Glutathione Turnover in Cultured Astrocytes: Studies with [15N]Glutamate

The incorporation of [15N]glutamic acid into glutathione was studied in primary cultures of astrocytes. Turnover of the intracellular glutathione pool was rapid, attaining a steady state value of 30.0 atom% excess in 180 min. The intracellular glutathione concentration was high (20-40 nmol/mg protein) and the tripeptide was released rapidly into the incubation medium. Although labeling of glutathione (atom% excess) with [15N]glutamate occurred rapidly, little accumulation of 15N in glutathione was noted during the incubation compared with 15N in aspartate, glutamine, and alanine. Glutathione turnover was stimulated by incubating the astrocytes with diethylmaleate, an electrophile that caused a partial depletion of the glutathione pool(s). Diethylmaleate treatment also was associated with significant reductions of intraastrocytic glutamate, glycine, and cysteine, i.e., the constituents of glutathione. Glutathione synthesis could be stimulated by supplementing the steady-state incubation medium with 0.05 mM L-cysteine, such treatment again partially depleting intraastrocytic glutamate and causing significant reductions of 15N labeling of both alanine and glutamine, suggesting that glutamate had been diverted from the synthesis of these amino acids and toward the formation of glutathione. The current study underscores both the intensity of glutathione turnover in astrocytes and the relationship of this turnover to the metabolism of glutamate and other amino acids.     

Nash equilibrium and evolutionary stability in large- and finite-population "playing the field" models

This paper studies the correspondence between Nash equilibrium and evolutionary stability in large- and finite-population "playing the field" models. Whenever the fitness function is sufficiently continuous, any large-population ESS corresponds to a symmetric Nash equilibrium in the game that describes the simultaneous interaction of the individuals in the population, and any strict, symmetric Nash equilibrium in that game corresponds to a large-population ESS. This correspondence continues to hold, approximately, in finite populations; and it holds exactly for strict pure-strategy equilibria in sufficiently large finite populations. By contrast, a sequence of (mixed-strategy) finite-population ESSs can converge, as the population grows, to a limit that is not a large-population ESS, and a large-population ESS need not be the limit of any sequence of finite-population ESSs.     

Differential rates of secular increase of five major primary abilities

A number of studies have shown that mean intelligence levels have been increasing at a rate of around 3 IQ points per decade during the last half-century. This paper presents data for cohorts of 9-11 year olds in Northern Ireland tested in 1978 and 1988 and shows that the rates of increase differ markedly for different primary abilities. Verbal comprehension has shown virtually no increase, while there have been large increases in spatial relations and perceptual speed. Moderate increases have taken place in the numerical and reasoning primaries. The results are interpreted as supporting a nutrition theory of the secular increases in intelligence.     

Cellulose and lignin degradation in forest soils: Response to moisture,temperature, and acidity

The concentration of lignin in plant tissue is a major factor controlling organic matter degradation rates in forest ecosystems. Microbial biomass and lignin and cellulose decomposition were measured for six weeks in forest soil microcosms in order to determine the influence of pH, moisture, and temperature on organic matter decomposition. Microbial biomass was determined by chloroform fumigation; lignin and cellulose decomposition were measured radiometrically. The experiment was designed as a Latin square with soils of pH of 4.5, 5.5, and 6.5 adjusted to 20, 40, or 60% moisture content, and incubated at temperatures of 4, 12, or 24°C. Microbial biomass and lignin and cellulose decomposition were not significantly affected by soil acidity. Microbial biomass was greater at higher soil moisture contents. Lignin and cellulose decomposition significantly increased at higher soil temperatures and moisture contents. Soil moisture was more important in affecting microbial biomass than either soil temperature or soil pH.     

Bioremediation of soils contaminated with the herbicide 2-sec-butyl-4,6-dinitrophenol (dinoseb).

A novel soil treatment method for achieving the removal of dinoseb (2-sec-butyl-4,6-dinitrophenol) from contaminated soils was investigated. One soil contained dinoseb as the major contaminant, although several other hazardous compounds were also present. A second soil was highly contaminated with dinoseb. Dinoseb was not degraded in these soils under the aerobic conditions at each site. Pretreatment of the soils by the addition of a starchy potato-processing by-product and flooding with phosphate buffer stimulated the consumption of oxygen and nitrate from the soils, thereby lowering the redox potential and creating anaerobic conditions. Anaerobiosis (Eh less than -200 mV) promoted the establishment of an anaerobic microbial consortium that degraded dinoseb completely, without the formation of the polymerization products seen under aerobic or microaerophilic conditions. When dinoseb was present at low concentrations in a chronically contaminated soil, the natural microflora was capable of establishing anaerobic conditions and degrading dinoseb as a result of starch degradation. Inoculation of this soil with an aerobic starch-degrading microorganism and then an acclimated, anaerobic, dinoseb-degrading consortium did not improve dinoseb degradation. In a second acutely contaminated soil, these inoculations improved dinoseb degradation rates over those of uninoculated controls.     

