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Background: Mediterranean temporary water bodies are important reservoirs of biodiversity and host a unique assemblage of diapausing aquatic invertebrates. These environments are currently vanishing because of increasing human pressure. Chirocephalus kerkyrensis is a fairy shrimp typical of temporary water bodies in Mediterranean plain forests and has undergone a substantial decline in number of populations in recent years due to habitat loss. We assessed patterns of genetic connectivity and phylogeographic history in the seven extant populations of the species from Albania, Corfu Is. (Greece), Southern and Central Italy.
Methodology/Principal Findings: We analyzed sequence variation at two mitochondrial DNA genes (Cytochrome Oxidase I and 16s rRNA) in all the known populations of C. kerkyrensis. We used multiple phylogenetic, phylogeographic and coalescence-based approaches to assess connectivity and historical demography across the whole distribution range of the species. C. kerkyrensis is genetically subdivided into three main mitochondrial lineages; two of them are geographically localized (Corfu Is. and Central Italy) and one encompasses a wide geographic area (Albania and Southern Italy). Most of the detected genetic variation (approximate to 81%) is apportioned among the aforementioned lineages.
Conclusions/Significance: Multiple analyses of mismatch distributions consistently supported both past demographic and spatial expansions with the former predating the latter; demographic expansions were consistently placed during interglacial warm phases of the Pleistocene while spatial expansions were restricted to cold periods. Coalescence methods revealed a scenario of past isolation with low levels of gene flow in line with what is already known for other co-distributed fairy shrimps and suggest drift as the prevailing force in promoting local divergence. We recommend that these evolutionary trajectories should be taken in proper consideration in any effort aimed at protecting Mediterranean temporary water bodies.
In this study we used molecular markers to screen for the occurrence and prevalence of the three most common haemosporidian genera (Haemoproteus, Plasmodium, and Leucocytozoon) in blood samples of the Philippine Bulbul (Hypsipetes philippinus), a thrush-size passerine bird endemic to the Philippine Archipelago. We then used molecular data to ask whether the phylogeographic patterns in this insular host-parasite system might follow similar evolutionary trajectories or not. We took advantage of a previous study describing the pattern of genetic structuring in the Philippine Bulbul across the Central Philippine Archipelago (6 islands, 7 populations and 58 individuals; three mitochondrial DNA genes). The very same birds were here screened for the occurrence of parasites by species-specific PCR assays of the mitochondrial cytochrome b gene (471 base pairs). Twenty-eight out of the 58 analysed birds had Haemoproteus (48%) infections while just 2% of the birds were infected with either Leucocytozoon or Plasmodium. Sixteen of the 28 birds carrying Haemoproteus had multiple infections. The phylogeography of the Philippine Bulbul mostly reflects the geographical origin of samples and it is consistent with the occurrence of two different subspecies on (1) Semirara and (2) Carabao, Boracay, North Gigante, Panay, and Negros, respectively. Haemoproteus phylogeography shows very little geographical structure, suggesting extensive gene flow among locations. While movements of birds among islands seem very sporadic, we found co-occurring evolutionary divergent parasite lineages. We conclude that historical processes have played a major role in shaping the host phylogeography, while they have left no signature in that of the parasites. Here ongoing population processes, possibly multiple reinvasions mediated by other hosts, are predominant.
Hidden diversity in diatoms of Kenyan Lake Naivasha a genetic approach detects temporal variation
(2012)
This study provides insights into the morphological and genetic diversity in diatoms occurring in core sediments from tropical lakes in Kenya. We developed a genetic survey technique specific for diatoms utilizing a short region (7667 bp) of the ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit (rbcL) gene as genetic barcode. Our analyses (i) validated the use of rbcL as a barcoding marker for diatoms, applied to sediment samples, (ii) showed a significant correlation between the results obtained by morphological and molecular data and (iii) indicated temporal variation in diatom assemblages on the inter- and intra-specific level. Diatom assemblages from a short core from Lake Naivasha show a drastic shift over the last 200 years, as littoral species (e.g. Navicula) are replaced by more planktonic ones (e.g. Aulacoseira). Within that same period, we detected periodic changes in the respective frequencies of distinct haplotype groups of Navicula, which coincide with wet and dry periods of Lake Naivasha between 1820 and 1938 AD. Our genetic analyses on historical lake sediments revealed inter- and intra-specific variation in diatoms, which is partially hidden behind single morphotypes. The occurrence of particular genetic lineages is probably correlated with environmental factors.
In this study, we report the genetic population structure of the Fire-bellied toad Bombina bombina in Brandenburg (East Germany) in the context of conservation. We analysed 298 samples originating from 11 populations in Brandenburg using mitochondrial control region sequences and six polymorphic microsatellite loci. For comparison, we included one population each from Poland and Ukraine into our analysis. Within Brandenburg, we detected a moderate variability in the mitochondrial control region (19 different haplotypes) and at microsatellite loci (9-12 alleles per locus). These polymorphisms revealed a clear population structure among toads in Brandenburg, despite a relatively high overall population density and the moderate size of single populations (100-2000 individuals). The overall genetic population structure is consistent with a postglacial colonization from South East-Europe and a subsequent population expansion. Based on genetic connectivity, we infer Management Units (MUs) as targets for conservation. Our genetic survey identified MUs, within which human infrastructure is currently preventing any genetic exchange. We also detect an unintentional translocation from South East to North West Brandenburg, presumably in the course of fish stocking activities. Provided suitable conservation measures are taken, Brandenburg should continue to harbor large populations of this critically endangered species.
Fourteen microsatellite markers were isolated and characterized for the endangered Visayan tarictic hornbill (Penelopides panini, Aves: Bucerotidae). In an analysis of 76 individuals, the number of alleles per locus varied from one to 12. Expected and observed heterozygosity ranged from 0.00 to 0.87 and from 0.00 to 0.89, respectively. All primers also amplify microsatellite loci in Luzon tarictic hornbill (Penelopides manillae), Mindanao tarictic hornbill (Penelopides affinis), the critically endangered Walden's hornbill (Aceros waldeni) and the near-threatened writhed hornbill (Aceros leucocephalus). Two loci which are monomorphic in P. panini were found polymorphic in at least one of the other species. These 14 new microsatellite markers specifically developed for two genera of Philippine hornbills, in combination with those already available for the hornbill genera Buceros and Bucorvus, comprise a reasonable number of loci to genetically analyse wild and captive populations of these and probably other related, often endangered hornbills.
Rates of multiple paternities were investigated in the sailfin molly (Poecilia latipinna), using eight microsatellite loci. Genotyping was performed for offspring and mothers in 40 broods from four allopatric populations from the south-eastern U.S.A. along a geographic stretch of 1200 km in west-east direction and approximately 200 km from north to south. No significant differences regarding rates of multiple paternities were found between populations despite sample populations stemming from ecologically divergent habitats. Even the most conservative statistical approach revealed a minimum of 70% of the broods being sired by at least two males, with an average of 1.80-2.95 putative fathers per brood. Within broods, one male typically sired far more offspring than would be expected under an assumed equal probability of all detected males siring offspring.
We tested the utility of a 230 base pair intron fragment of the highly conserved nuclear gene Elongation Factor 1-alpha (EF1-alpha) as a proper marker to reconstruct the phylogeography of the marine amphipod Pontogammarus maeoticus (Sowinsky, 1894) from the Caspian and Black Seas. As a prerequisite for further analysis, we confirmed by Southern blot analysis that EF1-alpha is encoded at a single locus in P. maeoticus. We included 15 populations and 60 individuals in the study. Both the phylogeny of the 27 unique alleles found and population genetic analyses revealed a significant differentiation between populations from the aforementioned sea basins. Our results are in remarkable agreement with recent studies on a variety of species from the same area, which invariably support a major phylogeographic break between the Caspian and Black Seas. We thus conclude that our EF1-alpha intron is an informative marker for phylogeographic studies in amphipods at the shallow population level.
Background: The Visayan Tarictic Hornbill (Penelopides panini) and the Walden's Hornbill (Aceros waldeni) are two threatened hornbill species endemic to the western islands of the Visayas that constitute - between Luzon and Mindanao - the central island group of the Philippine archipelago. In order to evaluate their genetic diversity and to support efforts towards their conservation, we analyzed genetic variation in similar to 600 base pairs (bp) of the mitochondrial control region I and at 12-19 nuclear microsatellite loci. The sampling covered extant populations, still occurring only on two islands (P. panini: Panay and Negros, A. waldeni: only Panay), and it was augmented with museum specimens of extinct populations from neighboring islands. For comparison, their less endangered (= more abundant) sister taxa, the Luzon Tarictic Hornbill (P. manillae) from the Luzon and Polillo Islands and the Writhed Hornbill (A. leucocephalus) from Mindanao Island, were also included in the study. We reconstructed the population history of the two Penelopides species and assessed the genetic population structure of the remaining wild populations in all four species.
Results: Mitochondrial and nuclear data concordantly show a clear genetic separation according to the island of origin in both Penelopides species, but also unravel sporadic over-water movements between islands. We found evidence that deforestation in the last century influenced these migratory events. Both classes of markers and the comparison to museum specimens reveal a genetic diversity loss in both Visayan hornbill species, P. panini and A. waldeni, as compared to their more abundant relatives. This might have been caused by local extinction of genetically differentiated populations together with the dramatic decline in the abundance of the extant populations.
Conclusions: We demonstrated a loss in genetic diversity of P. panini and A. waldeni as compared to their sister taxa P. manillae and A. leucocephalus. Because of the low potential for gene flow and population exchange across islands, saving of the remaining birds of almost extinct local populations - be it in the wild or in captivity - is particularly important to preserve the species' genetic potential.
The ongoing global amphibian decline calls for an increase of habitat and population management efforts. Pond restoration and construction is more and more accompanied by breeding and translocation programs. However, the appropriateness of translocations as a tool for conservation has been widely debated, as it can cause biodiversity loss through genetic homogenization and can disrupt local adaptation, eventually leading to outbreeding depression. In this study, we investigated the genetic structure of two translocated populations of the critically endangered fire-bellied toad Bombina bombina at its north western distribution edge using supposedly neutral genetic markers (variation in the mitochondrial control region and microsatellites) as well as a marker under selection (major histocompatibility complex (MHC) genes). While one of the newly established populations showed the typical genetic composition of surrounding populations, the other was extremely diverged without clear affinity to its putative source. In this population we detected a profound impact of allochthonous individuals: 100% of the analyzed individuals exhibited a highly divergent mitochondrial haplotype which was otherwise found in Austria. 83% of them were also assigned to Austria by the analysis of microsatellites. Interestingly, for the adaptive marker (MHC) local alleles were predominant in this population, while only very few alleles were shared with the Austrian population. Probably Mendelian inheritance has reshuffled genotypes such that adaptive local alleles are maintained (here, MHC), while presumably neutral allochthonous alleles dominate at other loci. The release of allochthonous individuals generally increased the genetic variability of the affected population without wiping out locally adaptive genotypes. Thus, outbreeding depression might be less apparent than sometimes thought and natural selection appears strong enough to maintain locally adaptive alleles, at least in functionally important immune system genes.
Talitrids are semiterrestrial crustacean amphipods inhabiting sandy and rocky beaches; they generally show limited active dispersal over long distances. In this study we assessed levels of population genetic structure and variability in the talitrid amphipod Orchestia montagui, a species strictly associated to stranded decaying heaps of the seagrass Posidonia oceanica. The study is based on six populations (153 individuals) and covers five basins of the Mediterranean Sea (Tyrrhenian, Ionian, Adriatic, Western and Eastern basins). Samples were screened for polymorphisms at a fragment of the mitochondrial DNA (mtDNA) coding for the cytochrome oxidase subunit I gene (COI; 571 base pairs) and at eight microsatellite loci. MtDNA revealed a relatively homogeneous haplogroup, which clustered together the populations from the Western, Tyrrhenian and Eastern basins, but not the populations from the Adriatic and Ionian ones; microsatellites detected two clusters, one including the Adriatic and Ionian populations, the second grouping all the others. We found a weak geographic pattern in the genetic structuring of the species, with a lack of isolation by distance at either class of markers. Results are discussed in terms of probability of passive dispersal over long distances through heaps of seagrass.
Neuronal computation of object distance from echo delay is an essential task that echolocating bats must master for spatial orientation and the capture of prey. In the dorsal auditory cortex of bats, neurons specifically respond to combinations of short frequency-modulated components of emitted call and delayed echo. These delay-tuned neurons are thought to serve in target range calculation. It is unknown whether neuronal correlates of active space perception are established by experience-dependent plasticity or by innate mechanisms. Here we demonstrate that in the first postnatal week, before onset of echolocation and flight, dorsal auditory cortex already contains functional circuits that calculate distance from the temporal separation of a simulated pulse and echo. This innate cortical implementation of a purely computational processing mechanism for sonar ranging should enhance survival of juvenile bats when they first engage in active echolocation behaviour and flight.
A lamin in lower eukaryotes?
(2012)
Lamins are the major components of the nuclear lamina and serve not only as a mechanical support, but are also involved in chromatin organization, epigenetic regulation, transcription and mitotic events. Despite these universal tasks, lamins have so far been found only in metazoans. Yet, recently we have identified Dictyostelium NE81 as the first lamin-like protein in a lower eukaryote. Based on the current knowledge, we draw a model for nuclear envelope organization in Dictyostelium in this Extra View and we review the experimental data that justified this classification. Furthermore we provide unpublished data underscoring the requirement of posttranslational CaaX-box processing for proper protein localization at the nuclear envelope. Sequence comparison of NE81 sequences from four Dictyostelia with bona fide lamins illustrates the evolutional relationship between these proteins. Under certain conditions these usually unicellular social amoebae congregate to form a multicellular body. We propose that the evolution of the lamin-like NE81 went along with the invention of multicellularity.
