Motohide Aoki

Size-dependent toxicity of silica nano-particles to Chlorella kessleri

SiO2 nano-particles were found to exhibit size-dependent toxicity toward the alga, Chlorella kessleri. Small SiO2 nano-particles exhibit stronger toxicity: 50% inhibitory concentrations (IC50) value for 5 nm = 0.8 ± 0.6%, 26 nm = 7.1 ± 2.8%, and 78 nm = 9.1 ± 4.7%. Enlargement of the cell body was observed by flow cytometry, which is due to the presence of structures that obstructed cell division. Optical and transmission microscopes were used to observe coagulated cells with incomplete division. Although the physiological effect of SiO2 nano-particles was not clear, SiO2 nano-particles are toxic, at least for algae in aquatic media. Under the transmission electron microscope, several amorphous structures appeared in the cells that were exposed to 5-nm silica nano-particles.

Involvement of sulfoquinovosyl diacylglycerol in DNA synthesis in Synechocystis sp. PCC 6803

BackgroundSulfoquinovosyl diacylglycerol (SQDG) is present in the membranes of cyanobacteria and their postulated progeny, plastids, in plants. A cyanobacterium, Synechocystis sp. PCC 6803, requires SQDG for growth: its mutant (SD1) with the sqdB gene for SQDG synthesis disrupted can grow with external supplementation of SQDG. However, upon removal of SQDG from the medium, its growth is retarded, with a decrease in the cellular content of SQDG throughout cell division, and finally ceases. Concomitantly with the decrease in SQDG, the maximal activity of photosynthesis at high-light intensity is repressed by 40%.FindingsWe investigated effects of SQDG-defect on physiological aspects in Synechocystis with the use of SD1. SD1 cells defective in SQDG exhibited normal photosynthesis at low-light intensity as on culturing. Meanwhile, SD1 cells defective in SQDG were impaired in light-activated heterotrophic growth as well as in photoautotrophic growth. Flow cytometric analysis of the photoautotrophically growing cells gave similar cell size histograms for the wild type and SD1 supplemented with SQDG. However, the profile of SD1 defective in SQDG changed such that large part of the cell population was increased in size. Of particular interest was the microscopic observation that the mitotic index, i.e., population of dumbbell-like cells with a septum, increased from 14 to 29% in the SD1 culture without SQDG. Flow cytometric analysis also showed that the enlarged cells of SD1 defective in SQDG contained high levels of Chl, however, the DNA content was low.ConclusionsOur experiments strongly support the idea that photosynthesis is not the limiting factor for the growth of SD1 defective in SQDG, and that SQDG is responsible for some physiologically fundamental process common to both photoautotrophic and light-activated heterotrophic growth. Our findings suggest that the SQDG-defect allows construction of the photosynthetic machinery at an elevated level for an increase in cell mass, but represses DNA synthesis. SQDG may be essential for normal replication of chromosomal DNA for completion of the cell cycle.

Profiling of lipid and glycogen accumulations under different growth conditions in the sulfothermophilic red alga Galdieria sulphuraria

The unicellular red alga Galdieria sulphuraria grows efficiently and produces a large amount of biomass in acidic conditions at high temperatures. It has great potential to produce biofuels and other beneficial compounds without becoming contaminated with other organisms. In G. sulphuraria, biomass measurements and glycogen and lipid analyses demonstrated that the amounts and compositions of glycogen and lipids differed when cells were grown under autotrophic, mixotrophic, and heterotrophic conditions. Maximum biomass production was obtained in the mixotrophic culture. High amounts of glycogen were obtained in the mixotrophic cultures, while the amounts of neutral lipids were similar between mixotrophic and heterotrophic cultures. The amounts of neutral lipids were highest in red algae, including thermophiles. Glycogen structure and fatty acids compositions largely depended on the growth conditions.