Comparison of extracellular peroxidase- and esterase-deficient mutants of Streptomyces viridosporus T7A.

Peroxidase-deficient mutants of the lignin-degrading bacterium Streptomyces viridosporus T7A were screened for their production of acid-precipitable polymeric lignin, extracellular peroxidases and esterases, and immunoreactivities against a polyclonal antibody produced against electrophoretically purified peroxidase isoform P3 of wild-type S. viridosporus. The mutants showed diminished abilities to solubilize lignin and produce acid-precipitable polymeric lignin. Their peroxidase activities were decreased, and their esterase production patterns were altered. Western immunoblots demonstrated that the mutants produced proteins immunologically reactive with the antibody, but with different mobilities from those of wild-type proteins. These findings confirm a direct role for peroxidases in lignin solubilization. They also indicate a possible role for esterases.     

Cloning and expression of a lignin peroxidase gene from Streptomyces viridosporus in Streptomyces lividans.

A lignin peroxidase gene was cloned from Streptomyces viridosporus T7A into Streptomyces lividans TK64 in plasmid pIJ702. BglII-digested genomic DNA (4-10 kb) of S. viridosporus was shotgun-cloned into S. lividans after insertion into the melanin (mel+) gene of pIJ702. Transformants expressing pIJ702 with insert DNA were selected based upon the appearance of thiostrepton resistant (tsrr)/mel-colonies on regeneration medium. Lignin peroxidase-expressing clones were isolated from this population by screening of transformants on a tsr-poly B-411 dye agar medium. In the presence of H2O2 excreted by S. lividans, colonies of lignin peroxidase-expressing clones decolorized the dye. Among 1000 transformants screened, 2 dye-decolorizing clones were found. One, pIJ702/TK64.1 (TK64.1), was further characterized. TK64.1 expressed significant extracellular 2,4-dichlorophenol (2.4-DCP) peroxidase activity (= assay for S. viridosporus lignin peroxidase). Under the cultural conditions employed, plasmidless S. lividans TK64 had a low background level of 2.4-DCP oxidizing activity. TK64.1 excreted an extracellular peroxidase not observed in S. lividans TK64, but similar to S. viridosporus lignin peroxidase ALip-P3, as shown by activity stain assays on nondenaturing polyacrylamide gels. The gene was located on a 4 kb fragment of S. viridosporus genomic DNA. When peroxidase-encoding plasmid, pIJ702.LP, was purified and used to transform three different S. lividans strains (TK64, TK23, TK24), all transformants tested decolorized poly B-411. When grown on lignocellulose in solid state processes, genetically engineered S. lividans TK64.1 degraded the lignocellulose slightly better than did S. lividans TK64. This is the first report of the cloning of a bacterial gene coding for a lignin-degrading enzyme.     

The translational stop signal: Codon with a context, or extended factor recognition element?

Wide ranging studies of the readthrough of translational stop codons within the last 25 years have suggested that the stop codon might be only part of the molecular signature for recognition of the termination signal. Such studies do not distinguish between effects on suppression and effects on termination, and so we have used a number of different approaches to deduce whether the stop signal is a codon with a context or an extended factor recognition element. A data base of natural termination sites from a wide range of organisms (148 organisms, 40000 sequences) shows a very marked bias in the bases surrounding the stop codon in the genes for all organisms examined, with the most dramatic bias in the base following the codon (+4). The nature of this base determines the efficiency of the stop signal in vivo, and in Escherichia coli this is reinforced by overexpressing the stimulatory factor, release factor-3. Strong signals, defined by their high relative rates of selecting the decoding release factors, are enhanced whereas weak signals respond relatively poorly. Site-directed cross-linking from the +1, and bases up to +6 but not beyond make close contact with the bacterial release factor-2. The translational stop signal is deduced to be an extended factor recognition sequence with a core element, rather than simply a factor recognition triplet codon influenced by context.