Converging forest community composition along an edaphic gradient threatens landscape‐level diversity

Aim Plant communities across the temperate zone are changing in response to successional processes and human‐induced disturbances. Here, we assess how upland forest under‐ and overstorey community composition has changed along an edaphic gradient. Location Northern Wisconsin, USA. Methods Forest sites initially sampled in the 1950s were resampled for overstorey composition and diversity, basal area, and understorey composition and diversity. We used clustering methods to identify groups of stands based on overstorey composition, and we used similarity indices, ordination and diversity indices to evaluate changes in species abundance and overall community structure. Results Sites clustered into four overstorey groups along the edaphic gradient: ‘hemlock’ sites dominated by hemlock in 1950, ‘mesic’ sites dominated by northern hardwoods, ‘dry’ sites with a significant pine inclusion in the canopy and diverse ‘dry‐mesic’ sites in the middle. Collectively, forests gained maple, ash and cherry while losing pines, birches and red oaks. The hemlock forest sites gained hardwoods, while the dry‐mesic sites shifted towards a more mesic hardwood composition. Only the driest sites have remained relatively stable in species composition. Main conclusions These trends reflect both ‘mesification’ and homogenization among northern forests. Highly diverse mid‐gradient and mesic hemlock‐dominated stands are transitioning to maple dominance. Fire suppression may be favouring invasions of more mesic plants into historically drier sites, while high deer abundance likely limits hemlock regeneration. If current trends continue, maples will dominate the majority of northern forests, with significant losses of local native species richness and substantial shifts in understorey composition.     

Scale‐dependent processes of community assemblyin an African rift lake

1. Ecologists continue to debate whether the assembly of communities of species is more strongly influenced by dispersal limitations or niche‐based factors. Analytical approaches that account for both mechanisms can help to resolve controls of community assembly. 2. We compared littoral snail assemblages in Lake Tanganyika at three different spatial scales (5–25 m, 0.5–10 km and 0.5–27 km) to test whether spatial distance or environmental differences are better predictors of community similarity. 3. At the finest scale (5–25 m), snail assemblages shifted strongly with depth but not across similar lateral distances, indicating a stronger response to environmental gradients than dispersal opportunities. 4. At the two larger scales (0.5–27 km), both environmental similarity and shoreline distance between sites predicted assemblage similarity across sites. Additionally, canonical correspondence analysis revealed that snail abundances were significantly correlated with algal carbon‐to‐nitrogen ratio and wave energy. 5. Our results indicate that the factors governing assemblage structure are scale dependent; niche‐based mechanisms act across all spatial scales, whereas community similarity declines with distance only at larger spatial separations.     

Mitotic spindle association of TACC3 requires Aurora‐A‐dependent stabilization of a cryptic α‐helix

Aurora‐A regulates the recruitment of TACC3 to the mitotic spindle through a phospho‐dependent interaction with clathrin heavy chain (CHC). Here, we describe the structural basis of these interactions, mediated by three motifs in a disordered region of TACC3. A hydrophobic docking motif binds to a previously uncharacterized pocket on Aurora‐A that is blocked in most kinases. Abrogation of the docking motif causes a delay in late mitosis, consistent with the cellular distribution of Aurora‐A complexes. Phosphorylation of Ser558 engages a conformational switch in a second motif from a disordered state, needed to bind the kinase active site, into a helical conformation. The helix extends into a third, adjacent motif that is recognized by a helical‐repeat region of CHC, not a recognized phospho‐reader domain. This potentially widespread mechanism of phospho‐recognition provides greater flexibility to tune the molecular details of the interaction than canonical recognition motifs that are dominated by phosphate binding.     

