Selection of Astaxanthin-Overproducing Mutants of Phaffia rhodozyma with beta-Ionone

beta-Ionone, an end ring analog of beta-carotene, inhibits astaxanthin production in the red yeast Phaffia rhodozyma. Astaxanthin-overproducing mutants of this yeast are easily spotted on beta-ionone-containing yeast malt agar plates. beta-Ionone appears to block astaxanthin synthesis at the beta-carotene level.     

Zooplankton composition and distribution off the coast of Galicia, Spain

During June and September 1984, zooplankton samples were collectedwith other hydrographic and biological data along the Galiciancoast (NW of Spain). In June copepods contributed {small tilde}60%to the total zooplankton community, with larvaceans, siphonophoresand cladocerans also abundant. In September >90% of the zooplanktonsampled were copepods. The dominant species of copepods in bothJune and September were Acartia clausi, Paracalanus parvus andTemora longicornis. The meroplankton was dominated by echinoderms,bryozoans, barnacle larvae and bivalve larvae. In June the averagezooplankton biomass was 31.08 mg C m^–3; the Septemberaverage was 41.69 mg C m^–3. The relationship between theslopes of the regression equations (biomass versus abundance)suggests that the zooplankton assemblage in June was composedby larger animals than in September. The major concentrationof zooplankton was between 0 and 50 m, with both June and Septemberdaytime surface samples having 6–7 times the amount oforganisms than the lower water column (50–100 m). Therewere no distinct differences in total zooplankton abundancesat the inshore and offshore stations; however, the inshore stationsoften had a higher percentage of meroplankton than the offshorestations. In June zooplankton abundance at the northern transectsand the western transects was similar. In September there weregreater concentrations of zooplankton in the western Galicianshelf as compared with the northern shelf. These differencesin the horizontal distribution of the zooplankton were relatedto upwelling events.     

Patterns of nucleotide change in mitochondrial ribosomal RNA genes and the phylogeny of piranhas

The patterns and rates of nucleotide substitution in mitochondrial ribosomal RNA genes are described and applied in a phylogenetic analysis of fishes of the subfamily Serrasalminae (Teleostei, Characiformes, Characidae). Fragments of 345 bp of the 12S and 535 bp of the 16S genes were sequenced for 37 taxa representing all but three genera in the subfamily. Secondary-structure models based on comparative sequence analysis were derived to characterize the pattern of change among paired and unpaired nucleotides, forming stem and loop regions, respectively. Base compositional biases were in the direction of A-rich loops and G-rich stems. Ninety-five percent of substitutions in stem regions were compensatory mutations, suggesting that selection for maintenance of base pairing is strong and that independence among characters cannot be assumed in phylogenetic analyses of stem characters. The relative rate of nucleotide substitution was similar in both fragments sequenced but higher in loop than in stem regions. In both genes, C-T transitions were the most common type of change, and overall transitions outnumbered transversions by a factor of two in 16S and four in 12S. Phylogenetic analysis of the mitochondrial DNA sequences suggests that a clade formed by the generaPiaractus, Colossoma, andMylossoma is the sister group to all other serrasalmins and that the generaMyleus, Serrasalmus, andPristobrycon are paraphyletic. A previous hypothesis concerning relationships for the serrasalmins, based on morphological evidence, is not supported by the molecular data. However, phylogenetic analysis of host-specific helminth parasites and cytogenetic data support the phylogeny of the Serrasalminae obtained in this study and provide evidence for coevolution between helminth parasites and their fish hosts.     

Rhizobium etli cytochrome mutants with derepressed expression of cytochrome terminal oxidases and enhanced symbiotic nitrogen accumulation

 A method to isolate mutants with derepressed expression of cytochrome oxidases and better symbiotic performance is presented. A mutant of Rhizobium etli, CFN030, isolated by its azide-resistant phenotype, was obtained by transposon Tn5-mob mutagenesis. This mutant has a derepressed expression of cytochrome aa₃, higher respiratory activities when cultured microaerobically and an improved symbiotic nitrogen fixation capacity. This phenotype was similar to the previously described mutant CFN037, which was isolated by its increased capacity to oxidize N,N,N′,N′-tetramethyl-p-phenylenediamine (TMPD) [Soberón M et al. (1990) J Bacteriol 172:1676–1680]. We show here that although both mutants have a similar symbiotic phenotype, they are affected in different genes. Strain CFN030 has the Tn5 inserted in the chromosome while in strain CFN037 the transposon was located in plasmid b. Cytochrome spectral analysis of both mutant strains in the post-exponential phase of growth, showed the expression of an additional terminal oxidase (cbb₃) that is not expressed in the wild-type strain. Received: 10 April 1995/Received revision: 21 August 1995/Accepted: 7 September1995     

