Regulation of ammonia homeostasis by the ammonium transporter AmtA in Dictyostelium discoideum

Ammonia has been shown to function as a morphogen at multiple steps during the development of the cellular slime mold Dictyostelium discoideum; however, it is largely unknown how intracellular ammonia levels are controlled. In the Dictyostelium genome, there are five genes that encode putative ammonium transporters: amtA, amtB, amtC, rhgA, and rhgB. Here, we show that AmtA regulates ammonia homeostasis during growth and development. We found that cells lacking amtA had increased levels of ammonia/ammonium, whereas their extracellular ammonia/ammonium levels were highly decreased. These results suggest that AmtA mediates the excretion of ammonium. In support of a role for AmtA in ammonia homeostasis, AmtA mRNA is expressed throughout the life cycle, and its expression level increases during development. Importantly, AmtA-mediated ammonia homeostasis is critical for many developmental processes. amtA(-) cells are more sensitive to NH(4)Cl than wild-type cells in inhibition of chemotaxis toward cyclic AMP and of formation of multicellular aggregates. Furthermore, even in the absence of exogenously added ammonia, we found that amtA(-) cells produced many small fruiting bodies and that the viability and germination of amtA(-) spores were dramatically compromised. Taken together, our data clearly demonstrate that AmtA regulates ammonia homeostasis and plays important roles in multiple developmental processes in Dictyostelium.     

Environmental DNA monitoring method of the commercially important and endangered fish Gnathopogon caerulescens

Limnology - Gnathopogon caerulescens is an endangered but commercially important fish in Lake Biwa, Japan. The population size of G. caerulescens has drastically reduced in the past decades, and...     

A method for estimating nucleotide diversity from AFLP data

A method for estimating the nucleotide diversity from AFLP data is developed by using the relationship between the number of nucleotide changes and the proportion of shared bands. The estimation equation is based on the assumption that GC-content is 0.5. Computer simulations, however, show that this method gives a reasonably accurate estimate even when GC-content deviates from 0.5, as long as the number of nucleotide changes per site (nucleotide diversity) is small. As an example, the nucleotide diversity of the wild yam, Dioscorea tokoro, was estimated. The estimated nucleotide diversity is 0.0055, which is larger than estimations from nucleotide sequence data for Adh and Pgi.     

Evaluation of parameters of oxidative stress after in vitro exposure to FMCW- and CDMA-modulated radiofrequency radiation fields

The goal of this study was to determine whether radiofrequency (RF) radiation is capable of inducing oxidative stress or affecting the response to oxidative stress in cultured mammalian cells. The two types of RF radiation investigated were frequency-modulated continuous-wave with a carrier frequency of 835.62 MHz (FMCW) and code division multiple access centered on 847.74 MHz (CDMA). To evaluate the effect of RF radiation on oxidative stress, J774.16 mouse macrophage cells were stimulated with gamma-interferon (IFN) and bacterial lipopolysaccharide (LPS) prior to exposure. Cell cultures were exposed for 20-22 h to a specific absorption rate of 0.8 W/kg at a temperature of 37.0 +/- 0.3 degrees C. Oxidative stress was evaluated by measuring oxidant levels, antioxidant levels, oxidative damage and nitric oxide production. Oxidation of thiols was measured by monitoring the accumulation of glutathione disulfide (GSSG). Cellular antioxidant defenses were evaluated by measuring superoxide dismutase activity (CuZnSOD and MnSOD) as well as catalase and glutathione peroxidase activity. The trypan blue dye exclusion assay was used to measure any changes in viability. The results of these studies indicated that FMCW- and CDMA-modulated RF radiation did not alter parameters indicative of oxidative stress in J774.16 cells. FMCW- and CDMA-modulated fields did not alter the level of intracellular oxidants, accumulation of GSSG or induction of antioxidant defenses in IFN/LPS-stimulated cells. Consistent with the lack of an effect on oxidative stress parameters, no change in toxicity was observed in J774.16 cells after either optimal (with or without inhibitors of nitric oxide synthase) or suboptimal stimulation.     

New analogue of arenastatin A, a potent cytotoxic spongean depsipeptide, with anti-tumor activity

Two analogues possessing steric hindered substituents on C-15 of arenastatin A (1), a potent cytotoxic spongean depsipeptide, were synthesized and shown to enhance stability in mouse serum. Notably, 15-tert-butylanalogue (6) with higher cytotoxicity exhibited in vivo anti-tumor activity through iv administration different from 1. Additionally, conformation analysis among the two analogues and arenastatin A (1) indicated that the torsion angle from C-14 to C-20 is a conclusive factor for the potent cytotoxicity of 1.     