Dictyostelium centrosomes consist of a layered core structure surrounded by a microtubule-nucleating corona. At the G2/M transition, the corona dissociates and the core structure duplicates, yielding two spindle pole bodies. Finally, in telophase, the spindle poles mature into two new, complete centrosomes. CP55 was identified in a centrosomal proteome analysis. It is a component of the centrosomal core structure, and persists at the centrosome throughout the entire cell cycle. FRAP experiments revealed that during interphase the majority of centrosomal GFP-CP55 is immobile, which indicates a structural task of CP55 at the centrosome. The CP55null mutant is characterized by increased ploidy, a less structured, slightly enlarged corona, and by supernumerary, cytosolic MTOCs, containing only corona proteins and lacking a core structure. Live cell imaging showed that supernumerary MTOCs arise in telophase. Lack of CP55 also caused premature recruitment of the corona organizer CP148 to mitotic spindle poles, already in metaphase instead of telophase. Forces transmitted through astral microtubules may expel prematurely acquired or loosely attached corona fragments into the cytosol, where they act as independent MTOCs. CP55null cells were also impaired in growth, most probably due to difficulties in centrosome splitting during prophase. Furthermore, although they were still capable of phagocytosis, they appeared unable to utilize phagocytosed nutrients. This inability may be attributed to their partially disorganized Golgi apparatus.
The centrosome consists of a layered core structure surrounded by a microtubule-nucleating corona. A tight linkage through the nuclear envelope connects the cytosolic centrosome with the clustered centromeres within the nuclear matrix. At G2/M the corona dissociates, and the core structure duplicates, yielding two spindle poles. CP148 is a novel coiled coil protein of the centrosomal corona. GFP-CP148 exhibited cell cycle-dependent presence and absence at the centrosome, which correlates with dissociation of the corona in prophase and its reformation in late telophase. During telophase, GFP-CP148 formed cytosolic foci, which coalesced and joined the centrosome. This explains the hypertrophic appearance of the corona upon strong overexpression of GFP-CP148. Depletion of CP148 by RNAi caused virtual loss of the corona and disorganization of interphase microtubules. Surprisingly, formation of the mitotic spindle and astral microtubules was unaffected. Thus, microtubule nucleation complexes associate with centrosomal core components through different means during interphase and mitosis. Furthermore, CP148 RNAi caused dispersal of centromeres and altered Sun1 distribution at the nuclear envelope, suggesting a role of CP148 in the linkage between centrosomes and centromeres. Taken together, CP148 is an essential factor for the formation of the centrosomal corona, which in turn is required for centrosome/centromere linkage.
Characterization of NE81, the first lamin-like nucleoskeleton protein in a unicellular organism
(2012)
Lamins build the nuclear lamina and are required for chromatin organization, gene expression, cell cycle progression, and mechanical stabilization. Despite these universal functions, lamins have so far been found only in metazoans. We have identified protein NE81 in Dictyostelium, which has properties that justify its denomination as a lamin-like protein in a lower eukaryote. This is based on its primary structure, subcellular localization, and regulation during mitosis, and its requirement of the C-terminal CaaX box as a posttranslational processing signal for proper localization. Our knockout and overexpression mutants revealed an important role for NE81 in nuclear integrity, chromatin organization, and mechanical stability of cells. All our results are in agreement with a role for NE81 in formation of a nuclear lamina. This function is corroborated by localization of Dictyostelium NE81 at the nuclear envelope in human cells. The discovery of a lamin-like protein in a unicellular organism is not only intriguing in light of evolution, it may also provide a simple experimental platform for studies of the molecular basis of laminopathies.
The present study reports a facile approach for sulfite biosensing, based on enhanced direct electron transfer of a human sulfite oxidase (hSO) immobilized on a gold nanoparticles modified electrode. The spherical core shell AuNPs were prepared via a new method by reduction of HAuCl4 with branched poly(ethyleneimine) in an ionic liquids resulting particles with a diameter less than 10 nm. These nanoparticles were covalently attached to a mercaptoundecanoic acid modified Au-electrode where then hSO was adsorbed and an enhanced interfacial electron transfer and electrocatalysis was achieved. UV/Vis and resonance Raman spectroscopy, in combination with direct protein voltammetry, are employed for the characterization of the system and reveal no perturbation of the structural integrity of the redox protein. The proposed biosensor exhibited a quick steady-state current response, within 2 s, a linear detection range between 0.5 and 5.4 mu M with a high sensitivity (1.85 nA mu M-1). The investigated system provides remarkable advantages in the possibility to work at low applied potential and at very high ionic strength. Therefore these properties could make the proposed system useful in the development of bioelectronic devices and its application in real samples.
As significant differences between sexes were found in the susceptibility to alcoholic liver disease in human and animal models, it was the aim of the present study to investigate whether female mice also are more susceptible to the development of nonalcoholic fatty liver disease (NAFLD). Male and female C57BL/6J mice were fed either water or 30% fructose solution ad libitum for 16 wks. Liver damage was evaluated by histological scoring. Portal endotoxin levels and markers of Kupffer cell activation and insulin resistance, plasminogen activator inhibitor 1 (PAI-1) and phosphorylated adenosine monophosphate-activated protein kinase (pAMPK) were measured in the liver. Adiponectin mRNA expression was determined in adipose tissue. Hepatic steatosis was almost similar between male and female mice; however, inflammation was markedly more pronounced in livers of female mice. Portal endotoxin levels, hepatic levels of myeloid differentiation primary response gene (88) (MyD88) protein and of 4-hydroxynonenal protein adducts were elevated in animals with NAFLD regardless of sex. Expression of insulin receptor substrate 1 and 2 was decreased to a similar extent in livers of male and female mice with NAFLD. The less pronounced susceptibility to liver damage in male mice was associated with a superinduction of hepatic pAMPK in these mice whereas, in livers of female mice with NAFLD, PAI-1 was markedly induced. Expression of adiponectin in visceral fat was significantly lower in female mice with NAFLD but unchanged in male mice compared with respective controls. In conclusion, our data suggest that the sex-specific differences in the susceptibility to NAFLD are associated with differences in the regulation of the adiponectin-AMPK-PAI-1 signaling cascade. Online address: http://www.molmed.Org doi: 10.2119/molmed.2012.00223
Ectoparasites of bats in Mongolia : Part 2 (Ischnopsyllidae, Nycteribiidae, Cimicidae and Acari)
(2012)
This study analyses ectoparasites found on Mongolian bats between 2008 and 2011. We examined 12 different bat species, with a total of 23 ectoparasite species present. Apart from reporting distributions, we also discuss specific host-parasite relationships. Owing to recent taxonomic changes splitting the Myotis mystacinus-group into several new taxa, their corresponding ectoparasite fauna could also be addressed in detail. Introducing ectoparasitic insects at length elsewhere (SCHEFFLER et al., 2010), this paper focuses on the analysis of parasitic Acari. Additional findings for Spinturnicidae (wing mites) and Macronyssidae broadened the spectrum of known parasites. Altogether, the knowledge of bat ectoparasites from Mongolia remains very sketchy. Based on different examples, we discuss current taxonomic problems regarding the species status of parasites, and suggest avenues for future research.
The present study provides a review of the published ectoparasitic species of European bats. On the basis of own researches in Germany, the abundance of each parasite was analysed. For every bat species the community of the dominant parasites and the quality of the host-parasite relation were summarized. Part 2 is concerned with the ectoparasites of the Greater mouse-eared bat (Myotis myotis), of the Lesser mouse-eared bat (Myotis blythii), of the Long-fingered bat (Myotis capaccinii), of the Whiskered bat (Myotis mystacinus), and of the Natterer`s bat (Myotis nattereri).
Die vorliegende Arbeit liefert eine Zusammenstellung der bisher bekannten Ektoparasitenarten europaeischer Fledermausarten. Fuer jede Wirtsart wird das Artenspektrum der haeufig vorkommenden Ektoparasiten angegeben und die Spezifitaet der Wirt-Parasit-Beziehung diskutiert. Teil 3 behandelt die Ektoparasiten der Wimperfledermaus (Myotis emarginatus), der Langfluegelfledermaus (Miniopterus schreibersii), der Großen Hufeisennase (Rhinolophus ferrumequinum), der Kleinen Hufeisennase (Rhinolophus hipposideros), der Blasius-Hufeisennase (Rhinolophus blasii), der Mittelmeerhufeisennase (Rhinolophus euryale) und der Mehely-Hufeisennase (Rhinolophus mehelyi).
Ellenberg indicator values are widely used ecological tools to elucidate relationships between vegetation and environment in ecological research and environmental planning. However, they are mainly deduced from expert knowledge on plant species and are thus subject of ongoing discussion. We researched if Ellenberg indicator values can be directly extracted from the vegetation biomass itself. Mean Ellenberg "moisture" (mF) and "nitrogen" (mN) values of 141 grassland plots were related to nutrient concentrations, fibre fractions and spectral information of the aboveground biomass. We developed calibration models for the prediction of mF and mN using spectral characteristics of biomass samples with near-infrared reflectance spectroscopy (NIRS). Prediction goodness was evaluated with internal cross-validations and with an external validation data set. NIRS could accurately predict Ellenberg mN, and with less accuracy Ellenberg mF. Predictions were not more precise for cover-weighted Ellenberg values compared with un-weighted values. Both Ellenberg mN and mF showed significant and strong correlations with some of the nutrient and fibre concentrations in the biomass. Against expectations, Ellenberg mN was more closely related to phosphorus than to nitrogen concentrations, suggesting that this value rather indicates productivity than solely nitrogen. To our knowledge we showed for the first time that mean Ellenberg indicator values could be directly predicted from the aboveground biomass, which underlines the usefulness of the NIRS technology for ecological studies, especially in grasslands ecosystems.
To identify genetic variants associated with head circumference in infancy, we performed a meta-analysis of seven genome-wide association studies (GWAS) (N = 10,768 individuals of European ancestry enrolled in pregnancy and/or birth cohorts) and followed up three lead signals in six replication studies (combined N = 19,089). rs7980687 on chromosome 12q24 (P = 8.1 x 10(-9)) and rs1042725 on chromosome 12q15 (P = 2.8 x 10(-10)) were robustly associated with head circumference in infancy. Although these loci have previously been associated with adult height(1), their effects on infant head circumference were largely independent of height (P = 3.8 x 10(-7) for rs7980687 and P = 1.3 x 10(-7) for rs1042725 after adjustment for infant height). A third signal, rs11655470 on chromosome 17q21, showed suggestive evidence of association with head circumference (P = 3.9 x 10(-6)). SNPs correlated to the 17q21 signal have shown genome-wide association with adult intracranial volume(2), Parkinson's disease and other neurodegenerative diseases(3-5), indicating that a common genetic variant in this region might link early brain growth with neurological disease in later life.
Habitat loss poses a severe threat to biodiversity. While many studies yield valuable information on how specific species cope with such environmental modification, the mechanistic understanding of how interacting species or whole communities are affected by habitat loss is still poor. Individual movement plays a crucial role for the space use characteristics of species, since it determines how individuals perceive and use their heterogeneous environment. At the community level, it is therefore essential to include individual movement and how it is influenced by resource sharing into the investigation of consequences of habitat loss. To elucidate the effects of foraging movement on communities in face of habitat loss, we here apply a recently published spatially-explicit and individual-based model of home range formation. This approach allows predicting the individual size distribution (ISD) of mammal communities in simulation landscapes that vary in the amount of suitable habitat. We apply three fundamentally different foraging movement approaches (central place forager (CPF), patrolling forager (PF) and body mass dependent nomadic forager (BNF)). Results show that the efficiency of the different foraging strategies depends on body mass, which again affects community structure in face of habitat loss. CPF is only efficient for small animals, and therefore yields steep ISD exponents on which habitat loss has little effect (due to a movement limitation of body mass). PF and particularly BNF are more efficient for larger animals, resulting in less steep ISDs with higher mass maxima, both showing a threshold behaviour with regard to loss of suitable habitat. These findings represent a new way of explaining observed extinction thresholds, and therefore indicate the importance of individual space use characterized by physiology and behaviour, i.e. foraging movement, for communities and their response to habitat loss. Findings also indicate the necessity to incorporate the crucial role of movement into future conservation efforts of terrestrial communities.
The uptake of potassium ions (K+) accompanied by an acidification of the apoplasm is a prerequisite for stomatal opening. The acidification (approximately 2-2.5 pH units) is perceived by voltage-gated inward potassium channels (K-in) that then can open their pores with lower energy cost. The sensory units for extracellular pH in stomatal K-in channels are proposed to be histidines exposed to the apoplasm. However, in the Arabidopsis thaliana stomatal K-in channel KAT1, mutations in the unique histidine exposed to the solvent (His(267)) do not affect the pH dependency. We demonstrate in the present study that His(267) of the KAT1 channel cannot sense pH changes since the neighbouring residue Phe(266) shifts its pK(a) to undetectable values through a cation-pi interaction. Instead, we show that Glu(240) placed in the extracellular loop between transmembrane segments S5 and S6 is involved in the extracellular acid activation mechanism. Based on structural models we propose that this region may serve as a molecular link between the pH- and the voltage-sensor. Like Glu(240), several other titratable residues could contribute to the pH-sensor of KAT1, interact with each other and even connect such residues far away from the voltage-sensor with the gating machinery of the channel.
1. Atmospheric nitrogen (N) deposition is expected to change forest understorey plant community composition and diversity, but results of experimental addition studies and observational studies are not yet conclusive. A shortcoming of observational studies, which are generally based on resurveys or sampling along large deposition gradients, is the occurrence of temporal or spatial confounding factors.
2. We were able to assess the contribution of N deposition versus other ecological drivers on forest understorey plant communities by combining a temporal and spatial approach. Data from 1205 (semi-)permanent vegetation plots taken from 23 rigorously selected understorey resurvey studies along a large deposition gradient across deciduous temperate forest in Europe were compiled and related to various local and regional driving factors, including the rate of atmospheric N deposition, the change in large herbivore densities and the change in canopy cover and composition.
3. Although no directional change in species richness occurred, there was considerable floristic turnover in the understorey plant community and a shift in species composition towards more shade-tolerant and nutrient-demanding species. However, atmospheric N deposition was not important in explaining the observed eutrophication signal. This signal seemed mainly related to a shift towards a denser canopy cover and a changed canopy species composition with a higher share of species with more easily decomposed litter.