Mapping protein interactions of sodium channel NaV1.7 using epitope‐tagged gene‐targeted mice

The voltage‐gated sodium channel Na_V1.7 plays a critical role in pain pathways. We generated an epitope‐tagged Na_V1.7 mouse that showed normal pain behaviours to identify channel‐interacting proteins. Analysis of Na_V1.7 complexes affinity‐purified under native conditions by mass spectrometry revealed 267 proteins associated with Nav1.7 in vivo. The sodium channel β3 (Scn3b), rather than the β1 subunit, complexes with Nav1.7, and we demonstrate an interaction between collapsing‐response mediator protein (Crmp2) and Nav1.7, through which the analgesic drug lacosamide regulates Nav1.7 current density. Novel Na_V1.7 protein interactors including membrane‐trafficking protein synaptotagmin‐2 (Syt2), L‐type amino acid transporter 1 (Lat1) and transmembrane P24‐trafficking protein 10 (Tmed10) together with Scn3b and Crmp2 were validated by co‐immunoprecipitation (Co‐IP) from sensory neuron extract. Nav1.7, known to regulate opioid receptor efficacy, interacts with the G protein‐regulated inducer of neurite outgrowth (Gprin1), an opioid receptor‐binding protein, demonstrating a physical and functional link between Nav1.7 and opioid signalling. Further information on physiological interactions provided with this normal epitope‐tagged mouse should provide useful insights into the many functions now associated with the Na_V1.7 channel.     

Scale‐dependent genetic structure of the Idaho giant salamander (Dicamptodon aterrimus) in stream networks

The network architecture of streams and rivers constrains evolutionary, demographic and ecological processes of freshwater organisms. This consistent architecture also makes stream networks useful for testing general models of population genetic structure and the scaling of gene flow. We examined genetic structure and gene flow in the facultatively paedomorphic Idaho giant salamander, Dicamptodon aterrimus, in stream networks of Idaho and Montana, USA. We used microsatellite data to test population structure models by (i) examining hierarchical partitioning of genetic variation in stream networks; and (ii) testing for genetic isolation by distance along stream corridors vs. overland pathways. Replicated sampling of streams within catchments within three river basins revealed that hierarchical scale had strong effects on genetic structure and gene flow. amova identified significant structure at all hierarchical scales (among streams, among catchments, among basins), but divergence among catchments had the greatest structural influence. Isolation by distance was detected within catchments, and in‐stream distance was a strong predictor of genetic divergence. Patterns of genetic divergence suggest that differentiation among streams within catchments was driven by limited migration, consistent with a stream hierarchy model of population structure. However, there was no evidence of migration among catchments within basins, or among basins, indicating that gene flow only counters the effects of genetic drift at smaller scales (within rather than among catchments). These results show the strong influence of stream networks on population structure and genetic divergence of a salamander, with contrasting effects at different hierarchical scales.     

PCR–DGGE‐based methodologies to assess diversity and dynamics of Aeromonas communities

Aims: Aeromonas is ubiquitous in aquatic environments and may cause infectious diseases in fish and humans. However, reliable and specific methods to evaluate the diversity and dynamics of Aeromonas populations are currently unavailable. This study aimed to develop PCR–DGGE methodologies for culture‐independent analysis of Aeromonas populations in water systems. Methods and Results: Three primer sets were designed to amplify selected sections of genes gyrB, rpoD and sodB from Aeromonas. Their specificity was confirmed by in silico analysis and by PCR on DNA from pure cultures. Estuarine water samples were analyzed by PCR–DGGE using those primers. DGGE patterns clearly clustered according to seasonal factors, and Aeromonas communities were surprisingly stable along a salinity gradient. Sequences of cloned amplicons affiliated to sequences belonging to seven Aeromonas species previously isolated from the same environment. Conclusions: The three systems used showed to be useful to describe the diversity of Aeromonas communities. However, the combined use of more than one primer set is advisable. Significance and Impact of the Study: The methods presented here can be applied to understand the natural pool of Aeromonas and also to monitor and control these bacteria in aquatic reservoirs.     

