CA with that by BR-CC-CA suggests that the BR-CC can differentially modulate the synthesis of these two homologs. The significance of these findings with regard to the possible endocrine function of JH III and the proposed neuroendocrine control of the CA is discussed.

Farnesoic acid and allatotropin stimulation in relation to locust allatal maturation.

Allatal maturation during the first gonotrophic cycle of adult female Locusta migratoria can be measured in vitro by the competence of these glands to produce juvenile hormone III (JH-III) when maximally stimulated by either farnesoic acid (FA) or allatotropin. Maturation precedes the full activation of the corpora allata (CA) during the first cycle of oogenesis. Basal activity of mature active female CA is highly correlated to stimulated activity. This suggests that between-gland variability reflects the inherent competence of each individual CA, and is not a consequence of a mere pulsatile on/off activation. Older glands may be inactive but their competence is equivalent to that of mature active glands. The kinetics of FA and allatotropic stimulation in vitro are equivalent, suggesting that allatotropin affects rate-limiting steps preceding FA biosynthesis.

Characterization and regulation of HMG-CoA reductase during a cycle of juvenile hormone synthesis.

The enzyme 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase was characterized in cockroach corpora allata which produce insect juvenile hormone III (methyl-(10R)10,11-epoxy-3,7,11-tri-methyl-2E,6E-dodecadienoate ). HMG-CoA reductase is a microsomal enzyme dependent on NADPH and dithiothreitol (or glutathione) for activity. The enzyme selectively reduced (3S)-HMG-CoA to (3R)-mevalonate with an apparent KM of 7.6 microM. Mevinolin was a competitive inhibitor of HMG-CoA reductase with a KI of 2.4 nM. No evidence for a modulation of enzyme activity by phosphorylation was obtained. Levels of HMG-CoA reductase were not altered after incubation of the corpora allata with either mevinolin (to decrease isoprenoid flux) or with mevalonate or farnesol (to increase isoprenoid flux). Split pairs of corpora allata were used to compare JH III synthetic activity with HMG-CoA reductase activity during the cycle of JH III synthesis that controls vitellogenesis and oocyte growth in adult females. Both activities changed over 10-fold and peaked on day 5 after emergence/mating, but JH III synthesis did not parallel HMG-CoA reductase activity precisely thereafter. The half-life of HMG-CoA reductase measured in the presence of cycloheximide was significantly different between low and high activity glands and was not related to the half-life of JH III synthesis. The results suggest that HMG-CoA reductase should not be considered 'the rate-limiting enzyme' in juvenile hormone synthesis by Diploptera punctata corpora allata.

Studies on the biosynthesis of terrecyclic acid A, an antitumor antibiotic from Aspergillus terreus.

The biosynthesis of terrecyclic acid A was investigated using 13C-labeled acetates and mevalonate. 13C NMR spectral analysis of isolated labeled terrecyclic acid demonstrated that the structure is assembled via an isoprene pathway.

Regulation of enzymes involved in the biosynthesis of the sesquiterpene antibiotic pentalenolactone in Streptomyces arenae.

The production of the sesquiterpenoid antibiotic pentalenolactone in the producer strain Streptomyces arenae TU 469 is controlled by the activity of the enzyme farnesylpyrophosphate cyclase. In contrast to the activity of this enzyme, the specific activities of all other enzymes of the mevalonoid pathway tested so far, proved to be not rate-limiting. Several metabolites of the pentalenolactone pathway were tested for inhibitory effects on the activity of the HMG-CoA reductase and farnesylpyrophosphate cyclase. The activity of the cyclase was inhibited by low concentrations of pentalenolactone and its derivatives, thus suggesting an end product inhibition of the starting enzyme of the pentalenolactone pathway. The activity of HMG-CoA reductase was not inhibited by pentalenene or any pentalenolactone-derivatives. According to these results, an end-product inhibition of the first enzyme which is specific for pentalenolactone synthesis seems to be a mechanism involved in the regulation of pentalenolactone biosynthesis.

Studies on biosynthesis of pentalenolactone. V isolation of deoxypentalenylglucuron.

Deoxypentalenylglucuron was isolated from the culture broths of three different strains, such as Streptomyces omiyaensis, S. albofaciens and S. viridifaciens. The structure of deoxypentalenylglucuron has been determined by 1H and 13C NMR, mass spectroscopy and by chemical correlation to be an oxidation product of pentalenene 1. Deoxypentalenylglucuron (1) has some antitumor activity against Sarcoma 180 in mice.

Specific inhibition of glyceraldehyde-3-phosphate dehydrogenase by koningic acid (heptelidic acid).

A sesquiterpene named koningic acid has been isolated from a strain of Trichoderma koningii as a potent inhibitor of ATP generation in the glycolytic pathway. From experiments with both cultured mouse carcinoma FM3A cells and isolated enzymes, it was shown that koningic acid is a specific inhibitor of glyceraldehyde 3-phosphate dehydrogenase that catalyzes the conversion of glyceraldehyde 3-phosphate to 3-phosphoglycerate.

A new sesquiterpene antibiotic, heptelidic acid producing organisms, fermentation, isolation and characterization.

A new sesquiterpene antibiotic, heptelidic acid, was found in the culture filtrate of three different strains of fungi isolated from soil samples. These strains were identified as Gliocladium virens, Chaetomium globosum and Trichoderma viride. Heptelidic acid was produced by conventional submerged culture and purified by successive column chromatography on silica gel and Sephadex LH-20 and finally by preparative TLC on silica gel. The molecular formula of heptelidic acid was determined as C15H20O5 on the basis of elementary analysis and high resolution mass spectrometry of its monomethyl ester. The antimicrobial spectrum of the antibiotic revealed its specific activity against anaerobic bacteria, especially against Bacteroides fragilis.

Metabolic conversions of trichothecene mycotoxins: biotransformation of 3-acetyldeoxynivalenol into fusarenon-X.

The trichothecene mycotoxin 3-acetyldeoxynivalenol was transformed by cultures of the diacetoxyscirpenol producer Fusarium sp. strain C37410-90 into four compounds, identified as deoxynivalenol, 15-acetyldeoxynivalenol, 3,15-diacetyldeoxynivalenol and fusarenon-X. The major transformations are the result of specific esterification and de-esterification processes, but the production of fusarenon-X involves in addition a novel 4 beta-hydroxylation of the trichothecene ring system itself.

[Detection of toxigenic Fusarium strains, producing T-2 toxin, in wheat grain mycoflora by microbiologic assay]. / Obnaruzhenie toksigennykh shtammov Fusarium, produtsiruiushchikh T-2 toksin, v mikoflore zerna pshenitsy mikrobiologicheskim metodom.

Twenty-three Fusarium strains were isolated from wheat grain harvested in the Moscow region. The ability of the fungi cultures isolated for producing T-2 toxin was studied by the microbiological assay with the use of Saccharomyces lactis culture (BKMU-459) susceptible to T-2 toxin. The toxigenic properties were shown by 9 cultures. Six strains with unestablished species appurtenance grown on A. Capek's agar in Perti dishes were found to produce T-2 toxin in an amount of 2 to 50 micrograms/ml agar. Three strains grown on sterilized wheat grain and attributed to Fusarium sporotrichiella v. poae according to the morphological characteristics were discovered to produce T-2 toxin in an amount from 50-100 to 400-600 micrograms/g. Production of T-2 toxin by the strains isolated was confirmed by thin-layer chromatography. Experiments made on young rats have demonstrated that extracts from F. sporotrichiella v. poae strains producing T-2 toxin appeared highly toxic for the animals.