4. Synthesis. Our multi-site approach clearly demonstrates that one should be cautious when drawing conclusions about the impact of atmospheric N deposition based on the interpretation of plant community shifts in single sites or regions due to other, concurrent, ecological changes. Even though the effects of chronically increased N deposition on the forest plant communities are apparently obscured by the effects of canopy changes, the accumulated N might still have a significant impact. However, more research is needed to assess whether this N time bomb will indeed explode when canopies will open up again.
Mesenchymal stem cells and glioma cells form a structural as well as a functional syncytium in vitro
(2012)
The interaction of human mesenchymal stem cells (hMSCs) and tumor cells has been investigated in various contexts. HMSCs are considered as cellular treatment vectors based on their capacity to migrate towards a malignant lesion. However, concerns about unpredictable behavior of transplanted hMSCs are accumulating. In malignant gliomas, the recruitment mechanism is driven by glioma-secreted factors which lead to accumulation of both, tissue specific stem cells as well as bone marrow derived hMSCs within the tumor. The aim of the present work was to study specific cellular interactions between hMSCs and glioma cells in vitro. We show, that glioma cells as well as hMSCs differentially express connexins. and that they interact via gap-junctional coupling. Besides this so-called functional syncytium formation, we also provide evidence of cell fusion events (structural syncytium). These complex cellular interactions led to an enhanced migration and altered proliferation of both, tumor and mesenchymal stem cell types in vitro. The presented work shows that glioma cells display signs of functional as well as structural syncytium formation with hMSCs in vitro. The described cellular phenomena provide new insight into the complexity of interaction patterns between tumor cells and host cells. Based on these findings, further studies are warranted to define the impact of a functional or structural syncytium formation on malignant tumors and cell based therapies in vivo.
Bacteriophages use specific tail proteins to recognize host cells. It is still not understood to molecular detail how the signal is transmitted over the tail to initiate infection. We have analysed in vitro DNA ejection in long-tailed siphovirus 9NA and short-tailed podovirus P22 upon incubation with Salmonella typhimurium lipopolysaccharide (LPS). We showed for the first time that LPS alone was sufficient to elicit DNA release from a siphovirus in vitro. Crystal structure analysis revealed that both phages use similar tailspike proteins for LPS recognition. Tailspike proteins hydrolyse LPS O antigen to position the phage on the cell surface. Thus we were able to compare in vitro DNA ejection processes from two phages with different morphologies with the same receptor under identical experimental conditions. Siphovirus 9NA ejected its DNA about 30 times faster than podovirus P22. DNA ejection is under control of the conformational opening of the particle and has a similar activation barrier in 9NA and P22. Our data suggest that tail morphology influences the efficiencies of particle opening given an identical initial receptor interaction event.
Plant community assembly in temperate forests along gradients of soil fertility and disturbance
(2012)
Plant community assembly from a regional pool is largely driven by two mechanisms: environmental filtering and niche partitioning, which result in trait convergence or divergence, respectively. Although empirical evidence for both assembly mechanisms exists, the environmental conditions and traits where each of the two assembly patterns is prevalent remain unclear. We studied community assembly mechanisms in herb layer communities of temperate forest patches in NW Germany, looking at distributions of competitive and reproductive traits along gradients of soil fertility and disturbance. We also examined how community assembly patterns changed over a time span of two decades. Canopy height converged toward taller species with increasing soil fertility and increasing light availability. Most reproductive traits diverged with an increasing degree of disturbance and with increasing fertility. Comparisons over time indicated that disturbance events induced the coexistence of species with different reproductive strategies and also selected for tall species as a result of enhanced competitive pressure. Our study demonstrates that in accordance with existing hypotheses, competitive traits (e.g., canopy height) can be convergent in favorable environments. However, this convergence is associated with a divergence of traits related to other challenges (e.g., reproduction), indicating that true functional redundancy within communities does not exist. Moreover, our study shows that the expected divergence of reproductive traits at disturbed sites can be accompanied by a convergence of other traits (e.g., canopy height), indicating that several assembly mechanisms can operate simultaneously.
The vesicle-inducing protein in plastids (VIPP1) was suggested to play a role in thylakoid membrane formation via membrane vesicles. As this functional assignment is under debate, we investigated the function of VIPP1 in Chlamydomonas reinhardtii. Using immunofluorescence, we localized VIPP1 to distinct spots within the chloroplast. In VIPP1-RNA interference/artificial microRNA cells, we consistently observed aberrant, prolamellar body-like structures at the origin of multiple thylakoid membrane layers, which appear to coincide with the immunofluorescent VIPP1 spots and suggest a defect in thylakoid membrane biogenesis. Accordingly, using quantitative shotgun proteomics, we found that unstressed vipp1 mutant cells accumulate 14 to 20% less photosystems, cytochrome b(6)f complex, and ATP synthase but 30% more light-harvesting complex II than control cells, while complex assembly, thylakoid membrane ultrastructure, and bulk lipid composition appeared unaltered. Photosystems in vipp1 mutants are sensitive to high light, which coincides with a lowered midpoint potential of the Q(A)/Q(A)(-) redox couple and increased thermosensitivity of photosystem II (PSII), suggesting structural defects in PSII. Moreover, swollen thylakoids, despite reduced membrane energization, in vipp1 mutants grown on ammonium suggest defects in the supermolecular organization of thylakoid membrane complexes. Overall, our data suggest a role of VIPP1 in the biogenesis/assembly of thylakoid membrane core complexes, most likely by supplying structural lipids.
The enzyme penicillin G acylase (EC 3.5.1.11) catalyzes amide-bond cleavage in benzylpenicillin (penicillin G) to yield 6-aminopenicillanic acid, an intermediate chemical used in the production of semisynthetic penicillins. A thermostable penicillin G acylase from Alcaligenes faecalis (AfPGA) has been crystallized using the hanging-drop vapour-diffusion method in two different space groups: C2221, with unit-cell parameters a = 72.9, b = 86.0, c = 260.2 angstrom, and P41212, with unit-cell parameters a = b = 85.6, c = 298.8 angstrom. Data were collected at 293 K and the structure was determined using the molecular-replacement method. Like other penicillin acylases, AfPGA belongs to the N-terminal nucleophilic hydrolase superfamily, has undergone post-translational processing and has a serine as the N-terminal residue of the beta-chain. A disulfide bridge has been identified in the structure that was not found in the other two known penicillin G acylase structures. The presence of the disulfide bridge is perceived to be one factor that confers higher stability to this enzyme.
The global expansion of species beyond their ancestral ranges can derive from mechanisms that are trait-based (e.g., post-establishment evolved differences compared to home populations) or circumstantial (e.g., propagule pressure, with no trait-based differences). These mechanisms can be difficult to distinguish following establishment, but each makes unique predictions regarding trait similarity between ancestral ('home') and introduced ('away') populations. Here, we tested for trait-based population differences across four continents for the globally distributed grass Dactylis glomerata, to assess the possible role of trait evolution in its worldwide expansion. We used a common-environment glasshouse experiment to quantify trait differences among home and away populations, and the potential relevance of these differences for competitive interactions. Few significant trait differences were found among continents, suggesting minimal change during global expansion. All populations were polyploids, with similar foliar carbon:nitrogen ratios (a proxy for defense), chlorophyll content, and biomass. Emergence time and growth rate favored home populations, resulting in their competitive superiority over away populations. Small but significant trait differences among away populations suggest different introductory histories or local adaptive responses following establishment. In summary, the worldwide distribution of this species appears to have arisen from its pre-adapted traits promoting growth, and its repeated introduction with cultivation and intense propagule pressure. Global expansion can thus occur without substantial shifts in growth, reproduction, or defense. Rather than focusing strictly on the invader, invasion success may also derive from the traits found (or lacking) in the recipient community and from environmental context including human disturbance.
Dynamic regulatory on/off minimization for biological systems under internal temporal perturbations
(2012)
Background: Flux balance analysis (FBA) together with its extension, dynamic FBA, have proven instrumental for analyzing the robustness and dynamics of metabolic networks by employing only the stoichiometry of the included reactions coupled with adequately chosen objective function. In addition, under the assumption of minimization of metabolic adjustment, dynamic FBA has recently been employed to analyze the transition between metabolic states.
Results: Here, we propose a suite of novel methods for analyzing the dynamics of (internally perturbed) metabolic networks and for quantifying their robustness with limited knowledge of kinetic parameters. Following the biochemically meaningful premise that metabolite concentrations exhibit smooth temporal changes, the proposed methods rely on minimizing the significant fluctuations of metabolic profiles to predict the time-resolved metabolic state, characterized by both fluxes and concentrations. By conducting a comparative analysis with a kinetic model of the Calvin-Benson cycle and a model of plant carbohydrate metabolism, we demonstrate that the principle of regulatory on/off minimization coupled with dynamic FBA can accurately predict the changes in metabolic states.
Conclusions: Our methods outperform the existing dynamic FBA-based modeling alternatives, and could help in revealing the mechanisms for maintaining robustness of dynamic processes in metabolic networks over time.
Biotic homogenization, the decrease in beta diversity among formerly distinct species assemblages, has been recognized as an important form of biotic impoverishment for more than a decade. Although researchers have stressed the importance of the functional dimension to understand its potential ecological consequences, biotic homogenization has mostly been studied at a taxonomic level. Here, we explore the relationship between taxonomic and functional homogenization using data on temperate forest herb layer communities in NW Germany, for which taxonomic homogenization has recently been demonstrated. We quantified beta diversity by partitioning Rao's quadratic entropy. We found a general positive relationship between changes in taxonomic and functional beta diversity. This relationship was stronger if multiple functional traits were taken into account. Averaged across sites, however, taxonomic homogenization was not consistently accompanied by functional homogenization. Depending on the traits considered, taxonomic homogenization occurred also together with functional differentiation or no change in functional beta diversity. The species shifts responsible for changes in beta diversity differed substantially between taxonomic and functional beta diversity measures and also among functional beta diversity measures based on different traits. We discuss likely environmental drivers for species shifts. Our study demonstrates that functional homogenization must be explicitly studied as an independent phenomenon that cannot be inferred from taxonomic homogenization.
'Biodiversity' is increasingly perceived as an important resource for research and conservation, but also for economy. Conservation, access and sustainable use of biodiversity (genetic resources, species, samples) are negotiated on different political levels, resulting in an internationally binding legal framework. Resulting legislation is binding for all parties involved in biological sampling, i.e. researches and (and in italics) countries, and especially applies for tissue or DNA samples and dervied products thereof. Understanding and awareness of export and import permits for biological samples is increasingly important for biologists to perform research projects legally and timely. Nevertheless, some biologists are still exporting and importing biological samples ignoring or noncompliant with national and international legislation, conventions, and regulations. Resulting difficulties may not only cause serious problems during field work, but may also delay the export, import or exchange of samples. Comprehensive a priori information regarding legal requirements helps to avoid or at least diminish potential problems. We identified four major factors facilitating export/import permits: (1) good personal (mutually trusted) contacts in the country of origin, (2) understanding and compliance with all relevant laws and regulations; (3) access to information regarding knowledge on permits, regulations and laws including their circulation within the researcher communities; and (4) access to consistent and up to date regulations
It has been predicted that Europe will experience a rise in temperature of 2.2-5.3 A degrees C within this century. This increase in temperature may lead to vegetation change along altitudinal gradients. To test whether vegetation composition has already changed in the recent decade due to current warming (and other concomitant environmental changes), we recorded plant species composition in 1995 and 2005/2006 in Swiss pre-alpine fen meadows (800-1,400 m a.s.l.). Despite no obvious changes in the management of these fens, overall, plant species richness (cumulative number of plant species at five plots per site) significantly increased over this period. This was mainly due to an increase in the number of thermophilous, rich-soil-indicator and shade-indicator species, which corresponded to increased community productivity and shading within the vegetation layer. In contrast, fen specialists significantly declined in species numbers. The strongest species shifts occurred at the lowest sites, which overall had a higher colonization rate by new species than did sites at higher altitudes. Vegetation change along the altitudinal gradient was also affected by different types of land management: early-flowering species and species with low habitat specificity had high colonization rates in grazed fens, especially at low altitudes.
The global warming potential of nitrous oxide (N2O) and its long atmospheric lifetime mean its presence in the atmosphere is of major concern, and that methods are required to measure and reduce emissions. Large spatial and temporal variations means, however, that simple extrapolation of measured data is inappropriate, and that other methods of quantification are required. Although process-based models have been developed to simulate these emissions, they often require a large amount of input data that is not available at a regional scale, making regional and global emission estimates difficult to achieve. The spatial extent of organic soils means that quantification of emissions from these soil types is also required, but will not be achievable using a process-based model that has not been developed to simulate soil water contents above field capacity or organic soils. The ECOSSE model was developed to overcome these limitations, and with a requirement for only input data that is readily available at a regional scale, it can be used to quantify regional emissions and directly inform land-use change decisions. ECOSSE includes the major processes of nitrogen (N) turnover, with material being exchanged between pools of SOM at rates modified by temperature, soil moisture, soil pH and crop cover. Evaluation of its performance at site-scale is presented to demonstrate its ability to adequately simulate soil N contents and N2O emissions from cropland soils in Europe. Mitigation scenarios and sensitivity analyses are also presented to demonstrate how ECOSSE can be used to estimate the impact of future climate and land-use change on N2O emissions.
The response of forest plant regeneration to temperature variation along a latitudinal gradient
(2012)
The response of forest herb regeneration from seed to temperature variations across latitudes was experimentally assessed in order to forecast the likely response of understorey community dynamics to climate warming.
Seeds of two characteristic forest plants (Anemone nemorosa and Milium effusum) were collected in natural populations along a latitudinal gradient from northern France to northern Sweden and exposed to three temperature regimes in growth chambers (first experiment). To test the importance of local adaptation, reciprocal transplants were also made of adult individuals that originated from the same populations in three common gardens located in southern, central and northern sites along the same gradient, and the resulting seeds were germinated (second experiment). Seedling establishment was quantified by measuring the timing and percentage of seedling emergence, and seedling biomass in both experiments.
