Microalgal lipids biochemistry and biotechnological perspectives

In the last few years, there has been an intense interest in using microalgal lipids in food, chemical and pharmaceutical industries and cosmetology, while a noteworthy research has been performed focusing on all aspects of microalgal lipid production. This includes basic research on the pathways of solar energy conversion and on lipid biosynthesis and catabolism, and applied research dealing with the various biological and technical bottlenecks of the lipid production process. In here, we review the current knowledge in microalgal lipids with respect to their metabolism and various biotechnological applications, and we discuss potential future perspectives.     

Boron bridging of rhamnogalacturonan‐II,monitored by gel electrophoresis,occurs during polysaccharide synthesis and secretion but not post‐secretion

The cell‐wall pectic domain rhamnogalacturonan‐II (RG‐II) is cross‐linked via borate diester bridges, which influence the expansion, thickness and porosity of the wall. Previously, little was known about the mechanism or subcellular site of this cross‐linking. Using polyacrylamide gel electrophoresis (PAGE) to separate monomeric from dimeric (boron‐bridged) RG‐II, we confirmed that Pb²⁺ promotes H₃BO₃‐dependent dimerisation in vitro. H₃BO₃ concentrations as high as 50 mm did not prevent cross‐linking. For in‐vivo experiments, we successfully cultured ‘Paul's Scarlet’ rose (Rosa sp.) cells in boron‐free medium: their wall‐bound pectin contained monomeric RG‐II domains but no detectable dimers. Thus pectins containing RG‐II domains can be held in the wall other than via boron bridges. Re‐addition of H₃BO₃ to 3.3 μm triggered a gradual appearance of RG‐II dimer over 24 h but without detectable loss of existing monomers, suggesting that only newly synthesised RG‐II was amenable to boron bridging. In agreement with this, Rosa cultures whose polysaccharide biosynthetic machinery had been compromised (by carbon starvation, respiratory inhibitors, anaerobiosis, freezing or boiling) lost the ability to generate RG‐II dimers. We conclude that RG‐II normally becomes boron‐bridged during synthesis or secretion but not post‐secretion. Supporting this conclusion, exogenous [³H]RG‐II was neither dimerised in the medium nor cross‐linked to existing wall‐associated RG‐II domains when added to Rosa cultures. In conclusion, in cultured Rosa cells RG‐II domains have a brief window of opportunity for boron‐bridging intraprotoplasmically or during secretion, but secretion into the apoplast is a point of no return beyond which additional boron‐bridging does not readily occur.     

Drosophila oogenesis as a bio-marker responding to EMF sources

The model biological organisms Drosophila melanogaster and Drosophila virilis have been utilized to assess effects on apoptotic cell death of follicles during oogenesis and reproductive capacity (fecundity) decline. A total of 280 different experiments were performed using newly emerged flies exposed for short time daily for 3–7?d to various EMF sources including: GSM 900/1800?MHz mobile phone, 1880–1900?MHz DECT wireless base, DECT wireless handset, mobile phone-DECT handset combination, 2.44?GHz wireless network (Wi-Fi), 2.44?GHz blue tooth, 92.8?MHz FM generator, 27.15?MHz baby monitor, 900?MHz CW RF generator and microwave oven’s 2.44?GHz RF and magnetic field components. Mobile phone was used as a reference exposure system for evaluating factors considered very important in dosimetry extending our published work with D. melanogaster to the insect D. virilis. Distance from the emitting source, the exposure duration and the repeatability were examined. All EMF sources used created statistically significant effects regarding fecundity and cell death-apoptosis induction, even at very low intensity levels (0.3?V/m blue tooth radiation), well below ICNIRP’s guidelines, suggesting that Drosophila oogenesis system is suitable to be used as a biomarker for exploring potential EMF bioactivity. Also, there is no linear cumulative effect when increasing the duration of exposure or using one EMF source after the other (i.e. mobile phone and DECT handset) at the specific conditions used. The role of the average versus the peak E-field values as measured by spectrum analyzers on the final effects is discussed.     

Mutations in the terminase genes of bacteriophage λ that bypass the necessity for FI

DNA maturation in bacteriophage λ is the process by which the concatemeric precursor DNA is cleaved at sites called cos to generate mature λ DNA molecules. These DNA molecules are then packaged into procapsids, the empty capsid precursors. The enzyme that catalyses these events is λ DNA terminase. It is composed of two subunits, made of 181 and 641 amino acids, the products of genes Nu1 and A, respectively. The product of the FI gene (gpFI ) stimulates the formation of an intermediate in capsid assembly called complex II, which contains a procapsid, terminase and DNA. The mechanism of stimulation remains unknown. It has been suggested that gpFI may also stimulate terminase-mediated cos cleavage, in the absence of procapsids, by increasing enzyme turnover. Mutants in FI fail to mature and package DNA but, in comparison with other capsid gene mutants, FI mutants are leaky. Second site mutants of FI phages, called ‘fin’ (for FI independence), bypass the necessity for gpFI. These mutants were originally localized to the region of Nu1 and A and are of two classes: finA includes those that induce the synthesis of fourfold more gene A product (gpA ) than wild-type phages, and finB includes those that produce normal amounts of gpA. Whereas all finA mutants analysed map to Nu1, finB mutants have been found both in E and in Nu1. The existence of E mutants able to bypass the necessity for gpFI in vivo shows that gpE and gpFI interact, directly or indirectly. Here we have analysed and sequenced two finA mutants and one finB mutant. All of these map in Nu1. Of the two finA mutants, one corresponds to an Ala163Ser change and the other is a silent mutation. It is likely that the finA mutations alter mRNA conformation in a manner that results in an increase in the efficiency of A mRNA translation. The fourfold increase in gpA synthesis translates into a 10-fold increase in terminase activity. These results show that terminase overproduction is sufficient to bypass the necessity for gpFI and that such an overproduction can be achieved by changes in the efficiency of translation of A due to subtle changes in the sequence upstream of the gene. The finBcs₁₀₃ mutation is a His-87→Tyr change in Nu1. Therefore, an alternative way in which to bypass the requirement for gpFI involves an alteration in the structure of gpNu1. It is likely that the altered gpNu1 would increase cleavage and packaging efficiency directly or indirectly. We have determined that DNA cleavage in vivo does not occur in the absence of gpFI. Therefore it seems that gpFI somehow facilitates an otherwise latent capacity of terminase to autoactivate its nucleolytic activity.     

