Can size distributions of European lake fish communities be predicted by trophic positions of their fish species?

An organism's body size plays an important role in ecological interactions such as predator-prey relationships. As predators are typically larger than their prey, this often leads to a strong positive relationship between body size and trophic position in aquatic ecosystems. The distribution of body sizes in a community can thus be an indicator of the strengths of predator-prey interactions. The aim of this study was to gain more insight into the relationship between fish body size distribution and trophic position in a wide range of European lakes. We used quantile regression to examine the relationship between fish species' trophic position and their log-transformed maximum body mass for 48 fish species found in 235 European lakes. Subsequently, we examined whether the slopes of the continuous community size distributions, estimated by maximum likelihood, were predicted by trophic position, predator-prey mass ratio (PPMR), or abundance (number per unit effort) of fish communities in these lakes. We found a positive linear relationship between species' maximum body mass and average trophic position in fishes only for the 75% quantile, contrasting our expectation that species' trophic position systematically increases with maximum body mass for fish species in European lakes. Consequently, the size spectrum slope was not related to the average community trophic position, but there were negative effects of community PPMR and total fish abundance on the size spectrum slope. We conclude that predator-prey interactions likely do not contribute strongly to shaping community size distributions in these lakes.

Selective activation of Gαob by an adenosine A1 receptor agonist elicits analgesia without cardiorespiratory depression.

The development of therapeutic agonists for G protein-coupled receptors (GPCRs) is hampered by the propensity of GPCRs to couple to multiple intracellular signalling pathways. This promiscuous coupling leads to numerous downstream cellular effects, some of which are therapeutically undesirable. This is especially the case for adenosine A1 receptors (A1Rs) whose clinical potential is undermined by the sedation and cardiorespiratory depression caused by conventional agonists. We have discovered that the A1R-selective agonist, benzyloxy-cyclopentyladenosine (BnOCPA), is a potent and powerful analgesic but does not cause sedation, bradycardia, hypotension or respiratory depression. This unprecedented discrimination between native A1Rs arises from BnOCPA's unique and exquisitely selective activation of Gob among the six Gαi/o subtypes, and in the absence of ß-arrestin recruitment. BnOCPA thus demonstrates a highly-specific Gα-selective activation of the native A1R, sheds new light on GPCR signalling, and reveals new possibilities for the development of novel therapeutics based on the far-reaching concept of selective Gα agonism.

Rapid adaptation through genomic and epigenomic responses following translocations in an endangered salmonid.

Identifying the molecular mechanisms facilitating adaptation to new environments is a key question in evolutionary biology, especially in the face of current rapid and human-induced changes. Translocations have become an important tool for species conservation, but the attendant small population sizes and new ecological pressures might affect phenotypic and genotypic variation and trajectories dramatically and in unknown ways. In Scotland, the European whitefish (Coregonus lavaretus) is native to only two lakes and vulnerable to extirpation. Six new refuge populations were established over the last 30 years as a conservation measure. In this study, we examined whether there is a predictable ecological and evolutionary response of these fishes to translocation. We found eco-morphological differences, as functional traits relating to body shape differed between source and refuge populations. Dual isotopic analyses suggested some ecological release, with the diets in refuge populations being more diverse than in source populations. Analyses of up to 9117 genome-mapped SNPs showed that refuge populations had reduced genetic diversity and elevated inbreeding and relatedness relative to source populations, though genomic differentiation was low (F ST = 0.002-0.030). We identified 14 genomic SNPs that showed shared signals of a selective response to translocations, including some located near or within genes involved in the immune system, nervous system and hepatic functions. Analysis of up to 120,897 epigenomic loci identified a component of consistent differential methylation between source and refuge populations. We found that epigenomic variation and genomic variation were associated with morphological variation, but we were not able to infer an effect of population age because the patterns were also linked with the methodology of the translocations. These results show that conservation-driven translocations affect evolutionary potential by impacting eco-morphological, genomic and epigenomic components of diversity, shedding light on acclimation and adaptation process in these contexts.

Complex and divergent histories gave rise to genome-wide divergence patterns amongst European whitefish (Coregonus lavaretus).