Spring warming increased emergence rates and seedling growth in the early-flowering forb A. nemorosa. Seedlings of the summer-flowering grass M. effusum originating from northern populations responded more strongly in terms of biomass growth to temperature than southern populations. The above-ground biomass of the seedlings of both species decreased with increasing latitude of origin, irrespective of whether seeds were collected from natural populations or from the common gardens. The emergence percentage decreased with increasing home-away distance in seeds from the transplant experiment, suggesting that the maternal plants were locally adapted.
Decreasing seedling emergence and growth were found from the centre to the northern edge of the distribution range for both species. Stronger responses to temperature variation in seedling growth of the grass M. effusum in the north may offer a way to cope with environmental change. The results further suggest that climate warming might differentially affect seedling establishment of understorey plants across their distribution range and thus alter future understorey plant dynamics.
Salivary gland cells of the blowfly Calliphora vicina have a vacuolar-type H+-ATPase (V-ATPase) that lies in their apical membrane and energizes the secretion of a KCl-rich primary saliva upon stimulation with serotonin (5-hydroxytryptamine). Whether and to what extent V-ATPase contributes to intracellular pH (pH(i)) regulation in unstimulated gland cells is unknown. We used the fluorescent dye BCECF to study intracellular pH(i) regulation microfluorometrically and show that: (1) under resting conditions, the application of Na+-free physiological saline induces an intracellular alkalinization attributable to the inhibition of the activity of a Na+-dependent glutamate transporter; (2) the maintenance of resting pHi is Na+, Cl-, concanamycin A and DIDS sensitive; (3) recovery from an intracellular acid load is Na+ sensitive and requires V-ATPase activity; (4) the Na+/H+ antiporter is not involved in pHi recovery after a NH4Cl prepulse; and (5) at least one Na+-dependent transporter and the V-ATPase maintain recovery from an intracellular acid load. Thus, under resting conditions, the V-ATPase and at least one Na+-dependent transporter maintain normal pH(i) values of pH.7.5. We have also detected the presence of a Na+-dependent glutamate transporter, which seems to act as an acid loader. Despite this not being a common pH(i)-regulating transporter, its activity affects steady-state pH(i) in C. vicina salivary gland cells.
Organisms eating each other are only one of many types of well documented and important interactions among species. Other such types include habitat modification, predator interference and facilitation. However, ecological network research has been typically limited to either pure food webs or to networks of only a few (<3) interaction types. The great diversity of non-trophic interactions observed in nature has been poorly addressed by ecologists and largely excluded from network theory. Herein, we propose a conceptual framework that organises this diversity into three main functional classes defined by how they modify specific parameters in a dynamic food web model. This approach provides a path forward for incorporating non-trophic interactions in traditional food web models and offers a new perspective on tackling ecological complexity that should stimulate both theoretical and empirical approaches to understanding the patterns and dynamics of diverse species interactions in nature.
The large scale production of a monoclonal anti-progesterone antibody in serum free medium followed by affinity chromatography on protein G lead to a contamination of the antibody sample with a protein of about 14 kDa. This protein was identified by mass spectrometry as secretory leukocyte protease inhibitor (SLPI). This SLPI contamination lead to a failure of the fiber-optic based competitive fluorescence assay to detect progesterone in milk. Purification of the monoclonal antibody using protein A columns circumvented this problem.
Background: There are alternative substrates to the mitochondrial respiration.
Results: Data-driven model-based analysis renders predictions of alternative substrates to the mitochondrial respiration.
Conclusion: Metabolomics data in conjunction with flux-based models can discriminate among hypotheses based on enzymology alone.
Significance: This analysis provides a basic framework for in silico studies of alternative pathways in metabolism.
The globally threatened Aquatic Warbler Acrocephalus paludicola is a Palearctic-African long-distance migrant that undergoes a complete moult while wintering in Africa. Little is known about the timing of moult and the birds' mobility during moulting periods. We conducted the first study on the moult of Aquatic Warblers, in the Djoudj area of Senegal, West Africa. Wing moult scores from 36 to 90 and raggedness scores from 0 to 25 were recorded in December and January. No moulting Aquatic Warblers were caught after January. Body-feather moult was observed during and shortly after wing moult until January. We conclude that Aquatic Warblers follow the typical sequence of passerine moult, with remige moult starting in October or November. To find out how moult affects their mobility, we measured the net distance that Aquatic Warblers equipped with radio transmitters travelled in 15-min intervals. In our small sample of eight birds, the mean path length was 34 m, and there was no obvious difference between the path lengths in moulting and non-moulting individuals. We conclude that, possibly, moult does not affect the mobility and flight ability of Aquatic Warblers in general. Further research is needed to locate other wintering grounds, e.g. in the Inner Niger Delta, and reproduce our study in other populations.
Indoor mesocosm experiments were conducted to test for potential climate change effects on the spring succession of Baltic Sea plankton. Two different temperature (Delta 0 A degrees C and Delta 6 A degrees C) and three light scenarios (62, 57 and 49 % of the natural surface light intensity on sunny days), mimicking increasing cloudiness as predicted for warmer winters in the Baltic Sea region, were simulated. By combining experimental and modeling approaches, we were able to test for a potential dietary mismatch between phytoplankton and zooplankton. Two general predator-prey models, one representing the community as a tri-trophic food chain and one as a 5-guild food web were applied to test for the consequences of different temperature sensitivities of heterotrophic components of the plankton. During the experiments, we observed reduced time-lags between the peaks of phytoplankton and protozoan biomass in response to warming. Microzooplankton peak biomass was reached by 2.5 day A degrees C-1 earlier and occurred almost synchronously with biomass peaks of phytoplankton in the warm mesocosms (Delta 6 A degrees C). The peak magnitudes of microzooplankton biomass remained unaffected by temperature, and growth rates of microzooplankton were higher at Delta 6 A degrees C (mu(a dagger 0 A degrees C) = 0.12 day(-1) and mu(a dagger 6 A degrees C) = 0.25 day(-1)). Furthermore, warming induced a shift in microzooplankton phenology leading to a faster species turnover and a shorter window of microzooplankton occurrence. Moderate differences in the light levels had no significant effect on the time-lags between autotrophic and heterotrophic biomass and on the timing, biomass maxima and growth rate of microzooplankton biomass. Both models predicted reduced time-lags between the biomass peaks of phytoplankton and its predators (both microzooplankton and copepods) with warming. The reduction of time-lags increased with increasing Q(10) values of copepods and protozoans in the tritrophic food chain. Indirect trophic effects modified this pattern in the 5-guild food web. Our study shows that instead of a mismatch, warming might lead to a stronger match between protist grazers and their prey altering in turn the transfer of matter and energy toward higher trophic levels.
We investigated the effects of warming on a natural phytoplankton community from the Baltic Sea, based on six mesocosm experiments conducted 2005-2009. We focused on differences in the dynamics of three phytoplankton size groups which are grazed to a variable extent by different zooplankton groups. While small-sized algae were mostly grazer-controlled, light and nutrient availability largely determined the growth of medium- and large-sized algae. Thus, the latter groups dominated at increased light levels. Warming increased mesozooplankton grazing on medium-sized algae, reducing their biomass. The biomass of small-sized algae was not affected by temperature, probably due to an interplay between indirect effects spreading through the food web. Thus, under the higher temperature and lower light levels anticipated for the next decades in the southern Baltic Sea, a higher share of smaller phytoplankton is expected. We conclude that considering the size structure of the phytoplankton community strongly improves the reliability of projections of climate change effects.
Recent declines in biodiversity have given new urgency to questions about the relationship between land-use change, biodiversity and ecosystem processes. Despite the existence of a large body of research on the effects of land use on species richness, it is unclear whether the effects of land use on species richness are principally direct or indirect, mediated by concomitant changes in ecosystem processes. Therefore, we compared the direct effects of land use (fertilization, mowing and grazing) on species richness with indirect ones (mediated via grassland productivity) for grasslands in central Europe. We measured the richness and above-ground biomass in 150 grassland plots in 3 regions of Germany (the so-called Biodiversity Exploratories). We used univariate and structural equation models to examine direct and indirect land-use effects. The direct effects of mowing (-0.37, effect size) and grazing (0.04) intensity on species richness were stronger compared with the indirect effects of mowing (-0.04) and grazing (-0.01). However, the strong negative effect of fertilization (-0.23) on species richness was mainly indirect, mediated by increased productivity compared with the weak direct negative effect (-0.07). Differences between regions in land-use effects showed five times weaker negative effects of mowing (-0.13) in the region with organic soils (Schorfheide-Chorin), strong overall negative effects of grazing (-0.29) for the region with organic soils opposed to a similar strong positive effect (0.30) in the Hainich-Dun region, whereas the Schwabische Alb region displayed a five times weaker positive effect (0.06) only. Further, fertilization effects on species richness were positive (0.03) for the region with organic soils compared to up to 25 times stronger negative effects in the other two regions. Synthesis. Our results clearly show the importance of studying both direct and indirect effects of land-use intensity. They demonstrate the indirect nature, via productivity, of the negative effect of fertilization intensity on plant species richness in the real-world context of management-induced gradients of intensity of fertilization, mowing and grazing. Finally, they highlight that careful consideration of regional environments is necessary before attempting to generalize land-use effects on species diversity.
We studied two pathways that involve the transfer of persulfide sulfur in humans, molybdenum cofactor biosynthesis and tRNA thiolation. Investigations using human cells showed that the two-domain protein MOCS3 is shared between both pathways. MOCS3 has an N-terminal adenylation domain and a C-terminal rhodanese-like domain. We showed that MOCS3 activates both MOCS2A and URM1 by adenylation and a subsequent sulfur transfer step for the formation of the thiocarboxylate group at the C terminus of each protein. MOCS2A and URM1 are beta-grasp fold proteins that contain a highly conserved C-terminal double glycine motif. The role of the terminal glycine of MOCS2A and URM1 was examined for the interaction and the cellular localization with MOCS3. Deletion of the C-terminal glycine of either MOCS2A or URM1 resulted in a loss of interaction with MOCS3. Enhanced cyan fluorescent protein and enhanced yellow fluorescent protein fusions of the proteins were constructed, and the fluorescence resonance energy transfer efficiency was determined by the decrease in the donor lifetime. The cellular localization results showed that extension of the C terminus with an additional glycine of MOCS2A and URM1 altered the localization of MOCS3 from the cytosol to the nucleus.
Aldehyde oxidase (AO) is a complex molybdo-flavoprotein that belongs to the xanthine oxidase family. AO is active as a homodimer, and each 150-kDa monomer binds two distinct [2Fe2S] clusters, FAD, and the molybdenum cofactor. AO has an important role in the metabolism of drugs based on its broad substrate specificity oxidizing aromatic aza-heterocycles, for example, N-1-methylnicotinamide and N-methylphthalazinium, or aldehydes, such as benzaldehyde, retinal, and vanillin. Sequencing the 35 coding exons of the human AOX1 gene in a sample of 180 Italian individuals led to the identification of relatively frequent, synonymous, missense and nonsense single-nucleotide polymorphisms (SNPs). Human aldehyde oxidase (hAOX1) was purified after heterologous expression in Escherichia coli. The recombinant protein was obtained with a purity of 95% and a yield of 50 mu g/l E. coli culture. Site-directed mutagenesis of the hAOX1 cDNA allowed the purification of protein variants bearing the amino acid changes R802C, R921H, N1135S, and H1297R, which correspond to some of the identified SNPs. The hAOX1 variants were purified and compared with the wild-type protein relative to activity, oligomerization state, and metal content. Our data show that the mutation of each amino acid residue has a variable impact on the ability of hAOX1 to metabolize selected substrates. Thus, the human population is characterized by the presence of functionally inactive hAOX1 allelic variants as well as variants encoding enzymes with different catalytic activities. Our results indicate that the presence of these allelic variants should be considered for the design of future drugs.
Metabolic networks are characterized by complex interactions and regulatory mechanisms between many individual components. These interactions determine whether a steady state is stable to perturbations. Structural kinetic modeling (SKM) is a framework to analyze the stability of metabolic steady states that allows the study of the system Jacobian without requiring detailed knowledge about individual rate equations. Stability criteria can be derived by generating a large number of structural kinetic models (SK-models) with randomly sampled parameter sets and evaluating the resulting Jacobian matrices. Until now, SKM experiments applied univariate tests to detect the network components with the largest influence on stability. In this work, we present an extended SKM approach relying on supervised machine learning to detect patterns of enzyme-metabolite interactions that act together in an orchestrated manner to ensure stability. We demonstrate its application on a detailed SK-model of the Calvin-Benson cycle and connected pathways. The identified stability patterns are highly complex reflecting that changes in dynamic properties depend on concerted interactions between several network components. In total, we find more patterns that reliably ensure stability than patterns ensuring instability. This shows that the design of this system is strongly targeted towards maintaining stability. We also investigate the effect of allosteric regulators revealing that the tendency to stability is significantly increased by including experimentally determined regulatory mechanisms that have not yet been integrated into existing kinetic models.
Background: Flux coupling analysis (FCA) has become a useful tool in the constraint-based analysis of genome-scale metabolic networks. FCA allows detecting dependencies between reaction fluxes of metabolic networks at steady-state. On the one hand, this can help in the curation of reconstructed metabolic networks by verifying whether the coupling between reactions is in agreement with the experimental findings. On the other hand, FCA can aid in defining intervention strategies to knock out target reactions.
Results: We present a new method F2C2 for FCA, which is orders of magnitude faster than previous approaches. As a consequence, FCA of genome-scale metabolic networks can now be performed in a routine manner.
Conclusions: We propose F2C2 as a fast tool for the computation of flux coupling in genome-scale metabolic networks. F2C2 is freely available for non-commercial use at https://sourceforge.net/projects/f2c2/files/.
