Spatial and niche-based ecological processes drive the distribution of endophytic Sebacinales in soil and root of grassland communities

The interest in endophytic sebacinalean communities has been increasing during the last decade due to the increased knowledge about their symbiotic life style and potential role for ecosystem functioning. Although they are present in many ecosystems, their abundance in individual plant roots is very limited. This fact affects their study: they are difficult to isolate and to detect in root DNA samples. To advance knowledge of the forces that shape their distribution, we approached the parallel study of sebacinalean communities in roots and soil of grassland. Using a small-scale spatially explicit sampling design, we analysed the contribution of spatial position, soil properties, plant community and phylogenetic components to the variation of sebacinalean communities. The results revealed the presence of 11 operational taxonomic units (OTUs) and a high coincidence between root and soil communities: on an average a single-OTU per sample was recorded for both sample types. Spatial distance was found to mainly drive the distribution of Sebacinales in soil, whereas phylogenetic plus environmental signatures mainly drove their presence in roots. Independently of the sample type, we found clear evidence of environmental filtering caused by soil pH which, furthermore, seemed to control the presence of a specialized sebacinalean OTU

Arbuscular mycorrhizal fungal communities are phylogenetically clustered at small scales

Next-generation sequencing technologies with markers covering the full Glomeromycota phylum were used to uncover phylogenetic community structure of arbuscular mycorrhizal fungi (AMF) associated with Festuca brevipila. The study system was a semi-arid grassland with high plant diversity and a steep environmental gradient in pH, C, N, P and soil water content. The AMF community in roots and rhizosphere soil were analyzed separately and consisted of 74 distinct operational taxonomic units (OTUs) in total. Community-level variance partitioning showed that the role of environmental factors in determining AM species composition was marginal when controlling for spatial autocorrelation at multiple scales. Instead, phylogenetic distance and spatial distance were major correlates of AMF communities: OTUs that were more closely related (and which therefore may have similar traits) were more likely to co-occur. This pattern was insensitive to phylogenetic sampling breadth. Given the minor effects of the environment, we propose that at small scales closely related AMF positively associate through biotic factors such as plant-AMF filtering and interactions within the soil biota

Structure of the molybdenum site in YedY, a sulfite oxidase homologue from escherichia coli

YedY from Escherichia coli is a new member of the sulfite oxidase family of molybdenum cofactor (Moco)-containing oxidoreductases. We investigated the atomic structure of the molybdenum site in YedY by X-ray absorption spectroscopy, in comparison to human sulfite oxidase (hSO) and to a MoIV model complex. The K-edge energy was indicative of MoV in YedY, in agreement with X-and Q-band electron paramagnetic resonance results, whereas the hSO protein contained MoVI. In YedY and hSO, molybdenum is coordinated by two sulfur ligands from the molybdopterin ligand of the Moco, one thiolate sulfur of a cysteine (average Mo-S bond length of∼2.4 A), and one (axial) oxo ligand (Mo=O,∼1.7 A). hSO contained a second oxo group at Mo as expected, but in YedY, two species in about a 1:1 ratio were found at the active site, corresponding to an equatorial Mo-OH bond (∼2.1 A) or possibly to a shorter Mo-O-bond. Yet another oxygen (or nitrogen) at a∼2.6 A distance to Mo in YedY was identified, which could originate from a water molecule in the substrate binding cavity or from an amino acid residue close to the molybdenum site, i.e., Glu104, that is replaced by a glycine in hSO, or Asn45. The addition of the poor substrate dimethyl sulfoxide to YedY left the molybdenum coordination unchanged at high pH. In contrast, we found indications that the better substrate trimethylamine N-oxide and the substrate analogue acetone were bound at a∼2.6 Ã distance to the molybdenum, presumably replacing the equatorial oxygen ligand. These findings were used to interpret the recent crystal structure of YedY and bear implications for its catalytic mechanism

A crystallographic and Mo K-edge XAS study of molybdenum oxo bis-, mono-, and non-dithiolene complexes : first-sphere coordination geometry and noninnocence of ligands

