Community composition of the rocky intertidal at Helgoland (German Bight,North Sea)

At the rocky island of Helgoland (North Sea), the distribution and abundances of intertidal communities were assessed and the effects of wave exposure and tidal height on the spatial distribution patterns of the communities were evaluated. Macroalgae and invertebrates were sampled quantitatively along line transects in three intertidal locations, a semi-exposed, an exposed and a sheltered one. The semi-exposed location was characterised by (1) Ulva spp. at the high intertidal (Ulva-community), (2) mussels and periwinkles at the mid intertidal (Mytilus-community) and (3) Corallina officinalis and mainly the large brown alga Fucus serratus at the low intertidal (Fucus-community). The exposed location encompassed the mid and low intertidal; at both zones the Fucus-community occurred. The sheltered location was characterised by (1) barnacles (Balanus-community) and (2) bryozoans, hydrozoans and mainly the large brown alga Ascophyllum nodosum (Ascophyllum-community). At the semi-exposed, but not at the exposed location the communities changed with the intertidal position. A relationship between wave exposure and the occurrence of specific communities was shown for the sheltered location; in contrast, communities of the semi-exposed and the exposed location appear to be little influenced by wave exposure directly. The community concept and the potential causes of distribution patterns of the defined communities are discussed and suggestions for a future monitoring are given. Variations in the communities at different spatial scales speak in favour of a multiple scale sampling design to monitor changes in the intertidal communities at Helgoland. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

The phosphoproteome of a Chlamydomonas reinhardtii eyespot fraction includes key proteins of the light signaling pathway

Flagellate green algae have developed a visual system, the eyespot apparatus, which allows the cell to phototax. In a recent proteomic approach, we identified 202 proteins from a fraction enriched in eyespot apparatuses of Chlamydomonas reinhardtii. Among these proteins, five protein kinases and two protein phosphatases were present, indicating that reversible protein phosphorylation occurs in the eyespot. About 20 major phosphoprotein bands were detected in immunoblots of eyespot proteins with an anti-phosphothreonine antibody. Toward the profiling of the targets of protein kinases in the eyespot fraction, we analyzed its phosphoproteome. The solubilized proteins of the eyespot fraction were treated with the endopeptidases LysC and trypsin prior to enrichment of phosphopeptides with immobilized metal-ion affinity chromatography. Phosphopeptides were analyzed by nano-liquid chromatography-electrospray ionization-mass spectrometry (MS) with MS/MS as well as neutral-loss-triggered MS/MS/MS spectra. We were able to identify 68 different phosphopeptides along with 52 precise in vivo phosphorylation sites corresponding to 32 known proteins of the eyespot fraction. Among the identified phosphoproteins are enzymes of carotenoid and fatty acid metabolism, putative signaling components, such as a SOUL heme-binding protein, a Ca(2+)-binding protein, and an unusual protein kinase, but also several proteins with unknown function. Notably, two unique photoreceptors, channelrhodopsin-1 and channelrhodopsin-2, contain three and one phosphorylation sites, respectively. Phosphorylation of both photoreceptors occurs in the cytoplasmatic loop next to their seven transmembrane regions in a similar distance to that observed in vertebrate rhodopsins, implying functional importance for regulation of these directly light-gated ion channels relevant for the photoresponses of C. reinhardtii.     

Association of common polymorphisms in GLUT9 gene with gout but not with coronary artery disease in a large case-control study

Background

Serum uric acid (UA) levels have recently been shown to be genetically influenced by common polymorphisms in the GLUT9 gene in two genome-wide association analyses of Italian and British populations. Elevated serum UA levels are often found in conjunction with the metabolic syndrome. Hyperuricemia is the major risk factor for gout and has been associated with increased cardiovascular morbidity and mortality. The aim of the present study was to further elucidate the association of polymorphisms in GLUT9 with gout and coronary artery disease (CAD) or myocardial infarction (MI). To test our hypotheses, we performed two large case-control association analyses of individuals from the German MI Family Study.

Methods and Findings

First, 665 patients with gout and 665 healthy controls, which were carefully matched for age and gender, were genotyped for four single nucleotide polymorphisms (SNPs) within or near the GLUT9 gene. All four SNPs demonstrated highly significant association with gout. SNP rs6855911, located within intron 7 of GLUT9, showed the strongest signal with a protective effect of the minor allele with an allelic odds ratio of 0.62 (95% confidence interval 0.52–0.75; p  =  3.2*10⁻⁷). Importantly, this finding was not influenced by adjustment for components of the metabolic syndrome or intake of diuretics. Secondly, 1,473 cases with severe CAD or MI and 1,241 healthy controls were tested for the same four GLUT9 SNPs. The analyses revealed, however, no significant association with CAD or with MI. Additional screening of genome-wide association data sets showed no signal for CAD or MI within the GLUT9 gene region.