Silver nanoparticles with identical plasmonic properties but different surface functionalities are synthesized and tested as chemically selective surface-enhanced resonance Raman (SERR) amplifiers in a two-component protein solution. The surface plasmon resonances of the particles are tuned to 413 nm to match the molecular resonance of protein heme cofactors. Biocompatible functionalization of the nanoparticles with a thin film of chitosan yields selective SERR enhancement of the anionic protein cytochrome b(5), whereas functionalization with SiO2 amplifies only the spectra of the cationic protein cytochrome c. As a result, subsequent addition of the two differently functionalized particles yields complementary information on the same mixed protein sample solution. Finally, the applicability of chitosan-coated Ag nanoparticles for protein separation was tested by in situ resonance Raman spectroscopy.
To contribute to a further insight into heterosis we applied an integrative analysis to a systems biological network approach and a quantitative genetics analysis towards biomass heterosis in early Arabidopsis thaliana development. The study was performed on the parental accessions C24 and Col-0 and the reciprocal crosses. In an over-representation analysis it was tested if the overlap between the resulting gene lists of the two approaches is significantly larger than expected by chance. Top ranked genes in the results list of the systems biological analysis were significantly over-represented in the heterotic QTL candidate regions for either hybrid as well as regarding mid-parent and best-parent heterosis. This suggests that not only a few but rather several genes that influence biomass heterosis are located within each heterotic QTL region. Furthermore, the overlapping resulting genes of the two integrated approaches were particularly enriched in biomass related pathways. A chromosome-wise over-representation analysis gave rise to the hypothesis that chromosomes number 2 and 4 probably carry a majority of the genes involved in biomass heterosis in the early development of Arabidopsis thaliana.
Inbreeding alters activities of the stress-related enzymes chitinases and beta-1,3-Glucanases
(2012)
Pathogenesis-related proteins, chitinases (CHT) and beta-1,3-glucanases (GLU), are stress proteins up-regulated as response to extrinsic environmental stress in plants. It is unknown whether these PR proteins are also influenced by inbreeding, which has been suggested to constitute intrinsic genetic stress, and which is also known to affect the ability of plants to cope with environmental stress. We investigated activities of CHT and GLU in response to inbreeding in plants from 13 Ragged Robin (Lychnis flos-cuculi) populations. We also studied whether activities of these enzymes were associated with levels of herbivore damage and pathogen infection in the populations from which the plants originated. We found an increase in pathogenesis-related protein activity in inbred plants from five out of the 13 investigated populations, which suggests that these proteins may play a role in how plants respond to intrinsic genetic stress brought about by inbreeding in some populations depending on the allele frequencies of loci affecting the expression of CHT and the past levels of inbreeding. More importantly, we found that CHT activities were higher in plants from populations with higher levels of herbivore or pathogen damage, but inbreeding reduced CHT activity in these populations disrupting the increased activities of this resistance-related enzyme in populations where high resistance is beneficial. These results provide novel information on the effects of plant inbreeding on plant-enemy interactions on a biochemical level.
Crosstalk between intracellular signalling pathways is a functionally important and widespread phenomenon in cell physiology across phyla. In the salivary gland of the blowfly, serotonin induces fluid secretion via parallel activation of both the InsP(3)/Ca2+ and the cAMP/PKA signalling pathways, which interact on multiple levels. We have determined the molecular identity of a link between both pathways that mediates a Ca2+-dependent rise of intracellular cAMP. Whereas hydrolysis of cAMP via phosphodiesterases is largely independent of Ca2+, cAMP synthesis by adenylyl cyclases (AC) is potentiated in a Ca2+/calmodulin (Ca2+/CaM)-dependent manner. The existence of a Ca2+/CaM-dependent AC is supported by physiological data and a molecular approach. We have cloned Cv rutabaga cDNA, encoding the first blowfly AC, and confirmed its expression in the salivary gland via reverse transcription followed by polymerase chain reaction. The putative gene product of Cv rutabaga is a Ca2+/CaM-dependent type I AC and shows highest homology to Rutabaga from Drosophila. Thus, a Ca2+/CaM-dependent AC serves as a link between the InsP(3)/Ca2+ and the cAMP/PKA signalling pathways in the salivary gland of the blowfly and might be important for the amplification and optimization of the secretory response.
Honeybees, Apis mellifera, can differ considerably in their birth weights but the consequences of these weight differences for behaviour are unknown. I investigated how these birth weight differences affected their cognitive skills when the bees reached foraging age. Individual sucrose responsiveness measured by the proboscis extension response is a strong determinant of appetitive olfactory learning performance in honeybees. Most of the observed learning differences between individuals or between genetic bee strains correlate with differences in their sucrose responsiveness. My second aim was therefore to investigate whether the sucrose responsiveness of newly emerged bees could predict the learning behaviour of the bees 3 weeks later. Both birth weight and sucrose responsiveness measured at emergence could predict olfactory learning scores as demonstrated by significant positive correlations. Heavy bees and bees with high sucrose responsiveness later learned better than lighter individuals or bees with lower responsiveness to sucrose at emergence. These results demonstrate for the first time a fundamental relationship between sensory responsiveness and morphology at emergence and later cognitive skills in insects. Because sensory responsiveness is closely linked to division of labour in honeybees, differences in weight and sucrose responsiveness at emergence might be involved in regulating division of labour.
We have analyzed the O-antigen polysaccharide of the previously uncharacterized Escherichia coli strain TD2158 which is a host of bacteriophage HK620. This bacteriophage recognizes and cleaves the polysaccharide with its tailspike protein (TSP). The polysaccharide preparation as well as oligosaccharides obtained from HK620TSP endoglycosidase digests were analyzed with NMR spectroscopy. Additionally, sugar analysis was performed on the O-antigen polysaccharide and MALDI-TOF MS was used in oligosaccharide analysis. The present study revealed a heterogeneous polysaccharide with a hexasaccharide repeating unit of the following structure:
alpha-D-Glcp-(1 -> 6) vertical bar vertical bar 2)-alpha-L-Rhap-(1 -> 6)-alpha-D-Glcp-(1 -> 4)-alpha-D-Galp-(1 -> 3)-alpha-D-GlcpNAc- (1 ->vertical bar beta-D-Glcp/beta-D-GlcpNAc-(1 -> 3)
A repeating unit with a D-GlcNAc substitution of D-Gal has been described earlier as characteristic for serogroup O18A1. Accordingly, we termed repeating units with D-Glc substitution at D-Gal as O18A2. NMR analyses of the polysaccharide confirmed that O18A1- and O18A2-type repeats were present in a 1:1 ratio. However, HK620TSP preferentially bound the D-GlcNAc- substituted O18A1-type repeating units in its high affinity binding pocket with a dissociation constant of 140 mu M and disfavored the O18A2-type having a beta-D-Glcp-(1 -> 3)-linked group. As a result, in hexasaccharide preparations, O18A1 and O18A2 repeats were present in a 9: 1 ratio stressing the clear preference of O18A1- type repeats to be cleaved by HK620TSP.
A direct competitive homogeneous immunoassay for progesterone - the Redox Quenching Immunoassay
(2012)
A direct competitive amperometric immunoassay format for the detection of haptens and proteins was developed. The method is based on the quenching of electroactivity of ferrocenium, which is coupled to the antigen and used as the primary reporter, upon binding to a monoclonal anti-ferrocenium antibody, which is coupled to the detection antibody and used as a secondary reporter. A separation-free progesterone immunoassay with a lower detection limit of 1 ng?mL-1 (3.18 nmol?L-1) in 1?:?2 diluted blood serum was realised by combining two bifunctional conjugates, a ferrocenium-PEG-progesterone tracer and a bioconjugate of one anti-progesterone and one anti-ferrocenium antibody. The immune complex is formed within 30 s upon addition of progesterone, resulting in a total analysis time of 1.5 min.
Sperm proteins of marine sessile invertebrates have been extensively studied to understand the molecular basis of reproductive isolation. Apart from molecules such as bindin of sea urchins or lysin of abalone species, the acrosomal protein M7 lysin of Mytilus edulis has been analyzed. M7 lysin was found to be under positive selection, but mechanisms driving the evolution of this protein are not fully understood. To explore functional aspects, this study investigated the protein expression pattern of M7 and M6 lysin in gametes and somatic tissue of male and female M. edulis. The study employs a previously published monoclonal antibody (G26-AG8) to investigate M6 and M7 lysin protein expression, and explores expression of both genes. It is shown that these proteins and their encoding genes are expressed in gametes and somatic tissue of both sexes. This is in contrast to sea urchin bindin and abalone lysin, in which gene expression is strictly limited to males. Although future studies need to clarify the functional importance of both acrosomal proteins in male and female somatic tissue, new insights into the evolution of sperm proteins in marine sessile invertebrates are possible. This is because proteins with male-specific expression (bindin, lysin) might evolve differently than proteins with expression in both sexes (M6/M7 lysin), and the putative function of both proteins in females opens the possibility that the evolution of M6/M7 lysin is under sexual antagonistic selection, for example, mutations beneficial to the acrosomal function that are less beneficial the function in somatic tissue of females.Mol. Reprod. Dev. 79: 517-524, 2012.
Background: Reconstruction of genome-scale metabolic networks has resulted in models capable of reproducing experimentally observed biomass yield/growth rates and predicting the effect of alterations in metabolism for biotechnological applications. The existing studies rely on modifying the metabolic network of an investigated organism by removing or inserting reactions taken either from evolutionary similar organisms or from databases of biochemical reactions (e.g., KEGG). A potential disadvantage of these knowledge-driven approaches is that the result is biased towards known reactions, as such approaches do not account for the possibility of including novel enzymes, together with the reactions they catalyze.
Results: Here, we explore the alternative of increasing biomass yield in three model organisms, namely Bacillus subtilis, Escherichia coil, and Hordeum vulgare, by applying small, chemically feasible network modifications. We use the predicted and experimentally confirmed growth rates of the wild-type networks as reference values and determine the effect of inserting mass-balanced, thermodynamically feasible reactions on predictions of growth rate by using flux balance analysis.
Conclusions: While many replacements of existing reactions naturally lead to a decrease or complete loss of biomass production ability, in all three investigated organisms we find feasible modifications which facilitate a significant increase in this biological function. We focus on modifications with feasible chemical properties and a significant increase in biomass yield. The results demonstrate that small modifications are sufficient to substantially alter biomass yield in the three organisms. The method can be used to predict the effect of targeted modifications on the yield of any set of metabolites (e.g., ethanol), thus providing a computational framework for synthetic metabolic engineering.
Background: The multidrug transporter P-glycoprotein (PGP) is expressed in the human placenta. In particular the C3435T ABCB1 polymorphism was associated with altered tissue expression of PGP in the human placenta. However, the potential functional impact of this polymorphism on the offspring is unknown so far.
Methods: We analyzed the impact of the ABCB1/C3435T polymorphism on fetal growth in 262 mother/child pairs. Fetal growth was assessed by differential ultrasound examination of the fetal body prior to birth and by measuring birth weight.
Results: The maternal ABCB1/C3435T polymorphism showed no trend for an association with birth weight or any ultrasound parameter describing late gestational fetal body shape. Genotyping the newborns, however, demonstrated that newborns carrying two copies of the T allele had a birth weight of 3176.39 g, whereas CT and CC newborns had a birth weight of 3345.04 g (p = 0.022). Adjusting for gestational age at delivery, child's gender, maternal BM1, maternal age and body weight at delivery confirmed this finding (p = 0.009). Considering gestational day of late ultrasound examination, gestational age at delivery, child's gender, maternal BMI, maternal age and maternal body weight at delivery, the fetal ABCB1/C3435T genotype revealed likewise a significant negative correlation with abdominal diameter and abdominal circumference (R-2 = 0.538, p = 0.010 and R-2 = 0.534, p = 0.005, respectively).
Conclusions: Low birth weight may be a risk factor for cardiovascular diseases in later life. The fetal ABCB1/C3435T gene polymorphism may contribute to this risk. Since PGP controls transport of various biological agents, we suggest that PGP is involved in the transport of biological agents to the fetus that are important for normal fetal growth.
Land use is increasingly recognized as a major driver of biodiversity and ecosystem functioning in many current research projects. In grasslands, land use is often classified by categorical descriptors such as pastures versus meadows or fertilized versus unfertilized sites. However, to account for the quantitative variation of multiple land-use types in heterogeneous landscapes, a quantitative, continuous index of land-use intensity (LUI) is desirable. Here we define such a compound, additive LUI index for managed grasslands including meadows and pastures. The LUI index summarizes the standardized intensity of three components of land use, namely fertilization, mowing, and livestock grazing at each site. We examined the performance of the LUI index to predict selected response variables on up to 150 grassland sites in the Biodiversity Exploratories in three regions in Germany(Alb, Hainich, Schorlheide). We tested the average Ellenberg nitrogen indicator values of the plant community, nitrogen and phosphorus concentration in the aboveground plant biomass, plant-available phosphorus concentration in the top soil, and soil C/N ratio, and the first principle component of these five response variables.
The LUI index significantly predicted the principal component of all five response variables, as well as some of the individual responses. Moreover, vascular plant diversity decreased significantly with LUI in two regions (Alb and Hainich).
Inter-annual changes in management practice were pronounced from 2006 to 2008, particularly due to variation in grazing intensity. This rendered the selection of the appropriate reference year(s) an important decision for analyses of land-use effects, whereas details in the standardization of the index were of minor importance. We also tested several alternative calculations of a LUI index, but all are strongly linearly correlated to the proposed index.
The proposed LUI index reduces the complexity of agricultural practices to a single dimension and may serve as a baseline to test how different groups of organisms and processes respond to land use. In combination with more detailed analyses, this index may help to unravel whether and how land-use intensities, associated disturbance levels or other local or regional influences drive ecological processes.