Pleistocene glaciations dramatically affected species distribution in regions that were impacted by ice cover and subsequent postglacial range expansion impacted contemporary biodiversity in complex ways. The European whitefish, Coregonus lavaretus, is a widely distributed salmonid fish species on mainland Europe, but in Britain it has only seven native populations, all of which are found on the western extremes of the island. The origins and colonization routes of the species into Britain are unknown but likely contributed to contemporary genetic patterns and regional uniqueness. Here, we used up to 25,751 genome-wide polymorphic loci to reconstruct the history and to discern the demographic and evolutionary forces underpinning divergence between British populations. Overall, we found lower genetic diversity in Scottish populations but high differentiation (FST  = 0.433-0.712) from the English/Welsh and other European populations. Differentiation was elevated genome-wide rather than in particular genomic regions. Demographic modelling supported a postglacial colonization into western Scotland from northern refugia and a separate colonization route for the English/Welsh populations from southern refugia, with these two groups having been separated for more than ca. 50 Ky. We found cyto-nuclear discordance at a European scale, with the Scottish populations clustering closely with Baltic population in the mtDNA analysis but not in the nuclear data, and with the Norwegian and Alpine populations displaying the same mtDNA haplotype but being distantly related in the nuclear tree. These findings suggest that neutral processes, primarily drift and regionally distinct pre-glacial evolutionary histories, are important drivers of genomic divergence in British populations of European whitefish. This sheds new light on the establishment of the native British freshwater fauna after the last glacial maximum.

Structure-based identification of dual ligands at the A2AR and PDE10A with anti-proliferative effects in lung cancer cell-lines.

Enhanced/prolonged cAMP signalling has been suggested as a suppressor of cancer proliferation. Interestingly, two key modulators that elevate cAMP, the A2A receptor (A2AR) and phosphodiesterase 10A (PDE10A), are differentially co-expressed in various types of non-small lung cancer (NSCLC) cell-lines. Thus, finding dual-target compounds, which are simultaneously agonists at the A2AR whilst also inhibiting PDE10A, could be a novel anti-proliferative approach. Using ligand- and structure-based modelling combined with MD simulations (which identified Val84 displacement as a novel conformational descriptor of A2AR activation), a series of known PDE10A inhibitors were shown to dock to the orthosteric site of the A2AR. Subsequent in-vitro analysis confirmed that these compounds bind to the A2AR and exhibit dual-activity at both the A2AR and PDE10A. Furthermore, many of the compounds exhibited promising anti-proliferative effects upon NSCLC cell-lines, which directly correlated with the expression of both PDE10A and the A2AR. Thus, we propose a structure-based methodology, which has been validated in in-vitro binding and functional assays, and demonstrated a promising therapeutic value.

The Role of ICL1 and H8 in Class B1 GPCRs; Implications for Receptor Activation.

The first intracellular loop (ICL1) of G protein-coupled receptors (GPCRs) has received little attention, although there is evidence that, with the 8th helix (H8), it is involved in early conformational changes following receptor activation as well as contacting the G protein ß subunit. In class B1 GPCRs, the distal part of ICL1 contains a conserved R12.48KLRCxR2.46b motif that extends into the base of the second transmembrane helix; this is weakly conserved as a [R/H]12.48KL[R/H] motif in class A GPCRs. In the current study, the role of ICL1 and H8 in signaling through cAMP, iCa2+ and ERK1/2 has been examined in two class B1 GPCRs, using mutagenesis and molecular dynamics. Mutations throughout ICL1 can either enhance or disrupt cAMP production by CGRP at the CGRP receptor. Alanine mutagenesis identified subtle differences with regard elevation of iCa2+, with the distal end of the loop being particularly sensitive. ERK1/2 activation displayed little sensitivity to ICL1 mutation. A broadly similar pattern was observed with the glucagon receptor, although there were differences in significance of individual residues. Extending the study revealed that at the CRF1 receptor, an insertion in ICL1 switched signaling bias between iCa2+ and cAMP. Molecular dynamics suggested that changes in ICL1 altered the conformation of ICL2 and the H8/TM7 junction (ICL4). For H8, alanine mutagenesis showed the importance of E3908.49b for all three signal transduction pathways, for the CGRP receptor, but mutations of other residues largely just altered ERK1/2 activation. Thus, ICL1 may modulate GPCR bias via interactions with ICL2, ICL4 and the Gß subunit.

Invasive non-native species likely to threaten biodiversity and ecosystems in the Antarctic Peninsula region.