Hot‐Substrate Deposition of Hole‐ and Electron‐Transport Layers for Enhanced Performance in Perovskite Solar Cells

Charge transport layers play an important role in determining the power conversion efficiencies (PCEs) of perovskite solar cells (PSCs). However, it has proven challenging to produce thin and compact charge transport layers via solution processing techniques. Herein, a hot substrate deposition method capable of improving the morphology of high‐coverage hole‐transport layers (HTLs) and electron‐transport layers (ETLs) is reported. PSC devices using HTLs deposited on a hot substrate show improvement in the open‐circuit voltage (V_oc) from 1.041 to 1.070 V and the PCE from 17.00% to 18.01%. The overall device performance is then further enhanced with the hot substrate deposition of ETLs as the V_oc and PCE reach 1.105 V and 19.16%, respectively. The improved performance can be explained by the decreased current leakage and series resistance, which are present in PSCs with rough and discontinuous HTLs and ETLs.     

The maintenance of sex: host–parasite coevolution with density‐dependent virulence

Why don’t asexual females replace sexual females in most natural populations of eukaryotes? One promising explanation is that parasites could counter the reproductive advantages of asexual reproduction by exerting frequency‐dependent selection against common clones (the Red Queen hypothesis). One apparent limitation of the Red Queen theory, however, is that parasites would seem to be required by theory to be highly virulent. In the present study, I present a population‐dynamic view of competition between sexual females and asexual females that interact with co‐evolving parasites. The results show that asexual populations have higher carrying capacities, and more unstable population dynamics, than sexual populations. The results also suggest that the spread of a clone into a sexual population could increase the effective parasite virulence as population density increases. This combination of parasite‐mediated frequency‐dependent selection, and density‐dependent virulence, could lead to the coexistence of sexual and asexual reproductive strategies and the long‐term persistence of sex.     

Substrate‐driven conformational changes in ClC‐ec1 observed by fluorine NMR

The CLC ‘Cl⁻ channel'' family consists of both Cl⁻/H⁺ antiporters and Cl⁻ channels. Although CLC channels can undergo large, conformational changes involving cooperativity between the two protein subunits, it has been hypothesized that conformational changes in the antiporters may be limited to small movements localized near the Cl⁻ permeation pathway. However, to date few studies have directly addressed this issue, and therefore little is known about the molecular movements that underlie CLC-mediated antiport. The crystal structure of the Escherichia coli antiporter ClC-ec1 provides an invaluable molecular framework, but this static picture alone cannot depict the protein movements that must occur during ion transport. In this study we use fluorine nuclear magnetic resonance (NMR) to monitor substrate-induced conformational changes in ClC-ec1. Using mutational analysis, we show that substrate-dependent ¹⁹F spectral changes reflect functionally relevant protein movement occurring at the ClC-ec1 dimer interface. Our results show that conformational change in CLC antiporters is not restricted to the Cl⁻ permeation pathway and show the usefulness of ¹⁹F NMR for studying conformational changes in membrane proteins of known structure.     

Principles of membrane protein interactions with annular lipids deduced from aquaporin‐0 2D crystals

We have previously described the interactions of aquaporin‐0 (AQP0) with dimyristoyl phosphatidylcholine (DMPC) lipids. We have now determined the 2.5 Å structure of AQP0 in two‐dimensional (2D) crystals formed with Escherichia coli polar lipids (EPLs), which differ from DMPC both in headgroups and acyl chains. Comparison of the two structures shows that AQP0 does not adapt to the different length of the acyl chains in EPLs and that the distance between the phosphodiester groups in the two leaflets of the DMPC and EPL bilayers is almost identical. The EPL headgroups interact differently with AQP0 than do those of DMPC, but the acyl chains in the EPL and DMPC bilayers occupy similar positions. The interactions of annular lipids with membrane proteins seem to be driven by the propensity of the acyl chains to fill gaps in the protein surface. Interactions of the lipid headgroups may be responsible for the specific interactions found in tightly bound lipids but seem to have a negligible effect on interactions of generic annular lipids with membrane proteins.     

