Three manganese oxide-rich marine sediments harbor similar communities of acetate-oxidizing manganese-reducing bacteria

Dissimilatory manganese reduction dominates anaerobic carbon oxidation in marine sediments with high manganese oxide concentrations, but the microorganisms responsible for this process are largely unknown. In this study, the acetate-utilizing manganese-reducing microbiota in geographically well-separated, manganese oxide-rich sediments from Gullmar Fjord (Sweden), Skagerrak (Norway) and Ulleung Basin (Korea) were analyzed by 16S rRNA-stable isotope probing (SIP). Manganese reduction was the prevailing terminal electron-accepting process in anoxic incubations of surface sediments, and even the addition of acetate stimulated neither iron nor sulfate reduction. The three geographically distinct sediments harbored surprisingly similar communities of acetate-utilizing manganese-reducing bacteria: 16S rRNA of members of the genera Colwellia and Arcobacter and of novel genera within the Oceanospirillaceae and Alteromonadales were detected in heavy RNA-SIP fractions from these three sediments. Most probable number (MPN) analysis yielded up to 10⁶ acetate-utilizing manganese-reducing cells cm⁻³ in Gullmar Fjord sediment. A 16S rRNA gene clone library that was established from the highest MPN dilutions was dominated by sequences of Colwellia and Arcobacter species and members of the Oceanospirillaceae, supporting the obtained RNA-SIP results. In conclusion, these findings strongly suggest that (i) acetate-dependent manganese reduction in manganese oxide-rich sediments is catalyzed by members of taxa (Arcobacter, Colwellia and Oceanospirillaceae) previously not known to possess this physiological function, (ii) similar acetate-utilizing manganese reducers thrive in geographically distinct regions and (iii) the identified manganese reducers differ greatly from the extensively explored iron reducers in marine sediments.     

Synthesis of full length PB1-F2 influenza A virus proteins from 'Spanish flu' and 'bird flu'.

The proapoptotic influenza A virus PB1-F2 protein contributes to viral pathogenicity and is present in most human and avian isolates. Previous synthetic protocols have been improved to provide a synthetic full length H1N1 type PB1-F2 protein that is encoded by the 'Spanish flu' isolate and an equivalent protein from an avian host that is representative of a highly pathogenic H5N1 'bird flu' isolate, termed SF2 and BF2, respectively. Full length SF2, different mutants of BF2 and a number of fragments of these peptides have been synthesized by either the standard solid-phase peptide synthesis method or by native chemical ligation of unprotected N- and C-terminal peptide fragments. For SF2 chemical ligation made use of the histidine and the cysteine residues located in positions 41 and 42 of the native sequence, respectively, to afford a highly efficient synthesis of SF2 compared to the standard SPPS elongation method. By-product formation at the aspartic acid residue in position 23 was prevented by specific modifications of the SPPS protocol. As the native sequence of BF2 does not contain a cysteine residue two different mutants of BF2 (Y42C) and BF2 (S47C) with appropriate cysteine exchanges were produced. In addition to the full length molecules, fragments of the native sequences were synthesized for comparison of their physical characteristics with those from the H1N1 human isolate A/Puerto Rico/8/34 (H1N1). All peptides were analyzed by mass spectrometry, (1)H NMR spectroscopy, and SDS-PAGE. The protocols allow the synthesis of significant amounts of PB1-F2 and its related peptides. Copyright (c) 2008 European Peptide Society and John Wiley & Sons, Ltd.     

Hearing in Cichlid Fishes under Noise Conditions

Background

Hearing thresholds of fishes are typically acquired under laboratory conditions. This does not reflect the situation in natural habitats, where ambient noise may mask their hearing sensitivities. In the current study we investigate hearing in terms of sound pressure (SPL) and particle acceleration levels (PAL) of two cichlid species within the naturally occurring range of noise levels. This enabled us to determine whether species with and without hearing specializations are differently affected by noise.

Methodology/Principal Findings

We investigated auditory sensitivities in the orange chromide Etroplus maculatus, which possesses anterior swim bladder extensions, and the slender lionhead cichlid Steatocranus tinanti, in which the swim bladder is much smaller and lacks extensions. E. maculatus was tested between 0.2 and 3kHz and S. tinanti between 0.1 and 0.5 kHz using the auditory evoked potential (AEP) recording technique. In both species, SPL and PAL audiograms were determined in the presence of quiet laboratory conditions (baseline) and continuous white noise of 110 and 130 dB RMS. Baseline thresholds showed greatest hearing sensitivity around 0.5 kHz (SPL) and 0.2 kHz (PAL) in E. maculatus and 0.2 kHz in S. tinanti. White noise of 110 dB elevated the thresholds by 0–11 dB (SPL) and 7–11 dB (PAL) in E. maculatus and by 1–2 dB (SPL) and by 1–4 dB (PAL) in S. tinanti. White noise of 130 dB elevated hearing thresholds by 13–29 dB (SPL) and 26–32 dB (PAL) in E. maculatus and 6–16 dB (SPL) and 6–19 dB (PAL) in S. tinanti.