The Antarctic is considered to be a pristine environment relative to other regions of the Earth, but it is increasingly vulnerable to invasions by marine, freshwater and terrestrial non-native species. The Antarctic Peninsula region (APR), which encompasses the Antarctic Peninsula, South Shetland Islands and South Orkney Islands, is by far the most invaded part of the Antarctica continent. The risk of introduction of invasive non-native species to the APR is likely to increase with predicted increases in the intensity, diversity and distribution of human activities. Parties that are signatories to the Antarctic Treaty have called for regional assessments of non-native species risk. In response, taxonomic and Antarctic experts undertook a horizon scanning exercise using expert opinion and consensus approaches to identify the species that are likely to present the highest risk to biodiversity and ecosystems within the APR over the next 10 years. One hundred and three species, currently absent in the APR, were identified as relevant for review, with 13 species identified as presenting a high risk of invading the APR. Marine invertebrates dominated the list of highest risk species, with flowering plants and terrestrial invertebrates also represented; however, vertebrate species were thought unlikely to establish in the APR within the 10 year timeframe. We recommend (a) the further development and application of biosecurity measures by all stakeholders active in the APR, including surveillance for species such as those identified during this horizon scanning exercise, and (b) use of this methodology across the other regions of Antarctica. Without the application of appropriate biosecurity measures, rates of introductions and invasions within the APR are likely to increase, resulting in negative consequences for the biodiversity of the whole continent, as introduced species establish and spread further due to climate change and increasing human activity.

Elevated intracellular cAMP concentration mediates growth suppression in glioma cells.

Supressed levels of intracellular cAMP have been associated with malignancy. Thus, elevating cAMP through activation of adenylyl cyclase (AC) or by inhibition of phosphodiesterase (PDE) may be therapeutically beneficial. Here, we demonstrate that elevated cAMP levels suppress growth in C6 cells (a model of glioma) through treatment with forskolin, an AC activator, or a range of small molecule PDE inhibitors with differing selectivity profiles. Forskolin suppressed cell growth in a PKA-dependent manner by inducing a G2/M phase cell cycle arrest. In contrast, trequinsin (a non-selective PDE2/3/7 inhibitor), not only inhibited cell growth via PKA, but also stimulated (independent of PKA) caspase-3/-7 and induced an aneuploidy phenotype. Interestingly, a cocktail of individual PDE 2,3,7 inhibitors suppressed cell growth in a manner analogous to forskolin but not trequinsin. Finally, we demonstrate that concomitant targeting of both AC and PDEs synergistically elevated intracellular cAMP levels thereby potentiating their antiproliferative actions.

Effects of size- and sex-selective harvesting: An integral projection model approach.

Harvesting is often size-selective, and in species with sexual size dimorphism, it may also be sex-selective. A powerful approach to investigate potential consequences of size- and/or sex-selective harvesting is to simulate it in a demographic population model. We developed a population-based integral projection model for a size- and sex-structured species, the commonly exploited pike (Esox lucius). The model allows reproductive success to be proportional to body size and potentially limited by both sexes. We ran all harvest simulations with both lower size limits and slot limits, and to quantify the effects of selective harvesting, we calculated sex ratios and the long-term population growth rate (λ). In addition, we quantified to what degree purely size-selective harvesting was sex-selective, and determined when λ shifted from being female to male limited under size- and sex-selective harvesting. We found that purely size-selective harvest can be sex-selective, and that it depends on the harvest limits and the size distributions of the sexes. For the size- and sex-selective harvest simulations, λ increased with harvest intensity up to a threshold as females limited reproduction. Beyond this threshold, males became the limiting sex, and λ decreased as more males were harvested. The peak in λ, and the corresponding sex ratio in harvest, varied with both the selectivity and the intensity of the harvest simulation. Our model represents a useful extension of size-structured population models as it includes both sexes, relaxes the assumption of female dominance, and accounts for size-dependent fecundity. The consequences of selective harvesting presented here are especially relevant for size- and sex-structured exploited species, such as commercial fisheries. Thus, our model provides a useful contribution toward the development of more sustainable harvesting regimes.

Interactions between RAMP2 and CRF receptors: The effect of receptor subtypes, splice variants and cell context.