Disturbances are characteristic for many ecosystems. However, we still lack generalizations concerning their role in shaping communities, particularly when disturbances co-occur. To study such effects, we used a novel modeling approach that is unrestricted by a priori tradeoffs among specific plant traits, except for those generated by allocation principles. Thus, trait combinations were emergent properties associated with biotic and abiotic constraints. Specifically, we asked which traits dominate under specific disturbance regimes, whether single and combined disturbance regimes promote similar trait tradeoffs and how complex disturbance regimes affect species richness and functional diversity. Overall, disturbances' temporal properties governed the outcome of combined disturbances and were a stronger assortative force than spatial disturbance properties: low temporal predictability decreased seed-dispersability and dormancy, but increased competitive ability and disturbance tolerance. Evidence for tradeoffs between different colonization modes and between dormancy and disturbance tolerance were found, while surprisingly, the widely accepted colonization-competition tradeoff was not generated. Diversity was highest at intermediate disturbance intensity, but decreased monotonically with increasing unpredictability. In accordance with our results, future models should avoid restrictive assumptions about tradeoffs to generate robust and more general predictions about the role of disturbances for community dynamics.
The Antarctic terrestrial environment is under increasing pressure from human activities. The Fildes Region is characterized by high biodiversity, but is also a major logistic centre for the northern Antarctic Peninsula. Different interests, from scientific research, nature conservation, protection of geological and historical values, station operations, transport logistics and tourism, regularly overlap in space and time. This has led to increasing conflict among the multiple uses of the region and breaches of the legal requirements for environmental protection that apply in the area. The aim of this study was to assess the impacts of human activities in the Fildes Region by monitoring the distribution of bird and seal breeding sites and recording human activities and their associated environmental impacts. Data from an initial monitoring period 2003-06 were compared with data from 2008-10. We observed similar or increased levels of air, land and ship traffic, but fewer violations of overflight limits near Antarctic Specially Protected Area No. 150 Ardley Island. Open waste dumping and oil contamination are still major environmental impacts. Scientific and outdoor leisure activities undertaken by station personnel are more frequent than tourist activities and are likely to have a commensurate level of environmental impact. Despite the initial success of some existing management measures, it is essential that scientific and environmental values continue to be safeguarded, otherwise environmental impacts will increase and the habitat will be further degraded. We argue that the Fildes Region should be considered for designation as an Antarctic Specially Managed Area, a measure that has proven effective for environmental management of vulnerable areas of the Antarctic.
Today, serious health problems as overweight and obesity are not just constricted to the developed world, but also increase in the developing countries (Prentice 2006, Ramachandram et al. 2002). Focusing on this issue, BMI and percentage of body fat were compared in 2094 schoolchildren from two cross-sectional studies from India and Germany investigated in 2008 and 2009. The German children are in all age groups significantly taller, whereas the Indian children show higher values in BMI (e.g. 12 years: Indian: around 22 kg/m(2); German: around 19 kg/m(2)) and in the percentage of body fat (e.g. 12 years: Indian: around 27 %; German: around 18-20%) in most of the investigated age groups. The Indian children have significantly higher BMI between 10 and 13 (boys) respectively 14 years (girls). Indian children showed significant higher percentage of body fat between 10 and 15 years (boys) and between 8 and 16 years (girls). The difference in overweight between Indian and German children was strongest at 11 (boys) and 12 (girls) years: 70 % of the Indian but 20% of the German children were classified as overweight. In countries such as India that undergo nutritional transition, a rapid increase in obesity and overweight is observed. In contrast to the industrialized countries, the risk of overweight in developing countries is associated with high socioeconomic status. Other reasons of the rapid increase of overweight in the developing countries caused by different environmental or genetic factors are discussed.
The trade-off between predation risk and the need to feed is one of the major constraints animals have to cope with. Virtually all animals have a higher risk of being preyed upon when being active (e.g., searching for food or mating partners), compared with being inactive (e.g., staying at their burrows, nests, etc.). Yet, staying safe leads to a higher risk of starvation and may reduce reproductive success and body growth. Hence selection on behaviour optimizing the search, handling and digestion of food while avoiding the risk of predation is strong and should lead to strategies maximising survival chances and inclusive fitness. These facts call for integrative studies manipulating both, abundance of food and predation risk in a factorial set up, analysing the effects of both factors on behaviour and physiological parameters. We present results of a 2 x 2 factorial experiment, manipulating risk of predation and food abundance in guppies. We found that the two factors have an additive effect on body growth, but that predation risk by a pike cichlid is the main factor affecting the expression of behavioural strategies in guppies. Low food availability and high predation risk led to lower body growth. High predation risk affected swimming depth and risk sensitivity of guppies and might represent adaptive behavioural changes to the environmental context experienced in early life. Our study shows that integrative studies, analysing multiple interdependent and interconnected factors in the wild and in the lab are needed to further understand animal behaviour and development.
Impact of Land-Use intensity and productivity on bryophyte diversity in agricultural grasslands
(2012)
While bryophytes greatly contribute to plant diversity of semi-natural grasslands, little is known about the relationships between land-use intensity, productivity, and bryophyte diversity in these habitats. We recorded vascular plant and bryophyte vegetation in 85 agricultural used grasslands in two regions in northern and central Germany and gathered information on land-use intensity. To assess grassland productivity, we harvested aboveground vascular plant biomass and analyzed nutrient concentrations of N, P, K, Ca and Mg. Further we calculated mean Ellenberg indicator values of vascular plant vegetation. We tested for effects of land-use intensity and productivity on total bryophyte species richness and on the species richness of acrocarpous (small & erect) and pleurocarpous (creeping, including liverworts) growth forms separately. Bryophyte species were found in almost all studied grasslands, but species richness differed considerably between study regions in northern Germany (2.8 species per 16 m(2)) and central Germany (6.4 species per 16 m(2)) due environmental differences as well as land-use history. Increased fertilizer application, coinciding with high mowing frequency, reduced bryophyte species richness significantly. Accordingly, productivity estimates such as plant biomass and nitrogen concentration were strongly negatively related to bryophyte species richness, although productivity decreased only pleurocarpous species. Ellenberg indicator values for nutrients proved to be useful indicators of species richness and productivity. In conclusion, bryophyte composition was strongly dependent on productivity, with smaller bryophytes that were likely negatively affected by greater competition for light. Intensive land-use, however, can also indirectly decrease bryophyte species richness by promoting grassland productivity. Thus, increasing productivity is likely to cause a loss of bryophyte species and a decrease in species diversity.
Gaining understanding of food-web processes often requires a simplified representation of natural diversity. One such simplification can be based on functional traits, as functionally similar species may provide a similar contribution to ecosystem level-processes. However, understanding how similarity in functional traits actually translates into similar contributions to ecosystem-level properties remains a challenge due to the complex ways in which traits can influence species' dynamics. Moreover, in many communities, seasonality alters the abiotic and biotic forcing regime, causing ongoing changes to patterns of species' dominance; groups of species do not stay intact but are rather continuously subjected to changes throughout the year. Using long-term high frequency measurements of phytoplankton in Lake Constance, we investigated the effect of seasonal changes on the relationship between functional similarity and temporal dynamics similarity of 36 morphotypes, and the relative contribution of different functional traits during the different parts of the year. Our results revealed seasonal differences in the overall degree of synchronization of morphotypes' temporal dynamics and how combinations of functional traits influence the relationship between functional trait similarity and temporal dynamics similarity, showing that different forcing regimes change how species cope with their environment based on their functional traits. Moreover, we show that the individual functional traits matter at different periods of the year indicating that species which are dynamically similar at certain parts of the year may not be at others. The differential strength of the overall and individual impact of functional traits on species' temporal dynamics makes the cohesion of a pair of functionally similar species dependent on the different forcing. Hence, simplifying food webs based solely on functional traits may not provide consistent estimates of functional groups over all seasons.
The dimensions of unfolded and intrinsically disordered proteins are highly dependent on their amino acid composition and solution conditions, especially salt and denaturant concentration. However, the quantitative implications of this behavior have remained unclear, largely because the effective theta-state, the central reference point for the underlying polymer collapse transition, has eluded experimental determination. Here, we used single-molecule fluorescence spectroscopy and two-focus correlation spectroscopy to determine the theta points for six different proteins. While the scaling exponents of all proteins converge to 0.62 +/- 0.03 at high denaturant concentrations, as expected for a polymer in good solvent, the scaling regime in water strongly depends on sequence composition. The resulting average scaling exponent of 0.46 +/- 0.05 for the four foldable protein sequences in our study suggests that the aqueous cellular milieu is close to effective theta conditions for unfolded proteins. In contrast, two intrinsically disordered proteins do not reach the T-point under any of our solvent conditions, which may reflect the optimization of their expanded state for the interactions with cellular partners. Sequence analyses based on our results imply that foldable sequences with more compact unfolded states are a more recent result of protein evolution.
An electrochemical detection system specifically designed for multi-parameter real-time monitoring of stem cell culturing/differentiation in a microfluidic system is presented. It is composed of a very compact 24-channel electronic board, compatible with arrays of microelectrodes and coupled to a microfluidic cell culture system. A versatile data acquisition software enables performing amperometry, cyclic voltammetry and impedance spectroscopy in each of the 12 independent chambers over a 100 kHz bandwidth with current resolution down to 5 pA for 100 ms measuring time. The design of the platform, its realization and experimental characterization are reported, with emphasis on the analysis of impact of input capacitance (i.e., microelectrode size) and microfluidic pump operation on current noise. Programmable sequences of successive injections of analytes (ferricyanide and dopamine) and rinsing buffer solution as well as the impedimetric continuous tracking for seven days of the proliferation of a colony of PC12 cells are successfully demonstrated.
Aldehyde oxidases (AOXs) are homodimeric proteins belonging to the xanthine oxidase family of molybdenum-containing enzymes. Each 150-kDa monomer contains a FAD redox cofactor, two spectroscopically distinct [2Fe-2S] clusters, and a molybdenum cofactor located within the protein active site. AOXs are characterized by broad range substrate specificity, oxidizing different aldehydes and aromatic N-heterocycles. Despite increasing recognition of its role in the metabolism of drugs and xenobiotics, the physiological function of the protein is still largely unknown. We have crystallized and solved the crystal structure of mouse liver aldehyde oxidase 3 to 2.9 angstrom. This is the first mammalian AOX whose structure has been solved. The structure provides important insights into the protein active center and further evidence on the catalytic differences characterizing AOX and xanthine oxidoreductase. The mouse liver aldehyde oxidase 3 three-dimensional structure combined with kinetic, mutagenesis data, molecular docking, and molecular dynamics studies make a decisive contribution to understand the molecular basis of its rather broad substrate specificity.
The c-Fosc-Jun complex forms the activator protein 1 transcription factor, a therapeutic target in the treatment of cancer. Various synthetic peptides have been designed to try to selectively disrupt the interaction between c-Fos and c-Jun at its leucine zipper domain. To evaluate the binding affinity between these synthetic peptides and c-Fos, polarizable and nonpolarizable molecular dynamics (MD) simulations were conducted, and the resulting conformations were analyzed using the molecular mechanics generalized Born surface area (MM/GBSA) method to compute free energies of binding. In contrast to empirical and semiempirical approaches, the estimation of free energies of binding using a combination of MD simulations and the MM/GBSA approach takes into account dynamical properties such as conformational changes, as well as solvation effects and hydrophobic and hydrophilic interactions. The predicted binding affinities of the series of c-Jun-based peptides targeting the c-Fos peptide show good correlation with experimental melting temperatures. This provides the basis for the rational design of peptides based on internal, van der Waals, and electrostatic interactions.
In the light of ongoing land use changes, it is important to understand how multitrophic communities perform at different land use intensities. The paradox of enrichment predicts that fertilization leads to destabilization and extinction of predator-prey systems. We tested this prediction for a land use intensity gradient from natural to highly fertilized agricultural ecosystems. We included multiple aboveground and belowground trophic levels and land use-dependent searching efficiencies of insects. To overcome logistic constraints of field experiments, we used a successfully validated simulation model to investigate plant responses to removal of herbivores and their enemies. Consistent with our predictions, instability measured by herbivore-induced plant mortality increased with increasing land use intensity. Simultaneously, the balance between herbivores and natural enemies turned increasingly towards herbivore dominance and natural enemy failure. Under natural conditions, there were more frequently significant effects of belowground herbivores and their natural enemies on plant performance, whereas there were more aboveground effects in agroecosystems. This result was partly due to the "boom-bust'' behavior of the shoot herbivore population. Plant responses to herbivore or natural enemy removal were much more abrupt than the imposed smooth land use intensity gradient. This may be due to the presence of multiple trophic levels aboveground and belowground. Our model suggests that destabilization and extinction are more likely to occur in agroecosystems than in natural communities, but the shape of the relationship is nonlinear under the influence of multiple trophic interactions. Citation: Meyer KM, Vos M, Mooij WM, Hol WHG, Termorshuizen AJ, et al. (2012) Testing the Paradox of Enrichment along a Land Use Gradient in a Multitrophic Aboveground and Belowground Community.
Secretion in blowfly (Calliphora vicina) salivary glands is stimulated by the biogenic amine serotonin (5-hydroxytryptamine, 5-HT), which activates both inositol 1,4,5-trisphosphate (InsP(3))/Ca2+ and cyclic adenosine 3',5'-monophosphate (cAMP) signalling pathways in the secretory cells. In order to characterize the signal-inducing 5-HT receptors, we cloned two cDNAs (Cv5-ht2 alpha, Cv5-ht7) that share high similarity with mammalian 5-HT2 and 5-HT7 receptor genes, respectively. RT-PCR demonstrated that both receptors are expressed in the salivary glands and brain. Stimulation of Cv5-ht2 alpha-transfected mammalian cells with 5-HT elevates cytosolic [Ca2+] in a dose-dependent manner (EC50 = 24 nM). In Cv5-ht7-transfected cells, 5-HT produces a dose-dependent increase in [cAMP](i) (EC50 = 4 nM). We studied the pharmacological profile for both receptors. Substances that appear to act as specific ligands of either Cv5-HT2 alpha or Cv5-HT7 in the heterologous expression system were also tested in intact blowfly salivary gland preparations. We observed that 5-methoxytryptamine (100 nM) activates only the Cv(5)-HT2 alpha receptor, 5-carboxamidotryptamine (300 nM) activates only the Cv5-HT7 receptor, and clozapine (1 mu M) antagonizes the effects of 5-HT via Cv5-HT7 in blowfly salivary glands, providing means for the selective activation of each of the two 5-HT receptor subtypes. This study represents the first comprehensive molecular and pharmacological characterization of two 5-HT receptors in the blowfly and permits the analysis of the physiological role of these receptors, even when co-expressed in cells, and of the modes of interaction between the Ca2+- and cAMP-signalling cascades. Citation: Roser C, Jordan N, Balfanz S, Baumann A, Walz B, et al. (2012) Molecular and Pharmacological Characterization of Serotonin 5-HT2a and 5-HT7 Receptors in the Salivary Glands of the Blowfly Calliphora vicina.