Conclusions

Our data showed for the first time for SPL and PAL thresholds that the specialized species was masked by different noise regimes at almost all frequencies, whereas the non-specialized species was much less affected. This indicates that noise can limit sound detection and acoustic orientation differently within a single fish family.     

Influences of AT1 receptor blockade on tissue metabolism in obese men

ANG II applied to the interstitial space influences carbohydrate and lipid metabolism in a tissue-specific fashion. Thus endogenous ANG II may have a tonic effect on tissue metabolism that could be reversed with ANG II type 1 (AT1) receptor blockade, particularly during adrenergic stimulation. We studied 14 obese men. They were treated for 10 days with the AT1 receptor blocker irbesartan or with placebo in a double-blind and crossover fashion. At the end of each treatment period, we assessed skeletal muscle and adipose tissue metabolism using the microdialysis technique. The ethanol dilution technique was applied to follow changes in tissue blood flow. Measurements were obtained at baseline and during application of incremental isoproterenol concentrations through the microdialysis catheter. Blood pressure decreased from 133 +/- 3/84 +/- 3 to 128 +/- 3/79 +/- 2 mmHg for systolic and diastolic blood, respectively (P = 0.02 and 0.006, respectively) with AT1 receptor blockade. Isoproterenol perfusion caused a dose-dependent increase in dialysate glycerol in adipose tissue and in skeletal muscle. Irbesartan slightly reduced the isoproterenol-induced glycerol response in adipose tissue (P < 0.05 by ANOVA). Ethanol ratio, interstitial glucose supply, and lactate production in adipose tissue and skeletal muscle were similar with placebo and irbesartan. We conclude that AT1 receptor blockade in obese men does not reveal a major tonic ANG II effect on interstitial glucose supply, lipolysis, or glycolysis in skeletal muscle, either at rest or during beta-adrenergic stimulation. Endogeneous ANG II may slightly increase adipose tissue lipolysis. The mechanism may promote the redistribution of triglycerides from adipose tissue toward other organs.     

Elementary mode analysis: a useful metabolic pathway analysis tool for characterizing cellular metabolism

Elementary mode analysis is a useful metabolic pathway analysis tool to identify the structure of a metabolic network that links the cellular phenotype to the corresponding genotype. The analysis can decompose the intricate metabolic network comprised of highly interconnected reactions into uniquely organized pathways. These pathways consisting of a minimal set of enzymes that can support steady state operation of cellular metabolism represent independent cellular physiological states. Such pathway definition provides a rigorous basis to systematically characterize cellular phenotypes, metabolic network regulation, robustness, and fragility that facilitate understanding of cell physiology and implementation of metabolic engineering strategies. This mini-review aims to overview the development and application of elementary mode analysis as a metabolic pathway analysis tool in studying cell physiology and as a basis of metabolic engineering.     

Immunocytochemical localization of Na+-HCO3- cotransporters and carbonic anhydrase dependence of fluid transport in corneal endothelial cells

In corneal endothelium, there is evidence for basolateral entry of HCO(3)(-) into corneal endothelial cells via Na(+)-HCO(3)(-) cotransporter (NBC) proteins and for net HCO(3)(-) flux from the basolateral to the apical side. However, how HCO(3)(-) exits the cells through the apical membrane is unclear. We determined that cultured corneal endothelial cells transport HCO(3)(-) similarly to fresh tissue. In addition, Cl(-) channel inhibitors decreased fluid transport by at most 16%, and inhibition of membrane-bound carbonic anhydrase IV by benzolamide or dextran-bound sulfonamide decreased fluid transport by at most 29%. Therefore, more than half of the fluid transport cannot be accounted for by anion transport through apical Cl(-) channels, CO(2) diffusion across the apical membrane, or a combination of these two mechanisms. However, immunocytochemistry using optical sectioning by confocal microscopy and cryosections revealed the presence of NBC transporters in both the basolateral and apical cell membranes of cultured bovine corneal endothelial cells and freshly isolated rabbit endothelia. This newly detected presence of an apical NBC transporter is consistent with its being the missing mechanism sought. We discuss discrepancies with other reports and provide a model that accounts for the experimental observations by assuming different stoichiometries of the NBC transport proteins at the basolateral and apical sides of the cells. Such functional differences might arise either from the expression of different isoforms or from regulatory factors affecting the stoichiometry of a single isoform.     