Adapting subject specific motor imagery EEG patterns in space–time–frequency for a brain computer interface

In this paper we propose a new technique that adaptively extracts subject specific motor imagery related EEG patterns in the space–time–frequency plane for single trial classification. The proposed approach requires no prior knowledge of reactive frequency bands, their temporal behavior or cortical locations. For a given electrode array, it finds all these parameters by constructing electrode adaptive time–frequency segmentations that are optimized for discrimination. This is accomplished first by segmenting the EEG along the time axis with Local Cosine Packets. Next the most discriminant frequency subbands are selected in each time segment with a frequency axis clustering algorithm to achieve time and frequency band adaptation individually. Finally the subject adapted features are sorted according to their discrimination power to reduce dimensionality and the top subset is used for final classification. We provide experimental results for 5 subjects of the BCI competition 2005 dataset IVa to show the superior performance of the proposed method. In particular, we demonstrate that by using a linear support vector machine as a classifier, the classification accuracy of the proposed algorithm varied between 90.5% and 99.7% and the average classification accuracy was 96%.     

Adenosine‐A3 receptors in neutrophil microdomains promote the formation of bacteria‐tethering cytonemes

The A₃‐adenosine receptor (A₃AR) has recently emerged as a key regulator of neutrophil behaviour. Using a fluorescent A₃AR ligand, we show that A₃ARs aggregate in highly polarized immunomodulatory microdomains on human neutrophil membranes. In addition to regulating chemotaxis, A₃ARs promote the formation of filipodia‐like projections (cytonemes) that can extend up to 100 μm to tether and ‘reel in’ pathogens. Exposure to bacteria or an A₃AR agonist stimulates the formation of these projections and bacterial phagocytosis, whereas an A₃AR‐selective antagonist inhibits cytoneme formation. Our results shed new light on the behaviour of neutrophils and identify the A₃AR as a potential target for modulating their function.     

Structural evidence for Nap1‐dependent H2A–H2B deposition and nucleosome assembly

Nap1 is a histone chaperone involved in the nuclear import of H2A–H2B and nucleosome assembly. Here, we report the crystal structure of Nap1 bound to H2A–H2B together with in vitro and in vivo functional studies that elucidate the principles underlying Nap1‐mediated H2A–H2B chaperoning and nucleosome assembly. A Nap1 dimer provides an acidic binding surface and asymmetrically engages a single H2A–H2B heterodimer. Oligomerization of the Nap1–H2A–H2B complex results in burial of surfaces required for deposition of H2A–H2B into nucleosomes. Chromatin immunoprecipitation‐exonuclease (ChIP‐exo) analysis shows that Nap1 is required for H2A–H2B deposition across the genome. Mutants that interfere with Nap1 oligomerization exhibit severe nucleosome assembly defects showing that oligomerization is essential for the chaperone function. These findings establish the molecular basis for Nap1‐mediated H2A–H2B deposition and nucleosome assembly.     

LuTHy: a double‐readout bioluminescence‐based two‐hybrid technology for quantitative mapping of protein–protein interactions in mammalian cells

Information on protein–protein interactions (PPIs) is of critical importance for studying complex biological systems and developing therapeutic strategies. Here, we present a double‐readout bioluminescence‐based two‐hybrid technology, termed LuTHy, which provides two quantitative scores in one experimental procedure when testing binary interactions. PPIs are first monitored in cells by quantification of bioluminescence resonance energy transfer (BRET) and, following cell lysis, are again quantitatively assessed by luminescence‐based co‐precipitation (LuC). The double‐readout procedure detects interactions with higher sensitivity than traditional single‐readout methods and is broadly applicable, for example, for detecting the effects of small molecules or disease‐causing mutations on PPIs. Applying LuTHy in a focused screen, we identified 42 interactions for the presynaptic chaperone CSPα, causative to adult‐onset neuronal ceroid lipofuscinosis (ANCL), a progressive neurodegenerative disease. Nearly 50% of PPIs were found to be affected when studying the effect of the disease‐causing missense mutations L115R and ?L116 in CSPα with LuTHy. Our study presents a robust, sensitive research tool with high utility for investigating the molecular mechanisms by which disease‐associated mutations impair protein activity in biological systems.