Wood has an excellent mechanical performance, but wider utilization of this renewable resource as an engineering material is limited by unfavorable properties such as low dimensional stability upon moisture changes and a low durability. However, some wood species are known to produce a wood of higher quality by inserting mainly phenolic substances in the already formed cell walls a process so-called heartwood formation. In the present study, we used the heartwood formation in black locust (Robinia pseudoacacia) as a source of bioinspiration and transferred principles of the modification in order to improve spruce wood properties (Picea abies) by a chemical treatment with commercially available flavonoids. We were able to effectively insert hydrophobic flavonoids in the cell wall after a tosylation treatment for activation. The chemical treatment reduced the water uptake of the wood cell walls and increased the dimensional stability of the bulk spruce wood. Further analysis of the chemical interaction of the flavonoid with the structural cell wall components revealed the basic principle of this bioinspired modification. Contrary to established modification treatments, which mainly address the hydroxyl groups of the carbohydrates with hydrophilic substances, the hydrophobic flavonoids are effective by a physical bulking in the cell wall most probably stabilized by pi-pi interactions. A biomimetic transfer of the underlying principle may lead to alternative cell wall modification procedures and improve the performance of wood as an engineering material.
Isolation and characterization of bone marrow-derived progenitor cells from malignant gliomas
(2012)
Background: Malignant gliomas are highly-vascularised tumours. Neoangiogenesis is a crucial factor in the malignant behaviour of tumour and prognosis of patients. Several mechanisms are suspected to lead to neoangiogenesis, one of them is the recruitment of multipotent progenitor cells towards the tumour. Factors such as Vascular endothelial growth factor-A (VEGF-A) were described to recruit bone marrow-derived endothelial progenitor cells (EPCs) to the glioma stroma and vasculature. Little is known about isolating EPCs from normal or malignant tissues. Materials and Methods: In this study, we addressed the topic of characterization of tumour-isolated EPCs and re-defined the clonal relationship between EPCs and hematopoietic stem cells (HSCs) in gliomas. We first checked public gene expression data of glioma for putative marker expression, pointing towards a prevalence of EPCs and HSCs in glioma. Immunohistochemical staining of glioma tissue confirmed the higher expression of these progenitor markers in glioma tissue. EPCs and HSCs were consequently isolated and characterized at the phenotypic and functional levels. We applied a new isolation method, for the first time, to specimen from patients with high grade glioma including seven grade IV glioblastoma, five-grade III astrocytoma, and three grade III oligoastrocytoma. Results: In all samples, we were able to isolate the tumour-derived EPCs, which were positive for characteristic markers: CD31, CD34 and VEGFR2. The EPCs formed capillary networks in vitro and had the ability to take up acetylated low-density lipoprotein. Glioma-derived HSCs were positive for CD34 and CD45, but they were unable to form a capillary network in vitro. These findings on tumour-derived EPCs/HSCs were in concordance with the results, derived from peripheral blood of healthy volunteers. Conclusion: In our study, we established a new method for EPC/HSC isolation from human gliomas, defined the contribution of EPCs and HSCs to the tumour tissue, and highlighted the intense in vivo tumour host interaction.
Silvicultural practices lead to changes in forest composition and structure and may impact species diversity from the overall regional species pool to stand-level species occurrence. We explored to what extent fine-scale occupancy patterns in differently managed forest stands are driven by environment and ecological traits in three regions in Germany using a multi-species hierarchical model. We tested for the possible impact of environmental variables and ecological traits on occupancy dynamics in a joint modelling exercise while taking possible variation in coefficient estimates over years and plots into account. Bird species richness differed across regions and years, and trends in species richness across years were different in the three regions. On the species level, forest management affected occupancy of species in all regions, but only 35% of the total assemblage-level variation in occurrence probability was explained by either forest type and successional stage and <?1% by forest edge. On the assemblage level, bird occurrence decreased with body mass in all regions. Species with smaller breeding ranges had lower occurrence probabilities in one region, while later spring arrival decreased occurrence probabilities in the two other regions. Spatial variation in the effect size of trait covariates such as species phylogeny and breeding strata showed that variation in patch occupancy due to fine-scale differences in forest management is, to some extent, predictable from ecological traits. Our results show that environmental factors and ecological traits jointly predict variation in bird occupancy patterns and their response to forest management. Observations at the fine scale of forest stands, at which conservation efforts can be arranged along with forest management practices in heterogeneous environments, have been shown to provide meaningful insights despite the difficulties involved in monitoring mobile organisms such as birds at the plot level.
We studied peculiarities of the structural reconstruction within holographically recorded gratings on the surface of several different amorphous azobenzene-containing polymers. Under illumination with a light interference pattern, two processes take place in this type of polymer. The first process is the light-induced orientation of azobenzene units perpendicular to the polarization plane of the incident light. The second one is a transfer of macromolecules along the grating vector (i.e. perpendicular to the grating lines). These two processes result in the creation of a volume orientation grating (alternating regions of different direction or degree of molecular orientation) and a surface relief grating (SRG)-i.e. modulation of film thickness. One can assume that both orientation of molecules and their movement might change the local mechanical properties of the material. Therefore, formation of the SRG is expected to result also in modulation of the local stiffness of the polymer film. To reveal and investigate these stiffness changes within the grating, spin-coated polymer films were prepared and the gratings were recorded on them in two different ways: with an orthogonal circular or orthogonal linear polarization of two recording light beams. A combination of atomic force microscopy (AFM) and ultrasonic force microscopy (UFM) techniques was applied for SRG development monitoring. We demonstrate that formation of the phase gratings depends on the chemical structure of polymers being used, polymer film thickness, and recording parameters, with the height of grating structures (depth of modulation) increasing with both the exposure time and the film thickness. UFM images suggest that the slopes of the topographic peaks in the phase gratings exhibit an increased stiffness with respect to the grating depressions.
Determining covalent and charge-transfer contributions to bonding in solution has remained an experimental challenge. Here, the quenching of fluorescence decay channels as expressed in dips in the L-edge X-ray spectra of solvated 3d transition-metal ions and complexes was reported as a probe. With a full set of experimental and theoretical ab initio L-edge X-ray spectra of aqueous Cr3+, including resonant inelastic X-ray scattering, we address covalency and charge transfer for this prototypical transition-metal ion in solution. We dissect local atomic effects from intermolecular interactions and quantify X-ray optical effects. We find no evidence for the asserted ultrafast charge transfer to the solvent and show that the dips are readily explained by X-ray optical effects and local atomic state dependence of the fluorescence yield. Instead, we find, besides ionic interactions, a covalent contribution to the bonding in the aqueous complex of ligand-to-metal charge-transfer character.
This article gives an overview of the current status of knowledge concerning the role of nanoparticles (inorganic and organic) in deep geological host rocks and the potential influence of these nanoparticles on radionuclide migration in far-field systems. The manuscript is not intended to be a full review paper or overview paper concerning nanoparticles, here the intention is to refer to recent publications but to highlight the progress made in the 6th framework project IP FUNMIG (Fundamental processes of radionuclide migration) and the open literature over the past 5 a concerning the process understanding of nanoparticle related issues in the three host rock formations investigated, namely: claystones, crystalline rocks and salt rock overburden. The results show inter alia that the inorganic nanoparticle concentration in deep groundwaters of advection dominated systems rarely exceeds 1 mg L (1) and is expected to be in the ng L (1) range in diffusion controlled systems. For organic nanoparticles DOC concentrations up to tens of milligrams in diffusion-controlled indurated clays with molecular sizes mostly <500 Da have been found. Fulvic acid type organics have been identified in crystalline environments and plastic Clay formations (Boom Clay) with molecular sizes <= 300 kDa. Additional sources of inorganic nanoparticles from the repository near-field (compacted bentonite) were identified and the initial erosion rates were determined. The results indicate under stagnant conditions similar to 38 mg cm (2) a (1) for bi-distilled water, similar to 20 mg cm (2) a (1) for glacial melt water (Grimsel groundwater) and very low rates similar to 0.02 mg cm (2) a (1) for 5 mM CaCl2 contact water. The low critical coagulation concentration (CCC) indicative for purely diffusion controlled coagulation of 1 mM L (1) Ca2+ found in bentonite nanoparticle stability analysis matches the low nanoparticle mobilization from compacted bentonite found in these systems.
Aim The study and prediction of speciesenvironment relationships is currently mainly based on species distribution models. These purely correlative models neglect spatial population dynamics and assume that species distributions are in equilibrium with their environment. This causes biased estimates of species niches and handicaps forecasts of range dynamics under environmental change. Here we aim to develop an approach that statistically estimates process-based models of range dynamics from data on species distributions and permits a more comprehensive quantification of forecast uncertainties.
Innovation We present an approach for the statistical estimation of process-based dynamic range models (DRMs) that integrate Hutchinson's niche concept with spatial population dynamics. In a hierarchical Bayesian framework the environmental response of demographic rates, local population dynamics and dispersal are estimated conditional upon each other while accounting for various sources of uncertainty. The method thus: (1) jointly infers species niches and spatiotemporal population dynamics from occurrence and abundance data, and (2) provides fully probabilistic forecasts of future range dynamics under environmental change. In a simulation study, we investigate the performance of DRMs for a variety of scenarios that differ in both ecological dynamics and the data used for model estimation.
Main conclusions Our results demonstrate the importance of considering dynamic aspects in the collection and analysis of biodiversity data. In combination with informative data, the presented framework has the potential to markedly improve the quantification of ecological niches, the process-based understanding of range dynamics and the forecasting of species responses to environmental change. It thereby strengthens links between biogeography, population biology and theoretical and applied ecology.
The interaction of diverse biomaterials with surfaces is more crucial than ever for biomedical applications to ensure efficiency and reproducibility. Very interesting surface materials are micrometer-thick polyelectrolyte multilayers. Not only their surface but also the bulk can be loaded with biomaterials like proteins or DNA for various purposes. Therefore, we established a method to analyze the lateral and vertical distribution of fluorescently labelled proteins of various size and charge in polyelectrolyte films composed of poly(L-lysine) and hyaluronic acid by confocal laser scanning microscopy. This approach enables us to measure the diffusion coefficients of the proteins via fluorescence recovery after photobleaching as a function of their vertical position in the film and facilitates the understanding of molecular interactions in the film with a high resolution in both space and time. As a result, we confirm that protein loading in the film is driven by electrostatic interactions - uncharged dextran molecules of 10 and 500 kDa do not diffuse into the film. Proteins of different sizes (3-11 nm) can diffuse relatively fast (D = 2-4 mm(2) s(-1)) independent of their net charge, indicating complex interpolymer interactions. This approach is a new powerful experimental tool to design the polyelectrolyte multilayers for bio-applications by finding a relationship between intermolecular interactions and mobility and availability of biomolecules to biological samples (e.g. cells) or detection units (e.g. biosensors).
Coastal regions of the North East Atlantic and Mediterranean Seas have four known species of driftwood talitrids. Records are extremely scanty, often limited to the type locality and dating to 1950. We were able to study three of them, all belonging to the genus Macarorchestia, using fresh and archived samples including type material. Allometric and molecular analyses support: (1) a close relationship among all the three classically defined Macarorchestia species, (2) Macarorchestia was well separated from non-driftwood taxa, and (3) a putative new driftwood talitrid discovered during this study was not closely related to Macarorchestia. Genetic divergence between the new species and Macarorchestia remyi is as high as the average distance among a number of talitrid species included in the study for comparison. A key is provided to identify all three of the presently known species of Macarorchestia, using morphological characters employed in the allometric study.
Restoration is gaining importance in the management of plant invasions. As the success of restoration projects is frequently determined by factors other than ecological ones, we explored the ecological and financial feasibility of active restoration on three different invaded sites in South Africa's Cape Floristic Region. The aim of our study was to identify cost-effective ways of restoring functional native ecosystems following invasion by alien plants. Over three years we evaluated different restoration approaches using field trials and experimental manipulations (i.e. mechanical clearing, burning, different soil restoration techniques and sowing of native species) to reduce elevated soil nutrient levels and to re-establish native fynbos communities. Furthermore we investigated the possibility of introducing native fynbos species that can be used for sustainable harvesting to create an incentive for restoration on private land.
Diversity and evenness of native plant species increased significantly after restoration at all three sites, whereas cover of alien plants decreased significantly, confirming that active restoration was successful. However, sowing of native fynbos species had no significant effect on native cover, species richness, diversity or evenness in the Acacia thicket and Kikuyu field, implying that the ecosystem was sufficiently resilient to allow autogenic recovery following clearing and burning of the invasive species. Soil restoration treatments resulted in an increase of available nitrogen in the Acacia thicket, but had no significant effects in the Eucalyptus plantation. However, despite elevated available soil nitrogen levels, native species germinated irrespective whether sown or unsown (i.e. regeneration from the soil seed bank).
Without active introduction of native species, native grasses, forbs and other shrubs would have dominated, and proteoids and ericoids (the major fynbos growth forms) would have been under-represented.
The financial analysis shows that income from flower harvesting following active restoration consistently outweighs income following passive restoration, but that the associated increase in income does not always justify the higher costs. We conclude that active restoration can be effective and financially feasible when compared to passive restoration, depending on the density of invasion. Active restoration of densely invaded sites may therefore only be justifiable if the target area is in a region of high conservation priority.
