Ammonium photo-production by heterocytous cyanobacteria: potentials and constraints

Over the last decades, production of microalgae and cyanobacteria has been developed for several applications, including novel foods, cosmetic ingredients and more recently biofuel. The sustainability of these promising developments can be hindered by some constraints, such as water and nutrient footprints. This review surveys data on N₂-fixing cyanobacteria for biomass production and ways to induce and improve the excretion of ammonium within cultures under aerobic conditions. The nitrogenase complex is oxygen sensitive. Nevertheless, nitrogen fixation occurs under oxic conditions due to cyanobacteria-specific characteristics. For instance, in some cyanobacteria, the vegetative cell differentiation in heterocyts provides a well-adapted anaerobic microenvironment for nitrogenase protection. Therefore, cell cultures of oxygenic cyanobacteria have been grown in laboratory and pilot photobioreactors (Dasgupta et al., 2010; Fontes et al., 1987; Moreno et al., 2003; Nayak & Das, 2013). Biomass production under diazotrophic conditions has been shown to be controlled by environmental factors such as light intensity, temperature, aeration rate, and inorganic carbon concentration, also, more specifically, by the concentration of dissolved oxygen in the culture medium. Currently, there is little information regarding the production of extracellular ammonium by heterocytous cyanobacteria. This review compares the available data on maximum ammonium concentrations and analyses the specific rate production in cultures grown as free or immobilized filamentous cyanobacteria. Extracellular production of ammonium could be coupled, as suggested by recent research on non-diazotrophic cyanobacteria, to that of other high value metabolites. There is little information available regarding the possibility for using diazotrophic cyanobacteria as cellular factories may be in regard of the constraints due to nitrogen fixation.     

Albino inflorescence proliferation of Dendrocalamus latiflorus

Summary Inflorescence proliferation is a plant tissue culture technique that, can be used to obtain in vitro inflorescences year-round without the intervening development of vegetative organs. In this study, we used albino mutant inflorescences of Dendrocalamus latiflorus as the original explant material to investigate, the effect of plant growth regulators on long-term inflorescence proliferation. The albino inflorescences proliferated on solidified Murashige and Skoog (MS) basal medium supplemented with thidiazuron (TDZ), and the optimal concentration for successful long-term inflorescence proliferation was 0.45 μM TDZ. A combination of α-naphthaleneacetic acid (NAA) with 0.45 μM TDZ inhibited the inflorescence proliferation. Inflorescences cultured on a TDZ-free medium supplemented with 26.82 μM NAA rooted in 21 d, vegetative shoots formed by 42 d and, in one case, flowering occurred after 63 d. The auxins 2,4-dichlorophenoxyacetic acid (2,4-D, 4.52 μM) and pieloram (4.14 μM) induced shoot formation. The protocol described can be used to produce large numbers of mutant inflorescences within a relatively short period of time.     

In vitro propagation of <Emphasis Type="Italic">Helleborus</Emphasis> species

A general in vitro cloning system was established for four Helleborus species: H. argutifolius, H. foetidus, H. niger and H. orientalis. The plant material was introduced in vitro from axillary buds. A Murashige and Skoog (MS)—based medium (Murashige and Skoog 1962) was used supplemented with 2% (w/v) sucrose, 2-isopentenyladenine (2-iP) and 6-benzylaminopurine (BA). Multiplication rates depended on the genotype and varied from 1.3 for H. foetidus till 3.8 for H. niger. The first results showed that the rooting phase could be done ex vitro. Rooting was induced by a drench for one week in a solution of indole-3-butyric acid (IBA -3 mg l⁻¹) and 1-naphthaleneacetic acid (NAA-1 mg l⁻¹) at 5°C.     

Unbalanced ribosome assembly in Saccharomyces cerevisiae expressing mutant 5 S rRNAs.

Mutant 5 S rRNA genes were expressed in Saccharomyces cerevisiae to further define the function of the ribosomal 5 S RNA. RNA synthesis and utilization were assayed using previously constructed markers which have been shown to be functionally neutral and easily detected by gel electrophoresis. Most mutations were found not to affect the growth rate because they were poorly expressed or could be accommodated effectively in the ribosomal structure. Two of the mutants, Y5A99U56U57 and Y5U90i5 adversely affected cell growth as well as protein synthesis in vitro. Polyribosome profiles in both of these mutants were substantially shorter, and an analysis of the ribosomal subunit composition revealed a significant imbalance with a 25-35% excess in 40 S subunits. Kinetic analyses of RNA labeling indicated very low cellular levels of mutant RNA either because it was poorly expressed (Y5U90i5) or rapidly degraded before being incorporated into mature 60 subunits (Y5A99U56U57). The results suggest that the 5 S RNA is required for the assembly of stable ribosomal 60 S subunits and raise the possibility that this RNA or, more likely, its corresponding ribonucleoprotein complex is critical for subunit assembly or even RNA processing.     