Ten square-based pyramidal molybdenum complexes with different sulfur donor ligands, that is, a variety of dithiolenes and sulfides, were prepared, which mimic coordination motifs of the molybdenum cofactors of molybdenum-dependent oxidoreductases. The model compounds were investigated by Mo K-edge X-ray absorption spectroscopy (XAS) and (with one exception) their molecular structures were analyzed by X-ray diffraction to derive detailed information on bond lengths and geometries of the first coordination shell of molybdenum. Only small variations in Mo=O and Mo-S bond lengths and their respective coordination angles were observed for all complexes including those containing Mo(CO) 2 or Mo(μ-S)2Mo motifs. XAS analysis (edge energy) revealed higher relative oxidation levels in the molybdenum ion in compounds with innocent sulfur-based ligands relative to those in dithiolene complexes, which are known to exhibit noninnocence, that is, donation of substantial electron density from ligand to metal. In addition, longer average Mo-S and Mo=O bonds and consequently lower ν(Mo=O) stretching frequencies in the IR spectra were observed for complexes with dithiolene-derived ligands. The results emphasize that the noninnocent character of the dithiolene ligand influences the electronic structure of the model compounds, but does not significantly affect their metal coordination geometry, which is largely determined by the Mo(IV) or (V) ion itself. The latter conclusion also holds for the molybdenum site geometries in the oxidized MoVI cofactor of DMSO reductase and the reduced MoIV cofactor of arsenite oxidase. The innocent behavior of the dithiolene molybdopterin ligands observed in the enzymes is likely to be related to cofactor-protein interactions

Identification of a bis-molybdopterin intermediate in molybdenum cofactor biosynthesis in Escherichia coli

The molybdenum cofactor is an important cofactor, and its biosynthesis is essential for many organisms, including humans. Its basic form comprises a single molybdopterin (MPT) unit, which binds a molybdenum ion bearing three oxygen ligands via a dithiolene function, thus forming Mo-MPT. In bacteria, this form is modified to form the bis-MPT guanine dinucleotide cofactor with two MPT units coordinated at one molybdenum atom, which additionally contains GMPs bound to the terminal phosphate group of the MPTs (bis-MGD). The MobA protein catalyzes the nucleotide addition to MPT, but the mechanism of the biosynthesis of the bis-MGD cofactor has remained enigmatic. We have established an in vitro system for studying bis-MGD assembly using purified compounds. Quantification of the MPT/molybdenum and molybdenum/ phosphorus ratios, time-dependent assays for MPT and MGD detection, and determination of the numbers and lengths of Mo-S and Mo-O bonds by X-ray absorption spectroscopy enabled identification of a novel bis-Mo-MPT intermediate on MobA prior to nucleotide attachment. The addition of Mg-GTP to MobA loaded with bis-Mo-MPT resulted in formation and release of the final bis-MGD product. This cofactor was fully functional and reconstituted the catalytic activity of apo-TMAO reductase (TorA). We propose a reaction sequence for bis-MGD formation, which involves 1) the formation of bis-Mo-MPT, 2) the addition of two GMP units to form bis-MGD on MobA, and 3) the release and transfer of the mature cofactor to the target protein TorA, in a reaction that is supported by the specific chaperone TorD, resulting in an active molybdoenzyme

Metabarcoding mites : three years of elevated CO2 has no effect on oribatid assemblages in a Eucalyptus woodland

Elevated atmospheric carbon dioxide concentrations (eCO2) are known to significantly impact aboveground vegetation, with cascading effects on belowground systems. However, the effects of altered biogeochemical and hydrological cycles on belowground faunal assemblages remain unclear. Oribatid mites are abundant and hyperdiverse litter decomposers, with the potential to act as bioindicators of ecosystem disturbance. Here, we assessed oribatid mite assemblages under ambient and eCO2 (ambient +150 ppm) at the Eucalyptus Free-Air CO2 Enrichment (EucFACE) facility using high-throughput sequencing of the mitochondrial cytochrome oxidase 1 (COI) gene. Our aims were to i) quantify the effect of eCO2 on mite abundance and diversity, and ii) evaluate metabarcoding as a reliable approach for the rapid assessment of oribatid mite assemblages. Three years of eCO2 exposure resulted in lower soil pH but had no significant impact on mite abundances or oribatid mite assemblage structure. Instead, mite densities were found to correlate negatively with soil NH4+, while oribatid species richness using both morphological and metabarcoding datasets were positively correlated with total soil nitrogen content. Additionally, metabarcoding showed that oribatid mite assemblage structure was correlated with total soil carbon and nitrogen content, and soil pH. However, dissimilarity matrices based on the two approaches were not correlated, indicating that metabarcoding can be used as a proxy for shifts in assemblage structure but cannot be directly compared with morphological datasets. This study demonstrates how metabarcoding can rapidly assess belowground assemblages, providing a useful monitoring tool under current and future climate scenarios