Conclusion

Thus, our results provide compelling evidence that common genetic variations within the GLUT9 gene strongly influence the risk for gout but are unlikely to have a major effect on CAD or MI in a German population.     

Repeated neonatal separation stress alters the composition of neurochemically characterized interneuron subpopulations in the rodent dentate gyrus and basolateral amygdala

Emotional experience during early life has been shown to interfere with the development of excitatory synaptic networks in the prefrontal cortex, hippocampus, and the amygdala of rodents and primates. The aim of the present study was to investigate a developmental "homoeostatic synaptic plasticity" hypothesis and to test whether stress-induced changes of excitatory synaptic composition are counterbalanced by parallel changes of inhibitory synaptic networks. The impact of repeated early separation stress on the development of two GABAergic neuronal subpopulations was quantitatively analyzed in the brain of the semiprecocial rodent Octodon degus. Assuming that PARV- and CaBP-D28k-expression are negatively correlated to the level of inhibitory activity, the previously described reduced density of excitatory spine synapses in the dentate gyrus of stressed animals appears to be "amplified" by elevated GABAergic inhibition, reflected by reduced PARV- (down to 85%) and CaBP-D28k-immunoreactivity (down to 74%). In opposite direction, the previously observed elevated excitatory spine density in the CA1 region of stressed animals appears to be amplified by reduced inhibition, reflected by elevated CaPB-D28k-immunoreactivity (up to 149%). In the (baso)lateral amygdala, the previously described reduction of excitatory spine synapses appears to be "compensated" by reduced inhibitory activity, reflected by dramatically elevated PARV- (up to 395%) and CaPB-D28k-immunoreactivity (up to 327%). No significant differences were found in the central nucleus of the amygdala, the piriform, and somatosensory cortices and in the hypothalamic paraventricular nucleus. Thus during stress-evoked neuronal and synaptic reorganization, a homeostatic balance between excitation and inhibition is not maintained in all regions of the juvenile brain.     

Polyvinylamine‐coated polyester fibers as a carrier matrix for the immobilization of peroxidases

Biocatalytic transformations that employ immobilized enzymes become increasingly important for industrial applications. Synthetic or natural textile fiber materials such as polyester, polyamide or viscose are support materials that are comparatively inexpensive. Contrary to traditional support materials, their flexibility enables their use in reactors of any geometry and a fast and residue‐free removal from batch reactors. In this study a permanently immobilized peroxidase (Baylase®) has been investigated on polyester felt as a solid support as a new heterogeneous catalyst system. The polyester felt was functionalized by coating with polyvinylamine and subsequent activation with glutaraldehyde as a crosslinking agent. The enzyme load on the textile surface, the activity of the immobilized protein after repeated use as well as the storage stability was evaluated. Scanning electron micrographs and UV Vis spectroscopy made it possible to verify the enzyme immobilization on the textile surface. Furthermore, the load of immobilized peroxidase was determined by ICP OE spectrometry to be 9–12 mg per gram of textile. The activity of immobilized Baylase® remained high over 35 reaction cycles and a storage period of 8 weeks.     

Fitness change in relation to mutation number in spontaneous mutation accumulation lines of Chlamydomonas reinhardtii

Although all genetic variation ultimately stems from mutations, their properties are difficult to study directly. Here, we used multiple mutation accumulation (MA) lines derived from five genetic backgrounds of the green algae Chlamydomonas reinhardtii that have been previously subjected to whole genome sequencing to investigate the relationship between the number of spontaneous mutations and change in fitness from a nonevolved ancestor. MA lines were on average less fit than their ancestors and we detected a significantly negative correlation between the change in fitness and the total number of accumulated mutations in the genome. Likewise, the number of mutations located within coding regions significantly and negatively impacted MA line fitness. We used the fitness data to parameterize a maximum likelihood model to estimate discrete categories of mutational effects, and found that models containing one to two mutational effect categories (one neutral and one deleterious category) fitted the data best. However, the best‐fitting mutational effects models were highly dependent on the genetic background of the ancestral strain.     

