GABA-like immunoreactivity of identified interneurons in the flight system of the locust,Locusta migratoria

Summary The transmitter content of identified inhibitory interneurons in the flight system of the locust, Locusta migratoria, has been characterized using antibodies raised against protein-conjugated gamma aminobutyric acid. Identified flight neurons were filled with the fluorescent dye, Lucifer Yellow. Serial sections of dye-filled neurons were incubated with an antibody to gamma aminobutyric acid which was subsequently tagged with a fluorescent marker. Excitatory motoneurons to wing muscles and 13 flight interneurons (3 excitatory, 7 inhibitory, and 3 with unknown synaptic effect) were examined. Neither the moto-neurons nor any of the 3 excitatory interneurons contained immunoreactive material. Six of the 7 inhibitory interneurons did contain immunoreactive material. All the neurons which contained immunoreactive material and whose synaptic effect is known were inhibitory. We conclude that most of the inhibitory flight interneurons which have been described use gamma aminobutyric acid as their transmitter. Interestingly, at least 1 set of interneurons known to be inhibitory does not use gamma aminobutyric acid. We predict that the 2 interneurons which do contain immunoreactive material and whose synaptic effect is not yet known will be found to have inhibitory roles in the operation of the flight circuitry.     

Chromosomal localization of the gene for human glucosamine-6-sulphatase to 12q14

Summary Glucosamine-6-sulphatase (G6S), a lysosomal enzyme found in all cells, is involved in the catabolism of heparin, heparan sulphate, and keratan sulphate. Deficiency of G6S results in the accumulation of undegraded substrate and the lysosomal storage disorder mucopolysaccharidosis type IIID (Sanfilippo D syndrome). Regional mapping by in situ hybridization of a ³H-labelled human G6S cDNA probe to human metaphase chromosomes indicated that the G6S gene is localized to chromosome 12 at q14. The localization of the G6S gene to chromosome 12 was confirmed using the G6S cDNA clone in Southern blot hybridization analysis of DNA from human x mouse hybrid cell lines.     

Glucocorticoid regulation of enkephalins in cultured rat adrenal medulla

The effect of dexamethasone on enkephalin-containing (EC) peptide levels and preproenkephalin mRNA levels was determined in adrenal medullary explants (glands) from sham and hypophysectomized (hypox) rats. Culture for 4 days in serum-free medium without dexamethasone resulted in a 13- and 4-fold increase in EC peptide levels in sham and hypox glands, respectively. The addition of dexamethasone (10(-5) M) produced a 20- to 26-fold increase in EC peptides in sham and hypox glands. In serum free medium, hypox glands showed a concentration dependent increase in EC peptides with the ED50 for dexamethasone equal to 5.7 x 10(-7) M. Since the glucocorticoid antagonist RU486 partially blocked the rise in EC peptides in sham glands, it appears that the increase in EC peptides in sham glands in the absence of dexamethasone is a result of a higher concentration of endogenous corticosterone in sham compared to hypox glands. Dexamethasone resulted in a 6-fold increase in preproenkephalin mRNA in hypox glands cultured for 2 days. This increase was approximately proportional to the increase in EC peptides seen at 4 days. In serum free medium progesterone, testosterone, and deoxycorticosterone failed to increase EC peptides in hypox glands. These results indicate that glucocorticoid treatment is required for maximal proenkephalin gene expression and EC peptide biosynthesis in cultured glands.     

Nutrient mobility in variable- and permanent-charge soils

Variable-charge (v-c) and permanent-charge (p-c) soils differ fundamentally with regard to many nutrient-cycling processes. Variable-charge soils are more common in the tropics than in temperature zones because their formation requires desilication, which proceeds fastest in warm, moist climates. The dynamics of nutrient mobility tend to be more complex in v-c than in p-c soils. For example, theory predicts that, as pH of v-c soils decreases, cation exchange capacity (CEC) also decreases and anion exchange capacity (AEC) increases. If AEC exceeds CEC, cations such as ammonium and potassium will be more mobile than anions such as nitrate; this is the reverse of the situation in p-c soils, on which most of our knowledge of nutrient cycling is based. Variable-charge surfaces sorb phosphorus, creating plant nutritional problems throughout large areas of the humid tropics. Desilication, the same process that creates v-c surfaces, results also in stable aggregation, creating soils that retain water, yet drain rapidly and resist erosion. The Soil Taxonomy system incorporates information on mineralogy, texture, and organic matter content, and therefore provides insights into patterns of charge chemistry and nutrient cycling across a wide range of soil types.     

Effects of bovine inhibin, transforming growth factor-beta and bovine Activin-A on granulosa cell differentiation

To determine whether inhibin and its related peptides might act locally to control granulosa cell function and differentiation, the dose- and time-dependent effects of bovine inhibin, the homo-dimer of the beta-chain of bovine inhibin (Activin-A) and porcine TGF beta on rat granulosa cell aromatase activity and progesterone synthesis were investigated in vitro. TGF beta enhanced FSH-induced aromatase activity and progesterone synthesis, and accelerated the peak response for progesterone synthesis. Activin-A on the other hand, augmented FSH-induced aromatase activity while arresting progesterone synthesis, and anti-luteinization effect. By contrast, exogenous inhibin had no detectable effect on the steroidogenic potential of these cells. Thus TGF beta and Activin, unlike their similar effects on the release of FSH by the pituitary, appear to affect ovarian granulosa cell function in different fashion, under conditions where inhibin itself has no effect.     

