5-HT(3) receptors mediate inflammation-induced unmasking of functional tachykinin responses in vitro.

Exogenously applied tachykinins produce no measurable electrophysiological responses in the somata of vagal afferent neurons [nodose ganglion neurons (NGNs)] isolated from naive guinea pigs. By contrast, after in vitro antigen challenge of nodose ganglia from guinea pigs immunized with chick ovalbumin, approximately 60% (53 of 89) of NGNs were depolarized an average of 13 +/- 1.2 mV by substance P (SP; 100 nM; n = 53). Receptor antagonists and enzyme inhibitors were utilized to screen a number of mast cell-derived mediators for their role in the uncovering or "unmasking" of functional tachykinin receptors after antigen challenge. Two chemically distinct 5-hydroxytryptamine-3-receptor antagonists significantly reduced the percentage of NGNs displaying depolarizing SP responses. Treatment with Y-25130 (1 or 10 microM) or tropisetron (1 microM) 15 min before and during antigen challenge reduced the percentage of SP-responsive neurons to approximately 20 and approximately 15% respectively. These results suggest that activation of 5-hydroxytryptamine-3 receptors plays an integral role in the unmasking of functional tachykinin receptors after specific antigen challenge of nodose ganglia. The mediator(s) underlying tachykinin-receptor unmasking in the remainder of the NGNs has yet to be characterized. However, it does not appear to be histamine, prostanoids, or peptidoleukotrienes.     

Mutations of the immunoglobulin heavy chain variable region gene in CD99-deficient BJAB cell line

Hodgkin's disease (HD) is a lymphoid neoplasm characterized by a low frequency of malignant giant tumor cells, known as Hodgkin's and Reed-Sternberg (HRS) cells. Sequence analysis of the immunoglobulin heavy chain hypervariable region (IgH V) genes of HRS cells revealed multiple nucleotide substitutions, indicating somatic mutations, and suggested that HRS cells originate from germinal center B cells or their progeny. We previously reported that CD99-antisense transfected B cell lines led to the generation of cells with a HRS phenotype. Because it is considered that HRS cells in HD carry somatic mutations of the IgH genes, we assume that somatic mutation may take place in the IgH genes of HRS-like cells which do not express CD99. Here we report that CD99 downregulated BJAB cell line has several mutations in IgH V genes. The frequency of mutation was 5.2 x 10(-4) mut.bp(-1) out of total sequenced cell clones. On the contrary, control vector transfected BJAB cell line or CD99 downregulated IM9 cell line did not show any mutations on single strand conformational polymorphism (SSCP) and sequence analysis. We expect that the analysis of the mutation pattern of the CD99-deficient BJAB cell line might be the basis for the understanding of the molecular and cellular mechanism that regulate somatic mutation and B cell selection.     

First success of catalytic epoxidation of olefins by an electron-rich iron(III) porphyrin complex and H2O2: imidazole effect on the activation of H2O2 by iron porphyrin complexes in aprotic solvent

An electron-rich iron(III) porphyrin complex (meso-tetramesitylporphinato)iron(III) chloride [Fe(TMP)Cl], was found to catalyze the epoxidation of olefins by aqueous 30% H₂O₂ when the reaction was carried out in the presence of 5-chloro-1-methylimidazole (5-Cl-1-MeIm) in aprotic solvent. Epoxides were the predominant products with trace amounts of allylic oxidation products, indicating that Fenton-type oxidation reactions were not involved in the olefin epoxidation reactions. cis-Stilbene was stereospecifically oxidized to cis-stilbene oxide without giving isomerized trans-stilbene oxide product, demonstrating that neither hydroperoxy radical (HOO·) nor oxoiron(IV) porphyrin [(TMP)Fe^IV=O] was responsible for the olefin epoxidations. We also found that the reactivities of other iron(III) porphyrin complexes such as (meso-tetrakis(2,6-dichlorophenyl)porphinato)iron(III) chloride [Fe(TDCPP)Cl], (meso-tetrakis(2,6-difluorophenyl)porphinato)iron(III) chloride [Fe(TDFPP)Cl], and (meso-tetrakis(pentafluorophenyl)porphinato)iron(III) chloride [Fe(TPFPP)Cl] were significantly affected by the presence of the imidazole in the epoxidation of olefins by H₂O₂. These iron porphyrin complexes did not yield cyclohexene oxide in the epoxidation of cyclohexene by H₂O₂ in the absence of 5-Cl-1-MeIm in aprotic solvent; however, addition of 5-Cl-1-MeIm to the reaction solutions gave high yields of cyclohexene oxide with the formation of trace amounts of allylic oxidation products. We proposed, on the basis of the results of mechanistic studies, that the role of the imidazole is to decelerate the O–O bond cleavage of an iron(III) hydroperoxide porphyrin (or H₂O₂–iron(III) porphyrin adduct) and that the intermediate transfers its oxygen to olefins prior to the O–O bond cleavage.     