Species diversity in native fish community in Japan: comparison between non-invaded and invaded ponds by exotic fish

The relationship between invasions by two exotic fishes (Micropterus salmoides and Lepomis macrochirus) and species diversity in native fish communities was studied in 14 Japanese farm ponds. We found that mean number of species in native fish communities was three times higher in the ponds without the exotic fish than in the ponds with them. Further, negative relationships were observed between abundance of the two exotic fish and the total abundance of native fish communities. Our results suggest that invasions by the two exotic fish caused serious depletion of native fish communities, although another process can also be considered , that is, that ponds with poor native fish communities were prone to colonization by these exotic fish.     

PCR-Based Approach for Sequencing Mitochondrial Genomes of Decapod Crustaceans,with a Practical Example from Kuruma Prawn (<Emphasis Type="Italic">Marsupenaeus japonicus</Emphasis>)

An approach for sequencing the entire mitochondrial genomes (mitogenomes) of decapod crustaceans using 79 newly designed and 7 published polymerase chain reaction (PCR) primers is described. The approach comprises the following steps: (1) the entire mitogenome is amplified in 2 or 3 long PCRs; (2) the 86 primers are used in different combinations to amplify contiguous, overlapping short segments of the entire mitogenome with the diluted long PCR products as templates; (3) direct cycle sequencing is conducted using the short PCR products. This strategy allows a more rapid determination of decapod mitogenomic sequences than a traditional method using cloned mitochondrial DNA and primer walking strategy. As a practical example, the mitogenomic sequence for a kuruma prawn Marsupenaeus japonicus (Crustacea: Decapoda), was determined using the PCR-based approach.     

Synthesis and antibacterial activity of novel and potent DNA gyrase inhibitors with azole ring

The 4-piperidyl moiety and the pyrazole ring in 1-(3-chlorophenyl)-5-(4-phenoxyphenyl)-3-(4-piperidyl)pyrazole 2, which has previously shown improved DNA gyrase inhibition and target-related antibacterial activity, were transformed to other groups and the in vitro antibacterial activity of the synthesized compounds was evaluated. The selected pyrazole, oxazole and imidazole derivatives showed moderate inhibition against DNA gyrase and topoisomerase IV with similar IC(50) values (IC(50)=9.4-25 microg/mL). In addition, many of the pyrazole, oxazole and imidazole derivatives synthesized in this study exhibited potent antibacterial activity against quinolone-resistant clinical isolates and coumarin-resistant laboratory isolates of Gram-positive bacteria with minimal inhibitory concentration values equivalent to those against susceptible strains.     

A selective inducible NOS dimerization inhibitor prevents systemic,cardiac, and pulmonary hemodynamic dysfunction in endotoxemic mice

Increased nitric oxide (NO) production by inducible NO synthase (NOS2), an obligate homodimer, is implicated in the cardiovascular sequelae of sepsis. We tested the ability of a highly selective NOS2 dimerization inhibitor (BBS-2) to prevent endotoxin-induced systemic hypotension, myocardial dysfunction, and impaired hypoxic pulmonary vasoconstriction (HPV) in mice. Mice were challenged with Escherichia coli endotoxin before treatment with BBS-2 or vehicle. Systemic blood pressure was measured before and 4 and 7 h after endotoxin challenge, and echocardiographic parameters of myocardial function were measured before and 7 h after endotoxin challenge. The pulmonary vasoconstrictor response to left mainstem bronchus occlusion, which is a measure of HPV, was studied 22 h after endotoxin challenge. BBS-2 treatment alone did not alter baseline hemodynamics. BBS-2 treatment blocked NOS2 dimerization and completely inhibited the endotoxin-induced increase of plasma nitrate and nitrite levels. Treatment with BBS-2 after endotoxin administration prevented systemic hypotension and attenuated myocardial dysfunction. BBS-2 also prevented endotoxin-induced impairment of HPV. In contrast, treatment with NG-nitro-l-arginine methyl ester, which is an inhibitor of all three NOS isoforms, prevented the systemic hypotension but further aggravated the myocardial dysfunction associated with endotoxin challenge. Treatment with BBS-2 prevented endotoxin from causing key features of cardiovascular dysfunction in endotoxemic mice. Selective inhibition of NOS2 dimerization with BBS-2, while sparing the activities of other NOS isoforms, may prove to be a useful treatment strategy in sepsis.