Oxidative damage to sarcoplasmic reticulum Ca2+-pump induced by Fe2+/H2O2/ascorbate is not mediated by lipid peroxidation or thiol oxidation and leads to protein fragmentation

The major protein in the sarcoplasmic reticulum (SR) membrane is the Ca²⁺ transporting ATPase which carries out active Ca²⁺ pumping at the expense of ATP hydrolysis. The aim of this work was to elucidate the mechanisms by which oxidative stress induced by Fenton's reaction (Fe²⁺ + H₂O₂ HO· + OH^–+ Fe³⁺) alters the function of SR. ATP hydrolysis by both SR vesicles (SRV) and purified ATPase was inhibited in a dose-dependent manner in the presence of 0–1.5 MM H₂O₂ plus 50 M Fe²⁺ and 6 mM ascorbate. Ca²⁺ uptake carried out by the Ca²⁺-ATPase in SRV was also inhibited in parallel. The inhibition of hydrolysis and Ca²⁺ uptake was not prevented by butylhydroxytoluene (BHT) at concentrations which significantly blocked formation of thiobarbituric acid-reactive substances (TBARS), suggesting that inhibition of the ATPase was not due to lipid peroxidation of the SR membrane. In addition, dithiothreitol (DTT) did not prevent inhibition of either ATPase activity or Ca²⁺ uptake, suggesting that inhibition was not related to oxidation of ATPase thiols. The passive efflux of ⁴⁵Ca²⁺ from pre-loaded SR vesicles was greatly increased by oxidative stress and this effect could be only partially prevented (ca 20%) by addition of BHT or DTT. Trifluoperazine (which specifically binds to the Ca²⁺-ATPase, causing conformational changes in the enzyme) fully protected the ATPase activity against oxidative damage. These results suggest that the alterations in function observed upon oxidation of SRV are mainly due to direct effects on the Ca²⁺-ATPase. Electrophoretic analysis of oxidized Ca²⁺-ATPase revealed a decrease in intensity of the silver-stained 110 kDa Ca²⁺-ATPase band and the appearance of low molecular weight peptides (MW < 100 kDa) and high molecular weight protein aggregates. Presence of DTT during oxidation prevented the appearance of protein aggregates and caused a simultaneous increase in the amount of low molecular weight peptides. We propose that impairment of function of the Ca²⁺-pump may be related to aminoacid oxidation and fragmentation of the protein.Abbreviations AcP acetylphosphate - BHT butylhydroxytoluene - DTT dithiothreitol - Hepes 4-(2-hydroxyethyl)-1-piperazine-ethanesulfonic acid - SDS sodium dodecyl sulfate - SDS-PAGE polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate - SR sarcoplasmic reticulum - SRV sarcoplasmic reticulum vesicles - TBA thiobarbituric acid - TBARS thiobarbituric acid-reactive substances - TFP trifluoperazine     

Transport of acetic acid in Zygosaccharomyces bailii: effects of ethanol and their implications on the resistance of the yeast to acidic environments.

Cells of Zygosaccharomyces bailii ISA 1307 grown in a medium with acetic acid, ethanol, or glycerol as the sole carbon and energy source transported acetic acid by a saturable transport system. This system accepted propionic and formic acids but not lactic, sorbic, and benzoic acids. When the carbon source was glucose or fructose, the cells displayed activity of a mediated transport system specific for acetic acid, apparently not being able to recognize other monocarboxylic acids. In both types of cells, ethanol inhibited the transport of labelled acetic acid. The inhibition was noncompetitive, and the dependence of the maximum transport rate on the ethanol concentration was found to be exponential. These results reinforced the belief that, under the referenced growth conditions, the acid entered the cells mainly through a transporter protein. The simple diffusion of the undissociated acid appeared to contribute, with a relatively low weight, to the overall acid uptake. It was concluded that in Z. bailii, ethanol plays a protective role against the possible negative effects of acetic acid by inhibiting its transport and accumulation. Thus, the intracellular concentration of the acid could be maintained at levels lower than those expected if the acid entered the cells only by simple diffusion.