Diadenosine tetraphosphate (Ap4A) is compartmentalized in nuclei of mammalian cells

The intracellular compartmentation of Ap4A in various growth and cell-cycle stages in mammalian cells was studied by applying a non-aqueous extraction procedure for cell nuclei. In both slowly and in exponentially growing Ehrlich ascites tumour cells from random cultures, more than 75% of the whole cellular Ap4A content is localized in the nuclei. In G1 and early S-phase cells of synchronized baby hamster kidney (BHK) fibroblast cultures, approx. 90% of the intracellular Ap4A pool is confined to the nuclear compartment. In contrast, Ap4A is distributed to nearly equal amounts between cytoplasm and nuclei during mid-S phase. After transition through the S-phase, increasing proportions of Ap4A (78% 18 h and 96% 22 h after serum replenishing, respectively) are again localized in the nuclear compartment.     

OmpW of Caulobacter crescentus Functions as an Outer Membrane Channel for Cations

Caulobacter crescentus is an oligotrophic bacterium that lives in dilute organic environments such as soil and freshwater. This bacterium represents an interesting model for cellular differentiation and regulation because daughter cells after division have different forms: one is motile while the other is non-motile and can adhere to surfaces. Interestingly, the known genome of C. crescentus does not contain genes predicted to code for outer membrane porins of the OmpF/C general diffusion type present in enteric bacteria or those coding for specific porins selective for classes of substrates. Instead, genes coding for 67 TonB-dependent outer membrane receptors have been identified, suggesting that active transport of specific nutrients may be the norm. Here, we report that high channel-forming activity was observed with crude outer membrane extracts of C. crescentus in lipid bilayer experiments, indicating that the outer membrane of C. crescentus contained an ion-permeable channel with a single-channel conductance of about 120 pS in 1M KCl. The channel-forming protein with an apparent molecular mass of about 20 kDa was purified to homogeneity. Partial protein sequencing of the protein indicated it was a member of the OmpW family of outer membrane proteins from Gram-negative bacteria. This channel was not observed in reconstitution experiments with crude outer membrane extracts of an OmpW deficient C. crescentus mutant. Biophysical analysis of the C. crescentus OmpW suggested that it has features that are special for general diffusion porins of Gram-negative outer membranes because it was not a wide aqueous channel. Furthermore, OmpW of C. crescentus seems to be different to known OmpW porins and has a preference for ions, in particular cations. A putative model for OmpW of C. crescentus was built on the basis of the known 3D-structures of OmpW of Escherichia coli and OprG of Pseudomonas aeruginosa using homology modeling. A comparison of the two known structures with the model of OmpW of C. crescentus suggested that it has a more hydrophilic interior and possibly a larger diameter.     

In vitro analysis of the oligodendrocyte lineage in mice during demyelination and remyelination

A demyelinating disease induced in C57B1/6N mice by intracranial injection of a coronavirus (murine hepatitis virus strain A59) is followed by functional recovery and efficient CNS myelin repair. To study the biological properties of the cells involved in this repair process, glial cells were isolated and cultured from spinal cords of these young adult mice during demyelination and remyelination. Using three-color immunofluorescence combined with [3H]thymidine autoradiography, we have analyzed the antigenic phenotype and mitotic potential of individual glial cells. We identified oligodendrocytes with an antibody to galactocerebroside, astrocytes with an antibody to glial fibrillary acidic protein, and oligodendrocyte-type 2 astrocyte (O-2A) progenitor cells with the O4 antibody. Cultures from demyelinated tissue differed in several ways from those of age-matched controls: first, the total number of O-2A lineage cells was strikingly increased; second, the O-2A population consisted of a higher proportion of O4-positive astrocytes and cells of mixed oligodendrocyte-astrocyte phenotype; and third, all the cell types within the O-2A lineage showed enhanced proliferation. This proliferation was not further enhanced by adding PDGF, basic fibroblast growth factor (bFGF), or insulin-like growth factor I (IGF-I) to the defined medium. However, bFGF and IGF-I seemed to influence the fate of O-2A lineage cells in cultures of demyelinated tissue. Basic FGF decreased the percentage of cells expressing galactocerebroside. In contrast, IGF-I increased the relative proportion of oligodendrocytes. Thus, O-2A lineage cells from adult mice display greater phenotypic plasticity and enhanced mitotic potential in response to an episode of demyelination. These properties may be linked to the efficient remyelination achieved in this demyelinating disease.