Physiological and Photosynthetic ToxiCity of Thallium in Synechocystis sp. PCC6803

The physiological and photosynthetic toxiCity mechanism of monovalent thallium, Tl(I), in a cyanobacteria, Synechocystis sp. PCC6803, was examined based on a series of batch culture experiments, determination of the pigments content and measurements of photosynthetic activities under the metal exposure conditions. Results showed that micro-molar level of Tl(I) drastically inhibit its growth, then 50% inhibitory concentration (IC50) was approximately 1 μM. An acclimating incubation with 0.5 μM Tl(I) for 72 h bring no significant changes in IC50 of thallium for growth. Chlorophyll a and phycobiliproteins content per cell basis decreased by 71% and 94% during 72 h incubation with 2.5 μM Tl(I), respectively. Results from pigments determination suggested that metabolic defect was rose by thallium exposure in Synechocystis sp. PCC6803. Then, to investigate the effect of thallium on energy generation process, acute dose-response of Tl(I) on photosynthetic O2 evolution activities were measured. No effect on net photosynthetic O2 evolution activity per chlorophyll basis was observed in 1 mM and below Tl(I) exposure, while 20 mM Tl(I) decrease the activity by 60%. Furthermore, 20 mM thallium did not affect 1,4-benzoquinone dependent PSII activity. These photosynthetic 50% inhibitory doses of thallium were approximately 2,000-fold higher than IC50 of growth. Thus, photosynthetic energy metabolism did not constitute a limiting factor of growth under the thallium exposure. These results suggested that substance metabolic defect and/or NADP reducing processes could be the main process involved in thallium toxiCity in Synechocystis sp. PCC6803.

Electrochemically active bacteria sense electrode potentials for regulating catabolic pathways

Electrochemically active bacteria (EAB) receive considerable attention for their utility in bioelectrochemical processes. Although electrode potentials are known to affect the metabolic activity of EAB, it is unclear whether EAB are able to sense and respond to electrode potentials. Here, we show that, in the presence of a high-potential electrode, a model EAB Shewanella oneidensis MR-1 can utilize NADH-dependent catabolic pathways and a background formate-dependent pathway to achieve high growth yield. We also show that an Arc regulatory system is involved in sensing electrode potentials and regulating the expression of catabolic genes, including those for NADH dehydrogenase. We suggest that these findings may facilitate the use of EAB in biotechnological processes and offer the molecular bases for their ecological strategies in natural habitats.Whether electrochemically active bacteria (EAB) can gain energy according to electrode potentials is still unclear. Here, the authors show through transcriptome and deletion mutant analyses that EAB can sense electrode potentials by the Arc system and activate NADH-dependent catabolic pathway to generate ATP.

Thallium Induces Morphological Changes in the Photosynthetic Apparatus of Synechocystis sp. PCC6803

The aim of this study was to elucidate the mechanism of thallium (Tl) ion toxicity in photosynthetic organisms. The physiological and biochemical responses to Tl exposure were analyzed in the cyanobacterium Synechocystis sp. PCC6803, which is a widely used model to study photosynthesis. We examined the photosynthetic activities of Tl+-exposed cells, the extent of Tl accumulation, and the properties of membrane lipids. Exposure to Tl+ at 2.0 and 5.0 for 24 h decreased the net photosynthetic activities of cells to 92% and 34%, respectively. After exposure to 2.5 μM Tl+, cells concentrated the Tl to 20.8 μM on a packed cell volume basis. Exposure of Synechocystis to 0–2.5 μM Tl+ resulted in an approximately 9-fold concentration factor. Treatment with 2.0 μM Tl+ for 48 h decreased the total lipid content of the cells by 38%. Further, we observed the ultrastructure of cells treated with Tl+. The cells exposed to 5 μM Tl+ for 24 h showed thylakoid membrane fragmentation and generated less-dense particles following osmium staining. During this time, the net photosynthetic oxygen evolution of the cells was reduced to 34%. These results suggest that the accumulation of Tl in cells affects the integrity of the photosynthetic apparatus.