Calcium-dependent binding of EDTA-extractable proteins to calf lens fiber membrane structures

Calf lens fiber membranes and fractions enriched in junction-like structures have been isolated in the absence and presence of EDTA. Their biochemical features have been studied. Sodium dodecyl sulfate polyacrylamide gel electrophoresis and immunoblotting experiments have provided evidence that a distinct group of EDTA-extractable proteins, being one of the main protein components of calf lens fiber membranes and very likely also of junction-like structures, is bound to these membranes via calcium ions. In addition to these proteins, four polypeptides with apparent molecular weights between 14 000 and 17 000 are characteristic for detergent-insoluble lens fiber structures prepared in calcium-rich medium. The absence of EDTA-extractable proteins in the urea-soluble calcium-containing fraction implies that they are not components of the cytoskeleton and that the calcium-dependent binding of these proteins to the membrane is urea-resistant. The use of EDTA throughout the whole membrane isolation procedure results in their complete removal from the membranes which already starts during buffer washing. This indicates that EDTA-extractable proteins exclusively consist of extrinsic membrane proteins which probably are not involved in cytoskeleton binding.     

Alkyl-dihydroxyacetone phosphate synthase and dihydroxyacetone phosphate acyltransferase form a protein complex in peroxisomes.

Dihydroxyacetone phosphate (GrnP) acyltransferase and alkyl-GrnP synthase are the key enzymes involved in the biosynthesis of ether phospholipids. Both enzymes are located on the inside of the peroxisomal membrane. Here we report evidence for a direct interaction between these enzymes obtained by the use of chemical cross-linking. After cross-linking and immunoblot analysis alkyl-GrnP synthase could be detected in a 210-kDa complex which was located entirely on the lumenal side of the peroxisomal membrane. Two-dimensional SDS/PAGE demonstrated that GrnP-acyltransferase is also cross-linked in a 210-kDa complex. Co-immunoprecipitation confirmed that the two enzymes interact, in a heterotrimeric complex. Furthermore, alkyl-GrnP synthase can form a homotrimeric complex in the absence of GrnP-acyltransferase as was demonstrated by immunoblot analysis after cross-linking experiments with either GrnP-acyltransferase deficient human fibroblast homogenates or recombinant (His)6-tagged alkyl-GrnP synthase. We conclude that alkyl-GrnP synthase interacts selectively with GrnP-acyltransferase in a heterotrimeric complex and in the absence of GrnP-acyltransferase can also form a homotrimeric complex.     

Mitochondrial heredity of resistance to 3-(3,4-dichlorophenyl)-1,1-dimethylurea, an inhibitor of cytochrome b oxidation, in Saccharomyces cerevisiae.

3-(3,4-dichlorophenyl)-1,1-dimethylurea (diuron), an inhibitor of cytochrome b oxidation, has been used for the selection of three resistant mutants (diur) of Saccharomyces cerevisiae. The mutant diur-64 exhibits in vivo cross-resistance to antimycin A while diur-34 and diur-1 are more sensitive to antimycin A than the parental strain. The three mutants exhibit mitochondrial inheritance according to the following criteria: mitotic segregation of diuron-resistant and diuron-sensitive diploids is obtained among the diploid progeny of a cross between diur and dius; non-Mendelian segregation of diuron resistance (4:0) is observed in spores of tetrads issued from diuron-resistant diploid; extensive ethidium bromide treatment leads to the formation of Q- mutants which no longer transmit diur and dius alleles. Evidence for two distinct diuron-resistant loci were obtained by allelism tests. Recombination analysis shows that diuron-resistance is not located in the polar region of the mitochondrial genome. The diur loci are not linked to the erythromycin locus since the upper limit in recombinants frequency (26%) for a non-polar region is obtained between diur and eryr. A low recombinants frequency (3%) is observed in crosses between diur-34 mutation and the two mutants cob1 and cob2 suggesting that diur-34 might be located between these two cytochrome-b-deficient loci. The resistance to diuron is also expressed in vitro since the oxidation rates of succinate by sonicated submitochondrial particles from the mutants are clearly less sensitive to diuron than that of the wild type.     

Resistance of Anhydrobiotic Aphelenchus avenae to Methyl Bromide Fumigation

The effect of methyl bromide (MB) was tested on active and anhydrobiotic Aphelenchus avenae. A. avenae was induced into anhydrobiosis by three different techniques. Both active and anhydrobiotic nematodes were subjected to 3,000 μ1 MB/liter air for 14 periods from 0 to 82 h. Anhydrobiotic nematodes were more resistant to fumigation than active nematodes, regardless of the technique used to induce anhydrobiosis. The percent survival decreased with increasing MB exposures (μ1 MB × h). For an LD₉₅ of 45,000-54,000 μ1/1 × h were required for active nematodes and >279,000 μ1/1 × h for anhydrobiotic nematodes.