Characterizing Escherichia coli DH5alpha growth and metabolism in a complex medium using genome-scale flux analysis

Genome-scale flux analysis of Escherichia coli DH5alpha growth in a complex medium was performed to investigate the relationship between the uptake of various nutrients and their metabolic outcomes. During the exponential growth phase, we observed a sequential consumption order of serine, aspartate and glutamate in the complex medium as well as the complete consumption of key carbohydrate nutrients, glucose and trehalose. Based on the consumption and production rates of the measured metabolites, constraints-based flux analysis of a genome-scale E. coli model was then conducted to elucidate their utilization in the metabolism. The in silico analysis revealed that the cell exploited biosynthetic precursors taken up directly from the complex medium, through growth-related anabolic pathways. This suggests that the cell could be functioning in an energetically more efficient manner by reducing the energy needed to produce amino acids. The in silico simulation also allowed us to explain the observed rapid consumption of serine: excessively consumed external serine from the complex medium was mainly converted into pyruvate and glycine, which in turn, led to the acetate accumulation. The present work demonstrates the application of an in silico modeling approach to characterizing microbial metabolism under complex medium condition. This work further illustrates the use of in silico genome-scale analysis for developing better strategies related to improving microbial growth and enhancing the productivity of desirable metabolites.     

Complete mitochondrial genome of the rabbitfish Siganus fuscescens (Perciformes, Siganidae).

We determined the complete nucleotide sequence of the mitochondrial genome for the rabbitfish Siganus fuscescens (Perciformes, Siganidae). This mitochondrial genome, consisting of 16,491 base pairs (bp), included 13 protein-coding genes, 2 ribosomal RNAs, 22 transfer RNAs, and a noncoding control region similar those found in other vertebrates; the gene order was identical to that of typical vertebrates. Most of the genes of S. fuscescens were encoded on the H-strand, while the ND6 and eight tRNA (Gln, Ala, Asn, Cys, Tyr, Ser [UCN], Glu, and Pro) genes were encoded on the L-strand. The reading frames of ATPase 8 and 6 and those of ND4L and ND4 overlapped by ten and seven nucleotides, respectively. All mitochondrial protein-coding genes began with an ATG start codon, except for CO1, which started with GTG. Open reading frames of S. fuscescens ended with TAA (ND1, CO1, ATPase 8, ND4L, ND5 and ND6), and the remainder had incomplete stop codons, either TA (ATPase 6 and CO3) or T (ND2, CO2, ND3, ND4, and Cytb). The origin of L-strand replication in S. fuscescens was located in a cluster of five tRNA genes (WANCY) and was 34 nucleotides in length. A major noncoding region between the tRNA-Pro and tRNA-Phe genes (828 bp) was considered to be the control region (D-loop). Within this sequence, we identified a conserved sequence block characteristic of this region. The rabbitfish was grouped with Siganus canaliculatus in most parsimony analyses, which showed 100% bootstrap support for their divergence. These findings are useful for inferring phylogenetic relationships and identification within the suborder Acanthuroidei.     

Dominant role of the cbb3 oxidase in regulation of photosynthesis gene expression through the PrrBA system in Rhodobacter sphaeroides 2.4.1

In this study, the H303A mutant form of the cbb(3) oxidase (H303A oxidase), which has the H303A mutation in its catalytic subunit (CcoN), was purified from Rhodobacter sphaeroides. The H303A oxidase showed the same catalytic activity as did the wild-type form of the oxidase (WT oxidase). The heme contents of the mutant and WT forms of the cbb(3) oxidase were also comparable. However, the puf and puc operons, which are under the control of the PrrBA two-component system, were shown to be derepressed aerobically in the R. sphaeroides strain expressing the H303A oxidase. Since the strain harboring the H303A oxidase exhibited the same cytochrome c oxidase activity as the stain harboring the WT oxidase did, the aerobic derepression of photosynthesis gene expression observed in the H303A mutant appears to be the result of a defective signaling function of the H303A oxidase rather than reflecting any redox changes in the ubiquinone/ubiquinol pool. It was also demonstrated that ubiquinone inhibits not only the autokinase activity of full-length PrrB but also that of the truncated form of PrrB lacking its transmembrane domain, including the proposed quinone binding sequence. These results imply that the suggested ubiquinone binding site within the PrrB transmembrane domain is not necessary for the inhibition of PrrB kinase activity by ubiquinone. Instead, it is probable that signaling through H303 of the CcoN subunit of the cbb(3) oxidase is part of the pathway through which the cbb(3) oxidase affects the relative kinase/phosphatase activity of the membrane-bound PrrB.     

Effect of various ions, pH, and osmotic pressure on oxidation of elemental sulfur by Thiobacillus thiooxidans.

The oxidation of elemental sulfur by Thiobacillus thiooxidans was studied at pH 2.3, 4.5, and 7.0 in the presence of different concentrations of various anions (sulfate, phosphate, chloride, nitrate, and fluoride) and cations (potassium, sodium, lithium, rubidium, and cesium). The results agree with the expected response of this acidophilic bacterium to charge neutralization of colloids by ions, pH-dependent membrane permeability of ions, and osmotic pressure.