Diversity of Ferrous Iron-Oxidizing,Nitrate-Reducing Bacteria and their Involvement in Oxygen-Independent Iron Cycling

In previous studies, three different strains (BrG1, BrG2, and BrG3) of ferrous iron-oxidizing, nitrate-reducing bacteria were obtained from freshwater sediments. All three strains were facultative anaerobes and utilized a variety of organic substrates and molecular hydrogen with nitrate as electron acceptor. In this study, analyses of 16S rDNA sequences showed that strain BrG1 was affiliated with the genus Acidovorax, strain BrG2 with the genus Aquabacterium, and strain BrG3 with the genus Thermomonas. Previously, bacteria similar to these three strains were detected with molecular techniques in MPN dilution series for ferrous iron-oxidizing, nitrate-reducing bacteria inoculated with different freshwater sediment samples. In the present study, further molecular analyses of these MPN cultures indicated that the ability to oxidize ferrous iron with nitrate is widespread amongst the Proteobacteria and may also be found among the Gram-positive bacteria with high GC content of DNA. Nitrate-reducing bacteria oxidized ferrous iron to poorly crystallized ferrihydrite that was suitable as an electron acceptor for ferric iron-reducing bacteria. Biologically produced ferrihydrite and synthetically produced ferrihydrite were both well suited as electron acceptors in MPN dilution cultures. Repeated anaerobic cycling of iron was shown in a coculture of ferrous iron-oxidizing bacteria and the ferric iron-reducing bacterium Geobacter bremensis. The results indicate that iron can be cycled between its oxidation states +II and +III by microbial activities in anoxic sediments.     

How Natalizumab Binds and Antagonizes α4 Integrins

Natalizumab antibody to α₄-integrins is used in therapy of multiple sclerosis and Crohn''s disease. A crystal structure of the Fab bound to an α₄ integrin β-propeller and thigh domain fragment shows that natalizumab recognizes human-mouse differences on the circumference of the β-propeller domain. The epitope is adjacent to but outside of a ligand-binding groove formed at the interface with the β-subunit βI domain and shows no difference in structure when bound to Fab. Competition between Fab and the ligand vascular cell adhesion molecule (VCAM) for binding to cell surface α₄β₁ shows noncompetitive antagonism. In agreement, VCAM docking models suggest that binding of domain 1 of VCAM to α₄-integrins is unimpeded by the Fab, and that bound Fab requires a change in orientation between domains 1 and 2 of VCAM for binding to α₄β₁. Mapping of species-specific differences onto α₄β₁ and α₄β₇ shows that their ligand-binding sites are highly conserved. Skewing away from these conserved regions of the epitopes recognized by current therapeutic function-blocking antibodies has resulted in previously unanticipated mechanisms of action.     

Benzoic acid and specific 2-oxo acids activate hepatic efflux of glutamate at OAT2

The liver is the principal source of glutamate in blood plasma. Recently we have discovered that efflux of glutamate from hepatocytes is catalyzed by the transporter OAT2 (human gene symbol SLC22A7). Organic anion transporter 2 (OAT2) is an integral membrane protein of the sinusoidal membrane domain; it is primarily expressed in liver and much less in kidney, both in rats and humans. Many years ago, Häussinger and coworkers have demonstrated in isolated perfused rat liver that benzoic acid or specific 2-oxo acid analogs of amino acids like e.g. 2-oxo-4-methyl-pentanoate (‘2-oxo-leucine’) strongly stimulate release of glutamate (up to 7-fold); ‘2-oxo-valine’ and the corresponding amino acids were without effect. The molecular mechanism of efflux stimulation has remained unclear. In the present study, OAT2 from human and rat were heterologously expressed in 293 cells. Addition of 1 mmol/l benzoic acid to the external medium increased OAT2-specific efflux of glutamate up to 20-fold; ‘2-oxo-leucine’ was also effective, but not ‘2-oxo-valine’. Similar effects were seen for efflux of radiolabeled orotic acid. Expression of OAT2 did not increase uptake of benzoic acid; thus, benzoic acid is no substrate, and trans-stimulation can be excluded. Instead, further experiments suggest that increased efflux of glutamate is caused by direct interaction of benzoic acid and specific 2-oxo acids with OAT2. We propose that stimulators bind to a distinct extracellular site and thereby accelerate relocation of the empty substrate binding site to the intracellular face. Increased glutamate efflux at OAT2 could be the main benefit of benzoate treatment in patients with urea cycle defects.     

Cytochrome cd1 Nitrite Reductase NirS Is Involved in Anaerobic Magnetite Biomineralization in Magnetospirillum gryphiswaldense and Requires NirN for Proper d1 Heme Assembly

The alphaproteobacterium Magnetospirillum gryphiswaldense synthesizes magnetosomes, which are membrane-enveloped crystals of magnetite. Here we show that nitrite reduction is involved in redox control during anaerobic biomineralization of the mixed-valence iron oxide magnetite. The cytochrome cd₁-type nitrite reductase NirS shares conspicuous sequence similarity with NirN, which is also encoded within a larger nir cluster. Deletion of any one of these two nir genes resulted in impaired growth and smaller, fewer, and aberrantly shaped magnetite crystals during nitrate reduction. However, whereas nitrite reduction was completely abolished in the ΔnirS mutant, attenuated but significant nitrite reduction occurred in the ΔnirN mutant, indicating that only NirS is a nitrite reductase in M. gryphiswaldense. However, the ΔnirN mutant produced a different form of periplasmic d₁ heme that was not noncovalently bound to NirS, indicating that NirN is required for full reductase activity by maintaining a proper form of d₁ heme for holo-cytochrome cd₁ assembly. In conclusion, we assign for the first time a physiological function to NirN and demonstrate that effective nitrite reduction is required for biomineralization of wild-type crystals, probably by contributing to oxidation of ferrous iron under oxygen-limited conditions.     