Study of the antioxidant activity of Thymus sibthorpii Bentham (Lamiaceae)

Abstract

From the aerial parts of Thymus sibthorpii Bentham (Lamiaceae), five flavonoids apigenin (1), 7-methoxy-apigenin (2), naringenin (3), eriodictyol (4) and eriodictyol-7-glucoside (5), have been isolated together with caffeic acid methyl ester (6), rosmarinic acid (7) and rosmarinic acid methyl ester (8). The structures of the isolated compounds were established by spectroscopic methods. The extracts and the isolated compounds were tested for their free radical scavenging activity using the following in vitro assays: (i) interaction with the free stable radical of DPPH (1,1-diphenyl-2-picrylhydrazyl), (ii) inhibition of linoleic acid lipid peroxidation induced by the dihydrochloric acid of 2,2-azobis-2-amidinepropane (AAPH) and (iii) the scavenging activity of enzymatically produced superoxide anion. Their inhibitory activity toward soybean lipoxygenase was evaluated in vitro, using linoleic acid as a substrate. The antioxidant results of the extracts are discussed in terms of their constitution in phenolic compounds, which were determined following the Folin–Ciocalteu method.     

Simple chalcones and bis-chalcones ethers as possible pleiotropic agents

The synthesis, the antioxidative properties and the lipoxygenase (LOX) and acetylcholinesterase (AChE) inhibition of a number of 4-hydroxy-chalcones diversely substituted as well as of a series of bis-chalcones ether derivatives are reported. The chalcones derivatives were readily produced using a Claisen–Schmidt condensation in a ultra sound bath in good yields. The structures of the synthesized compounds were confirmed by spectral and elemental analysis. Their lipophilicity is experimentally determined by reversed-phase thin-layer chromatography method. Most of them are potent in vitro inhibitors of lipid peroxidation and of LOX. Compounds b2 and b3 were found to be the most potent LOX and AChE inhibitors among the tested derivatives with a significant anti-lipid peroxidation profile. The results led us to propose these enone derivatives as new multifunctional compounds against Alzheimer's disease. The results are discussed in terms of structural and physicochemical characteristics of the compounds. Moreover, the pharmacokinetic profile of these compounds was investigated using computational methods.     

Early and late passage C-6 glial cell growth: Similarities with primary glial cells in culture

Earlier studies in our laboratory have shown that C-6 glial cells in culture exhibit astrocytic properties with increasing cell passage. In this study, we tested the responsiveness of early and late passage C-6 glial cells to various cultures conditions: culture substrata (collagen, poly-L-lysine, plastic), or supplements for the culture medium, DMEM, [fetal calf, or heat inactivated (HI) serum, or media conditioned from mouse neuroblastoma cells (NBCM) or primary chick embryo cultured neurons (NCM)]. Glutamine synthetase (GS) and cyclic nucleotide phosphohydrolase (CNP), astrocytic and oligodendrocytic glial markers, were used. Cell numer and protein content increased exponentially with days in culture regardless of the type of the substratum or cell passage. Differences in cell morphology among the three types of substratum were also reflected on GS activity, which rose by three-fold on culture day 3 for cells grown on collagen; thereafter, GS profiles were similar for all substrata. This early rise in GS is interpreted to reflect differential cell adhesion processes on the substrata; specifically, cell adhesion on the collagen stimulated differentiation into astrocytic phenotype.Analogous to immature glia cells in primary cultures, early passage C-6 glial cells responded to neuronal factors supplied either from NCM or NBCM by expressing reduced GS activity, the astrocytic marker and enhanced CNP activity, the oligodendrocytic marker. Thus, early passage cells can be induced to express either astrocytic or oligodendrocytic phenotype. In accordance with our previous reports on primary glial cells, late passage C-6 cells exhibit their usual astrocytic behavior, responding to serum factors with GS activity. Moreover, whereas NCM or NBCM alone markedly lowered GS activity, a combination with serum restored activity. The present findings confirm our previous observations and further establish the C-6 glial cells as a reliable model to study immature glia.Special issue dedicated to Dr. Paola S. Timiras.     

Epigenetic chromatin modifiers in barley: IV. The study of barley Polycomb group (PcG) genes during seed development and in response to external ABA

Background  

Epigenetic phenomena have been associated with the regulation of active and silent chromatin states achieved by modifications of chromatin structure through DNA methylation, and histone post-translational modifications. The latter is accomplished, in part, through the action of PcG (Polycomb group) protein complexes which methylate nucleosomal histone tails at specific sites, ultimately leading to chromatin compaction and gene silencing. Different PcG complex variants operating during different developmental stages have been described in plants. In particular, the so-called FIE/MEA/FIS2 complex governs the expression of genes important in embryo and endosperm development in Arabidopsis. In our effort to understand the epigenetic mechanisms regulating seed development in barley (Hordeum vulgare), an agronomically important monocot plant cultivated for its endosperm, we set out to characterize the genes encoding barley PcG proteins.