Location and activity of ubiquinone 10 and ubiquinone analogues in model and biological membranes

Deuteriated analogues of ubiquinone 10 (Q10) have been dispersed with plasma membranes of Escherichia coli and with the inner membranes of beetroot mitochondria. Orientational order at various deuteriated sites was measured by solid-state deuterium nuclear magnetic resonance (2H NMR). Similar measurements were made, using the compounds dispersed in dimyristoylphosphatidylcholine (DMPC) and egg yolk lecithin and dispersions prepared from the lipid extracts of beetroot mitochondria. In all cases only a single unresolved 2H NMR spectrum (typically 1000-Hz full width at half-height) was observed at concentrations down to 0.02 mol % Q10 per membrane lipid. This result shows that most Q10 is in a mobile environment which is physically separate from the orientational constraints of the bilayer lipid chains. In contrast, a short-chain analogue of Q10, in which the 10 isoprene groups have been replaced by a perdeuteriated tridecyl chain, showed 2H NMR spectra with quadrupolar splittings typical of an ordered lipid that is intercalated into the bilayer. The NADH oxidase activity and O2 uptake in Escherichia coli and in mitochondria were independent of which analogue was incorporated into the membrane. Thus, despite the major difference in their physical association with membranes, or their lipid extracts, the electron transport function of the long- and short-chain ubiquinones is similar, suggesting that the bulk of the long-chain ubiquinone does not have a direct function in electron transporting activity. The physiologically active Q10 may only be a small fraction of the total ubiquinone, a fraction that is below the level of detection of the present NMR equipment.(ABSTRACT TRUNCATED AT 250 WORDS)     

A Stable Genomic Source of P Element Transposase in Drosophila Melanogaster

A single P element insert in Drosophila melanogaster, called P[ry+ delta 2-3](99B), is described that caused mobilization of other elements at unusually high frequencies, yet is itself remarkably stable. Its transposase activity is higher than that of an entire P strain, but it rarely undergoes internal deletion, excision or transposition. This element was constructed by F. Laski, D. Rio and G. Rubin for other purposes, but we have found it to be useful for experiments involving P elements. We demonstrate that together with a chromosome bearing numerous nonautonomous elements it can be used for P element mutagenesis. It can also substitute efficiently for "helper" plasmids in P element mediated transformation, and can be used to move transformed elements around the genome.     

Discontinuously synthesized mRNA from Trypanosoma brucei contains the highly methylated 5' cap structure, m7GpppA*A*C(2'-O)mU*A

Trypanosoma brucei mRNA is discontinuously synthesized via the 5' addition of a "mini-exon" sequence. The mini-exon-specific cap structure was purified from a complete RNase T2 and phosphatase digest of in vivo 32P-labeled poly(A)+RNA. The purified cap structure was sequenced by a series of partial and complete enzymatic digests by nuclease P1 and venom phosphodiesterase. This approach demonstrated that the T. brucei mini-exon cap structure consists of N7-methylguanosine linked in a conventional 5'-5' triphosphate bond to five nucleotides, in the sequence A*A*C(2'-O)mU*A (asterisks denote modifications that were not fully characterized in this work). 2'-O-methylations and other modifications appear to be present in this novel cap structure, which could have a functional role in the metabolism of the mini-exon.     

Two-dimensional NMR studies of Kazal proteinase inhibitors. 1. Sequence-specific assignments and secondary structure of turkey ovomucoid third domain

Two-dimensional proton NMR experiments have been used to sequentially assign resonances to all of the peptide backbone protons of turkey ovomucoid third domain (OMTKY3) except those of the N-terminal alpha-amino group whose signal was not resolved owing to exchange with the solvent. Assignments also have been made for more than 80% of the side-chain protons. Two-dimensional chemical shift correlated spectroscopy (COSY), relayed coherence transfer spectroscopy (RELAY), and two-dimensional homonuclear Hartmann-Hahn spectroscopy (HOHAHA) were used to identify the spin systems of almost half of the residues prior to sequential assignment. Assignments were based on two-dimensional nuclear Overhauser enhancements observed between adjacent residues. The secondary structure of OMTKY3 in solution was determined from additional assigned NOESY cross-peaks; it closely resembles the secondary structure determined by single-crystal X-ray diffraction of OMTKY3 in complex with Streptomyces griseus proteinase B [Fujinaga, M., Read, R.J., Sielecki, A., Ardelt, W., Laskowski, M., Jr., & James, M.N.G. (1982) Proc. Natl. Acad. Sci. U.S.A. 79, 4868-4872]. The NMR data provide evidence for three slowly exchanging amide protons that were not identified as hydrogen-bond donors in the crystal structure.     

Escherichia coli isocitrate lyase: properties and comparisons

The glyoxylate cycle was first discovered during studies on bacteria and fungi with the ability to grow on acetate or ethanol as the sole carbon source. Isocitrate lyase, the first enzyme unique to the glyoxylate cycle, has been studied in numerous prokaryotic and eukaryotic organisms. However, information on this enzyme from Escherichia coli is limited. We have recently reported the purification and in vitro phosphorylation of this enzyme. In the present study we have examined and characterized a variety of inhibitors, the divalent cation requirement and the amino acid composition of E. coli isocitrate lyase and compared these results to those obtained with other organisms.