Comprehensive Element Analysis of Prokaryotic and Eukaryotic Cells as well as Organelles by ICP-MS

Elemental abundances of prokaryotic and eukaryotic cells as well as organelles were provided as the number of atoms per cell (or organelle). In order to examine the specific uptake of vital elements by cells, E. coli cells were grown in different media and harvested at several time points during the cell growth and proliferation. The major-to-ultratrace elements in the cells were determined by micro-flow injection (μFI)-ICP-MS after microwave-assisted acid digestion of less than 10 mg of dry weight sample. Cyanobacteria and chlorella were also cultivated to compare elemental abundance differences among species. In addition, to further examine the localization and utilization of vital elements in the cells, chloroplasts and mitochondria were isolated from the tissues of spinach leaves, potato tuber, and bovine liver by density-gradient centrifugation using Percoll. The concentration data obtained by ICP-MS measurements were normalized to a per cell (or organelle) basis, taking into account the cell size and water content. Among about 60 elements detectable by ICP-MS, more than 30 elements were quantitatively determined, and it was clearly found that essential elements participating in specific vital functions such as redox reactions were highly accumulated in the microorganisms and organelles.

Effects of derivatization on solar-induced decomposition of polycyclic aromatic hydrocarbons in aqueous media

The solar-induced decomposition of 10 polycyclic aromatic hydrocarbons (PAHs) was observed in aqueous media. All 10 PAHs observed were half-decomposed within 120 min. Among anthracene derivatives, the decomposition rates were: anthracene = 1-methylanthracene < 2-methylanthracene < 9-methylanthracene < 9,10-dimethylanthracene ∼ 2-aminoanthracene. The addition of commercial humic acid had no effect on the decomposition rates of these PAHs. Deuterium water also hastened the decomposition of PAH. The products obtained by the solar radiation of PAH after extraction to DCM were mainly ketone and hydroxyl derivatives. To explaine these results, reactivities and electron charges at the constituent carbon atoms in each anthracene derivative were examined by an ab initio molecular orbital calculation method.

Short-lived long non-coding RNAs as surrogate indicators for chemical exposure and LINC00152 and MALAT1 modulate their neighboring genes

Whole transcriptome analyses have revealed a large number of novel long non-coding RNAs (lncRNAs). Although accumulating evidence demonstrates that lncRNAs play important roles in regulating gene expression, the detailed mechanisms of action of most lncRNAs remain unclear. We previously reported that a novel class of lncRNAs with a short half-life (t1/2 < 4 h) in HeLa cells, termed short-lived non-coding transcripts (SLiTs), are closely associated with physiological and pathological functions. In this study, we focused on 26 SLiTs and nuclear-enriched abundant lncRNA, MALAT1(t1/2 of 7.6 h in HeLa cells) in neural stem cells (NSCs) derived from human induced pluripotent stem cells, and identified four SLiTs (TUG1, GAS5, FAM222-AS1, and SNHG15) that were affected by the following typical chemical stresses (oxidative stress, heavy metal stress and protein synthesis stress). We also found the expression levels of LINC00152 (t1/2 of 2.1 h in NSCs), MALAT1 (t1/2 of 1.8 h in NSCs), and their neighboring genes were elevated proportionally to the chemical doses. Moreover, we confirmed that the overexpression of LINC00152 or MALAT1 upregulated the expressions of their neighboring genes even in the absence of chemical stress. These results reveal that LINC00152 and MALAT1 modulate their neighboring genes, and thus provide a deeper understanding of the functions of lncRNAs.

Protein cohesion induced by metal ions observed with fluorescence correlation spectroscopy

Nine metal ions were evaluated in the point of denaturating action of proteins. When some metal ions were added to the diluted protein solutions, aggregates appear: stronger denaturation causes the appearance of the larger-size aggregate. The size of the aggregatates are determined by fluorescence correlation spectroscopy (FCS). Green fluorescent protein (ZsGreen) and PE(phycoerythrin)-conjugated human-antibody monoclonal protein were employed as the target protein, of which solution was diluted 100–500 times and mixed with metal ions. According to this process, the denaturation power of metal ions is in the order of Mn2+≈ Fe2+< Co2+< Ni2+< Tl+< Cd2+< Cu+< Cu2+< Pb2+for ZsGreen, and Tl+≈ Ni2+< Cd2+< Fe2+< Cr3+≪ Pb2+for PE-conjugated antibody protein. Pb2+exhibits the strongest power of denaturation. In the case of ZsGreen, the denaturation power of metal ions is on the order of the Irving-Williams series, which provide the coordination tendency against ligands possessing nitrogen and oxygen. The present method with FCS is effective to evaluate the denaturation power of metal ions against proteins.