‘Real time’ genetic manipulation: a new tool for ecological field studies

Field experiments with transgenic plants often reveal the functional significance of genetic traits that are important for the performance of the plants in their natural environments. Until now, only constitutive overexpression, ectopic expression and gene silencing methods have been used to analyze gene‐related phenotypes in natural habitats. These methods do not allow sufficient control over gene expression for the study of ecological interactions in real time, of genetic traits that play essential roles in development, or of dose‐dependent effects. We applied the sensitive dexamethasone (DEX)‐inducible pOp6/LhGR expression system to the ecological model plant Nicotiana attenuata and established a lanolin‐based DEX application method to facilitate ectopic gene expression and RNA interference‐mediated gene silencing in the field and under challenging conditions (e.g. high temperature, wind and UV radiation). Fully established field‐grown plants were used to silence phytoene desaturase and thereby cause photobleaching only in specific plant sectors, and to activate expression of the cytokinin (CK) biosynthesis gene isopentenyl transferase (ipt). We used ipt expression to analyze the role of CKs in both the glasshouse and the field to understand resistance to the native herbivore Tupiocoris notatus, which attacks plants at small spatial scales. By spatially restricting ipt expression and elevating CK levels in single leaves, damage by T. notatus increased, demonstrating the role of CKs in this plant–herbivore interaction at a small scale. As the arena of most ecological interactions is highly constrained in time and space, these tools will advance the genetic analysis of dynamic traits that matter for plant performance in nature.     

Sex allocation and investment into pre- and post-copulatory traits in simultaneous hermaphrodites: the role of polyandry and local sperm competition

Sex allocation theory predicts the optimal allocation to male and female reproduction in sexual organisms. In animals, most work on sex allocation has focused on species with separate sexes and our understanding of simultaneous hermaphrodites is patchier. Recent theory predicts that sex allocation in simultaneous hermaphrodites should strongly be affected by post-copulatory sexual selection, while the role of pre-copulatory sexual selection is much less clear. Here, we review sex allocation and sexual selection theory for simultaneous hermaphrodites, and identify several strong and potentially unwarranted assumptions. We then present a model that treats allocation to sexually selected traits as components of sex allocation and explore patterns of allocation when some of these assumptions are relaxed. For example, when investment into a male sexually selected trait leads to skews in sperm competition, causing local sperm competition, this is expected to lead to a reduced allocation to sperm production. We conclude that understanding the evolution of sex allocation in simultaneous hermaphrodites requires detailed knowledge of the different sexual selection processes and their relative importance. However, little is currently known quantitatively about sexual selection in simultaneous hermaphrodites, about what the underlying traits are, and about what drives and constrains their evolution. Future work should therefore aim at quantifying sexual selection and identifying the underlying traits along the pre- to post-copulatory axis.     

A sphingosine 1-phosphate receptor 2 selective allosteric agonist

Molecular probe tool compounds for the Sphingosine 1-phosphate receptor 2 (S1PR2) are important for investigating the multiple biological processes in which the S1PR2 receptor has been implicated. Amongst these are NF-κB-mediated tumor cell survival and fibroblast chemotaxis to fibronectin. Here we report our efforts to identify selective chemical probes for S1PR2 and their characterization. We employed high throughput screening to identify two compounds which activate the S1PR2 receptor. SAR optimization led to compounds with high nanomolar potency. These compounds, XAX-162 and CYM-5520, are highly selective and do not activate other S1P receptors. Binding of CYM-5520 is not competitive with the antagonist JTE-013. Mutation of receptor residues responsible for binding to the zwitterionic headgroup of sphingosine 1-phosphate (S1P) abolishes S1P activation of the receptor, but not activation by CYM-5520. Competitive binding experiments with radiolabeled S1P demonstrate that CYM-5520 is an allosteric agonist and does not displace the native ligand. Computational modeling suggests that CYM-5520 binds lower in the orthosteric binding pocket, and that co-binding with S1P is energetically well tolerated. In summary, we have identified an allosteric S1PR2 selective agonist compound.     

An evidence-based consensus for the classification of arbuscular mycorrhizal fungi (Glomeromycota)

The publication of a large number of taxon names at all levels within the arbuscular mycorrhizal fungi (Glomeromycota) has resulted in conflicting systematic schemes and generated considerable confusion among biologists working with these important plant symbionts. A group of biologists with more than a century of collective experience in the systematics of Glomeromycota examined all available molecular–phylogenetic evidence within the framework of phylogenetic hypotheses, incorporating morphological characters when they were congruent. This study is the outcome, wherein the classification of Glomeromycota is revised by rejecting some new names on the grounds that they are founded in error and by synonymizing others that, while validly published, are not evidence-based. The proposed “consensus” will provide a framework for additional original research aimed at clarifying the evolutionary history of this important group of symbiotic fungi.