We studied the spreading of feature-based attention from attended to ignored motion fields (linear, circular, and combinations). When observers attended one of two superimposed motion fields on one side of the visual midline, sub-threshold priming by an ignored motion field was altered significantly on the opposite side of the midline. This attentional spreading was observed only when attended and ignored motion fields conformed to a complex global flow, not when they shared the same linear motion. These findings corroborate an earlier study (Festman & Braun, 2010), which obtained similar results with a complementary methodology. We conclude that feature-based attention is more complex than hitherto appreciated in that it spreads preferentially in an object-specific manner.
Unique properties of eukaryote-type actin and profilin horizontally transferred to cyanobacteria
(2012)
A eukaryote-type actin and its binding protein profilin encoded on a genomic island in the cyanobacterium Microcystis aeruginosa PCC 7806 co-localize to form a hollow, spherical enclosure occupying a considerable intracellular space as shown by in vivo fluorescence microscopy. Biochemical and biophysical characterization reveals key differences between these proteins and their eukaryotic homologs. Small-angle X-ray scattering shows that the actin assembles into elongated, filamentous polymers which can be visualized microscopically with fluorescent phalloidin. Whereas rabbit actin forms thin cylindrical filaments about 100 mu m in length, cyanobacterial actin polymers resemble a ribbon, arrest polymerization at 510 lam and tend to form irregular multi-strand assemblies. While eukaryotic profilin is a specific actin monomer binding protein, cyanobacterial profilin shows the unprecedented property of decorating actin filaments. Electron micrographs show that cyanobacterial profilin stimulates actin filament bundling and stabilizes their lateral alignment into heteropolymeric sheets from which the observed hollow enclosure may be formed. We hypothesize that adaptation to the confined space of a bacterial cell devoid of binding proteins usually regulating actin polymerization in eukaryotes has driven the co-evolution of cyanobacterial actin and profilin, giving rise to an intracellular entity.
Influence of drying on the secondary structure of intrinsically disordered and globular proteins
(2012)
Circular dichroism (CD) spectroscopy of five Arabidopsis late embryogenesis abundant (LEA) proteins constituting the plant specific families LEA_5 and LEA_6 showed that they are intrinsically disordered in solution and partially fold during drying. Structural predictions were comparable to these results for hydrated LEA_6, but not for LEA_5 proteins. FTIR spectroscopy showed that verbascose, but not sucrose, strongly affected the structure of the dry proteins. The four investigated globular proteins were only mildly affected by drying in the absence, but strongly in the presence of sugars. These data highlight the larger structural flexibility of disordered compared to globular proteins and the impact of sugars on the structure of both disordered and globular proteins during drying.
Background: Triblock copolymers from hydrophilic oligo(ethylene glycol) segment A and oligo(propylene glycol) segment B, providing an ABA structure (OEG-OPG-OEG triblock), are known to be biocompatible and are used as self-solidifying gels in drug depots. A complete removal of these depots would be helpful in cases of undesired side effects of a drug, but this remains a challenge as they liquefy below their transition temperature. Therefore we describe the synthesis of covalently cross-linked hydrogel networks.
Method: Triblock copolymer-based hydrogels were created by irradiating aqueous solutions of the corresponding macro-dimethacrylates with UV light. The degree of swelling, swelling kinetics, mechanical properties and morphology of the networks were investigated.
Results: Depending on precursor concentration, equilibrium degree of swelling of the films ranged between 500% and 880% and was reached in 1 hour. In addition, values for storage and loss moduli of the hydrogel networks were in the 100 Pa to 10 kPa range.
Conclusion: Although OEG-OPG-OEG triblocks are known for their micellization, which could hamper polymer network formation, reactive OEG-OPG-OEG triblock oligomers could be successfully polymerized into hydrogel networks. The degree of swelling of these hydrogels depends on their molecular weight and on the oligomer concentration used for hydrogel preparation. In combination with the temperature sensitivity of the ABA triblock copolymers, it is assumed that such hydrogels might be beneficial for future medical applications -e.g., removable drug release systems.
Preferential accumulation of mutant proteins in the nucleus has been suggested to be the molecular culprit that confers cellular toxicity in the neurodegenerative disorders caused by polyglutamine (polyQ) expansion. Here, we use dynamic imaging approaches, orthogonal cross-seeding, and composition analysis to examine the dynamics and structure of nuclear and cytoplasmic inclusions of atrophin-1, implicated in dentatorubropallidoluysian atrophy, a polyQ-based disease with complex clinical features. Our results reveal a large heterogeneity in the dynamics of the nuclear inclusions compared with the compact and immobile cytoplasmic aggregates. At least two types of inclusions of expanded atrophin-1 with different mobility of the molecular species and ability to exchange with the surrounding monomer pool coexist in the nucleus. Intriguingly, the enrichment of nuclear inclusions with slow dynamics parallels changes in the aggregate core architecture that are dominated by the polyQ stretch. We propose that the observed complexity in the dynamics of the nuclear inclusions provides a molecular explanation for the enhanced cellular toxicity of the nuclear aggregates in polyQ-based neurodegeneration.
Background: Different immunotherapy approaches for the treatment of cancer and autoimmune diseases are being developed and tested in clinical studies worldwide. Their resulting complex experimental data should be properly evaluated, therefore reliable normal healthy control baseline values are indispensable.
Methodology/Principal Findings: To assess intra- and inter-individual variability of various biomarkers, peripheral blood of 16 age and gender equilibrated healthy volunteers was sampled on 3 different days within a period of one month. Complex "crossomics'' analyses of plasma metabolite profiles, antibody concentrations and lymphocyte subset counts as well as whole genome expression profiling in CD4(+)T and NK cells were performed. Some of the observed age, gender and BMI dependences are in agreement with the existing knowledge, like negative correlation between sex hormone levels and age or BMI related increase in lipids and soluble sugars. Thus we can assume that the distribution of all 39.743 analysed markers is well representing the normal Caucasoid population. All lymphocyte subsets, 20% of metabolites and less than 10% of genes, were identified as highly variable in our dataset.
Conclusions/Significance: Our study shows that the intra- individual variability was at least two-fold lower compared to the inter-individual one at all investigated levels, showing the importance of personalised medicine approach from yet another perspective.
Rapid progress of experimental biology has provided a huge flow of quantitative data, which can be analyzed and understood only through the application of advanced techniques recently developed in theoretical sciences. On the other hand, synthetic biology enabled us to engineer biological models with reduced complexity. In this review we discuss that a multidisciplinary approach between this sciences can lead to deeper understanding of the underlying mechanisms behind complex processes in biology. Following the mini symposia "Noise and oscillations in biological systems" on Physcon 2011 we have collected different research examples from theoretical modeling, experimental and synthetic biology.
We report the mediated electrocatalytic voltammetry of the molybdoenzyme xanthine dehydrogenase (XDH) from Rhodobacter capsulatus at a thiol-modified Au electrode. The 2-electron acceptor N-methylphenazinium methanesulfonate (phenazine methosulfate, PMS) is an effective artificial electron transfer partner for XDH instead of its native electron acceptor NAD(+). XDH catalyzes the oxidative hydroxylation of hypoxanthine to xanthine and xanthine to uric acid. Cyclic voltammetry was used to generate the active (oxidized) form of the mediator. Simulation of the catalytic voltammetry across a broad range of substrate and PMS concentrations at different sweep rates was achieved with the program DigiSim to yield a set of consistent rate and equilibrium constants that describe the catalytic system. This provides the first example of the mediated electrochemistry of a xanthine dehydrogenase (or oxidase) that is uncomplicated by interference from product oxidation. A remarkable two-step, sequential oxidation of hypoxanthine to uric acid via xanthine by XDH is observed.
Effects of food quality and quantity on consumers are neither independent nor interchangeable. Although consumer growth and reproduction show strong variation in relation to both food quality and quantity, the effects of food quality or food quantity have usually been studied in isolation. In two experiments, we studied the growth and reproduction in three filter-feeding freshwater zooplankton species, i.e. Daphnia galeata x hyalina, D. pulicaria and D. magna, on their algal food (Scenedesmus obliquus), varying in carbon to phosphorus (C:P) ratios and quantities (concentrations). In the first experiment, we found a strong positive effect of the phosphorus content of food on growth of Daphnia, both in their early and late juvenile development. Variation in the relationship between the P-content of animals and their growth rate reflected interspecific differences in nutrient requirements. Although growth rates typically decreased as development neared maturation, this did not affect these species-specific couplings between growth rate and Daphnia P-content. In the second experiment, we examined the effects of food quality on Daphnia growth at different levels of food quantity. With the same decrease in P-content of food, species with higher estimated P-content at zero growth showed a larger increase in threshold food concentrations (i.e. food concentration sufficient to meet metabolic requirements but not growth). These results suggest that physiological processes such as maintenance and growth may in combination explain effects of food quality and quantity on consumers. Our study shows that differences in response to variation in food quality and quantity exist between species. As a consequence, species-specific effects of food quality on consumer growth will also determine how species deal with varying food levels, which has implications for resource-consumer interactions.
Clusters of codons pairing to low-abundance tRNAs synchronize the translation with co-translational folding of single domains in multidomain proteins. Although proven with some examples, the impact of the ribosomal speed on the folding and solubility on a global, cell-wide level remains elusive. Here we show that upregulation of three low-abundance tRNAs in Escherichia coil increased the aggregation propensity of several cellular proteins as a result of an accelerated elongation rate. Intriguingly, alterations in the concentration of the natural tRNA pool compromised the solubility of various chaperones consequently rendering the solubility of some chaperone-dependent proteins.
Motivation: Metabolic engineering aims at modulating the capabilities of metabolic networks by changing the activity of biochemical reactions. The existing constraint-based approaches for metabolic engineering have proven useful, but are limited only to reactions catalogued in various pathway databases.
Results: We consider the alternative of designing synthetic strategies which can be used not only to characterize the maximum theoretically possible product yield but also to engineer networks with optimal conversion capability by using a suitable biochemically feasible reaction called 'stoichiometric capacitance'. In addition, we provide a theoretical solution for decomposing a given stoichiometric capacitance over a set of known enzymatic reactions. We determine the stoichiometric capacitance for genome-scale metabolic networks of 10 organisms from different kingdoms of life and examine its implications for the alterations in flux variability patterns. Our empirical findings suggest that the theoretical capacity of metabolic networks comes at a cost of dramatic system's changes.
Very few principles have been unraveled that explain the relationship between soil properties and soil biota across large spatial scales and different land-use types. Here, we seek these general relationships using data from 52 differently managed grassland and forest soils in three study regions spanning a latitudinal gradient in Germany. We hypothesize that, after extraction of variation that is explained by location and land-use type, soil properties still explain significant proportions of variation in the abundance and diversity of soil biota. If the relationships between predictors and soil organisms were analyzed individually for each predictor group, soil properties explained the highest amount of variation in soil biota abundance and diversity, followed by land-use type and sampling location. After extraction of variation that originated from location or land-use, abiotic soil properties explained significant amounts of variation in fungal, meso-and macrofauna, but not in yeast or bacterial biomass or diversity. Nitrate or nitrogen concentration and fungal biomass were positively related, but nitrate concentration was negatively related to the abundances of Collembola and mites and to the myriapod species richness across a range of forest and grassland soils. The species richness of earthworms was positively correlated with clay content of soils independent of sample location and land-use type. Our study indicates that after accounting for heterogeneity resulting from large scale differences among sampling locations and land-use types, soil properties still explain significant proportions of variation in fungal and soil fauna abundance or diversity. However, soil biota was also related to processes that act at larger spatial scales and bacteria or soil yeasts only showed weak relationships to soil properties. We therefore argue that more general relationships between soil properties and soil biota can only be derived from future studies that consider larger spatial scales and different land-use types.
To identify copper homeostasis genes in Rhodobacter capsulatus, we performed random transposon Tn5 mutagenesis. Screening of more than 10,000 Tn5 mutants identified tellurite resistance gene trgB as a so far unrecognized major copper tolerance determinant. The trgB gene is flanked by tellurite resistance gene trgA and cysteine synthase gene cysK2. While growth of trgA mutants was only moderately restricted by tellurite, trgB and cysK2 mutants were severely affected by tellurite, which implies that viability under tellurite stress requires increased cysteine levels. Mutational analyses revealed that trgB was the only gene in this chromosomal region conferring cross-tolerance towards copper. Expression of the monocistronic trgB gene required promoter elements overlapping the trgA coding region as shown by nested deletions. Neither copper nor tellurite affected trgB transcription as demonstrated by reverse transcriptase PCR and trgB-lacZ fusions. Addition of tellurite or copper gave rise to increased cellular tellurium and copper concentrations, respectively, as determined by inductively coupled plasma-optical emission spectroscopy. By contrast, cellular iron concentrations remained fairly constant irrespective of tellurite or copper addition. This is the first study demonstrating a direct link between copper and tellurite response in bacteria.
Assessing polyglutamine conformation in the nucleating event by molecular dynamics simulations
(2012)
Polyglutamine (polyQ) diseases comprise a group of dominantly inherited pathology caused by an expansion of an unstable polyQ stretch which is presumed to form beta-sheets. Similar to other amyloid pathologies, polyQ amyloidogenesis occurs via a nucleated polymerization mechanism, and proceeds through energetically unfavorable nucleus whose existence and structure are difficult to detect. Here, we use atomistic molecular dynamics simulations in explicit solvent to assess the conformation of the polyQ stretch in the nucleus that initiates polyQ fibrillization. Comparison of the kinetic stability of various structures of polyQ peptide with a Q-length in the pathological range (Q(40)) revealed that steric zipper or nanotube-like structures (beta-nanotube or beta-pseudohelix) are not kinetically stable enough to serve as a template to initiate polyQ fibrillization as opposed to beta-hairpin-based (beta-sheet and beta-sheetstack) or alpha-helical conformations. The selection of different structures of the polyQ stretch in the aggregation-initiating event may provide an alternative explanation for polyQ aggregate polymorphism.