Incus facet morphology in carnivorous mammals from different ecosystems: Taxonomy vs. habitat

This study is prompted by the discovery of an incus of Hyaenodon, the first known auditory ossicle of this genus and thus of any hyaenodont mammal so far. A large set of incudes of recent Carnivora, including felids, hyaenids, viverrids, herpestids, nandiniid and canids of different ecosystems, was set up for morphological comparison. This study examines especially the incudo-mallear facet. Typically, the incudo-mallear facet is composed of: (1) three articular surfaces in felids, (2) a U-shaped surface in hyaenids and (3) four surfaces in canids. Both taxonomy (on family level) and habitat (open, closed or mixed habitat preference) might have an impact on the morphology of the incus facets, the former having a major impact in our sample. The Hyaenodon incus is small, delicate and possesses an incudo-mallear facet of a general saddle-shape with two articulation facets, a large superior articulation area and a circular, inferior articulation area. Herein, its general morphology and facet shape is most similar to the felid incus morphology.     

Enrichment of anaerobic nitrate-dependent methanotrophic ‘ Candidatus Methanoperedens nitroreducens’ archaea from an Italian paddy field soil

Paddy fields are a significant source of methane and contribute up to 20% of total methane emissions from wetland ecosystems. These inundated, anoxic soils featuring abundant nitrogen compounds and methane are an ideal niche for nitrate-dependent anaerobic methanotrophs. After 2 years of enrichment with a continuous supply of methane and nitrate as the sole electron donor and acceptor, a stable enrichment dominated by ‘Candidatus Methanoperedens nitroreducens’ archaea and ‘Candidatus Methylomirabilis oxyfera’ NC10 phylum bacteria was achieved. In this community, the methanotrophic archaea supplied the NC10 phylum bacteria with the necessary nitrite through nitrate reduction coupled to methane oxidation. The results of qPCR quantification of 16S ribosomal RNA (rRNA) gene copies, analysis of metagenomic 16S rRNA reads, and fluorescence in situ hybridization (FISH) correlated well and showed that after 2 years, ‘Candidatus Methanoperedens nitroreducens’ had the highest abundance of (2.2 ± 0.4 × 108) 16S rRNA copies per milliliter and constituted approximately 22% of the total microbial community. Phylogenetic analysis showed that the 16S rRNA genes of the dominant microorganisms clustered with previously described ‘Candidatus Methanoperedens nitroreducens ANME2D’ (96% identity) and ‘Candidatus Methylomirabilis oxyfera’ (99% identity) strains. The pooled metagenomic sequences resulted in a high-quality draft genome assembly of ‘Candidatus Methanoperedens nitroreducens Vercelli’ that contained all key functional genes for the reverse methanogenesis pathway and nitrate reduction. The diagnostic mcrA gene was 96% similar to ‘Candidatus Methanoperedens nitroreducens ANME2D’ (WP_048089615.1) at the protein level. The ‘Candidatus Methylomirabilis oxyfera’ draft genome contained the marker genes pmoCAB, mdh, and nirS and putative NO dismutase genes. Whole-reactor anaerobic activity measurements with methane and nitrate revealed an average methane oxidation rate of 0.012 mmol/h/L, with cell-specific methane oxidation rates up to 0.57 fmol/cell/day for ‘Candidatus Methanoperedens nitroreducens’. In summary, this study describes the first enrichment and draft genome of methanotrophic archaea from paddy field soil, where these organisms can contribute significantly to the mitigation of methane emissions.     

Bioinformatic analysis of fold-type III PLP-dependent enzymes discovers multimeric racemases

Pyridoxal-5′-phosphate (PLP)-dependent enzymes are ubiquitous in nature and catalyze a variety of important metabolic reactions. The fold-type III PLP-dependent enzyme family is primarily comprised of decarboxylases and alanine racemases. In the development of a multiple structural alignment database (3DM) for the enzyme family, a large subset of 5666 uncharacterized proteins with high structural, but low sequence similarity to alanine racemase and decarboxylases was found. Compared to these two classes of enzymes, the protein sequences being the object of this study completely lack the C-terminal domain, which has been reported important for the formation of the dimer interface in other fold-type III enzymes. The 5666 sequences cluster around four protein templates, which also share little sequence identity to each other. In this work, these four template proteins were solubly expressed in Escherichia coli, purified, and their substrate profiles were evaluated by HPLC analysis for racemase activity using a broader range of amino acids. They were found active only against alanine or serine, where they exhibited Michaelis constants within the range of typical bacterial alanine racemases, but with significantly lower turnover numbers. As the already described racemases were proposed to be active and appeared to be monomers as judged from their crystal structures, we also investigated this aspect for the four new enzymes. Here, size exclusion chromatography indicated the presence of oligomeric states of the enzymes and a native-PAGE in-gel assay showed that the racemase activity was present only in an oligomeric state but not as monomer. This suggests the likelihood of a different behavior of these enzymes in solution compared to the one observed in crystalline form.