Corticotrophin releasing factor (CRF) acts via two family B G-protein-coupled receptors, CRFR1 and CRFR2. Additional subtypes exist due to alternative splicing. CRFR1α is the most widely expressed subtype and lacks a 29-residue insert in the first intracellular loop that is present in CRFR1ß. It has been shown previously that co-expression of CRFR1ß with receptor activity modifying protein 2 (RAMP2) in HEK 293S cells increased the cell-surface expression of both proteins suggesting a physical interaction as seen with RAMPs and calcitonin receptor-like receptor (CLR). This study investigated the ability of CRFR1α, CRFR1ß and CRFR2ß to promote cell-surface expression of FLAG-tagged RAMP2. Four different cell-lines were utilised to investigate the effect of varying cellular context; COS-7, HEK 293T, HEK 293S and [ΔCTR]HEK 293 (which lacks endogenous calcitonin receptor). In all cell-lines, CRFR1α and CRFR1ß enhanced RAMP2 cell-surface expression. The magnitude of the effect on RAMP2 was dependent on the cell-line ([ΔCTR]HEK 293 > COS-7 > HEK 293T > HEK 293S). RT-PCR indicated this variation may relate to differences in endogenous RAMP expression between cell types. Furthermore, pre-treatment with CRF resulted in a loss of cell-surface FLAG-RAMP2 when it was co-expressed with CRFR1 subtypes. CRFR2ß co-expression had no effect on RAMP2 in any cell-line. Molecular modelling suggests that the potential contact interface between the extracellular domains of RAMP2 and CRF receptor subtypes is smaller than that of RAMP2 and CRL, the canonical receptor:RAMP pairing, assuming a physical interaction. Furthermore, a specific residue difference between CRFR1 subtypes (glutamate) and CRFR2ß (histidine) in this interface region may impair CRFR2ß:RAMP2 interaction by electrostatic repulsion.

Assessing the role of bed sediments in the persistence of red mud pollution in a shallow lake (Kinghorn Loch, UK).

Red mud is a by-product of alumina production. Little is known about the long-term fate of red mud constituents in fresh waters or of the processes regulating recovery of fresh waters following pollution control. In 1983, red mud leachate was diverted away from Kinghorn Loch, UK, after many years of polluting this shallow and monomictic lake. We hypothesised that the redox-sensitive constituents of red mud leachate, phosphorus (P), arsenic (As) and vanadium (V), would persist in the Kinghorn Loch for many years following pollution control as a result of cycling between the lake bed sediment and the overlying water column. To test this hypothesis, we conducted a 12-month field campaign in Kinghorn Loch between May 2012 and April 2013 to quantify the seasonal cycling of P, As, and V in relation to environmental conditions (e.g., dissolved oxygen (DO) concentration, pH, redox chemistry and temperature) in the lake surface and bottom waters. To confirm the mechanisms for P, As and V release, a sediment core incubation experiment was conducted using lake sediment sampled in July 2012, in which DO concentrations were manipulated to create either oxic or anoxic conditions similar to the bed conditions found in the lake. The effects on P, As, and V concentrations and species in the water column were measured daily over an eight-day incubation period. Phosphate (PO4-P) and dissolved As concentrations were significantly higher in the bottom waters (75.9 ± 30.2 µg L-1 and 23.5 ± 1.83 µg L-1, respectively) than in the surface waters (12.9 ± 1.50 µg L-1 and 14.1 ± 2.20 µg L-1, respectively) in Kinghorn Loch. Sediment release of As and P under anoxic conditions was confirmed by the incubation experiment and by the significant negative correlations between DO and P and As concentrations in the bottom waters of the lake. In contrast, the highest dissolved V concentrations occurred in the bottom waters of Kinghorn Loch under oxic conditions (15.0 ± 3.35 µg L-1), with the release from the bed sediment apparently being controlled by a combination of competitive ion concentrations, pH and redox conditions.

Receptor Activity-modifying Protein-directed G Protein Signaling Specificity for the Calcitonin Gene-related Peptide Family of Receptors.

The calcitonin gene-related peptide (CGRP) family of G protein-coupled receptors (GPCRs) is formed through the association of the calcitonin receptor-like receptor (CLR) and one of three receptor activity-modifying proteins (RAMPs). Binding of one of the three peptide ligands, CGRP, adrenomedullin (AM), and intermedin/adrenomedullin 2 (AM2), is well known to result in a Gαs-mediated increase in cAMP. Here we used modified yeast strains that couple receptor activation to cell growth, via chimeric yeast/Gα subunits, and HEK-293 cells to characterize the effect of different RAMP and ligand combinations on this pathway. We not only demonstrate functional couplings to both Gαs and Gαq but also identify a Gαi component to CLR signaling in both yeast and HEK-293 cells, which is absent in HEK-293S cells. We show that the CGRP family of receptors displays both ligand- and RAMP-dependent signaling bias among the Gαs, Gαi, and Gαq/11 pathways. The results are discussed in the context of RAMP interactions probed through molecular modeling and molecular dynamics simulations of the RAMP-GPCR-G protein complexes. This study further highlights the importance of RAMPs to CLR pharmacology and to bias in general, as well as identifying the importance of choosing an appropriate model system for the study of GPCR pharmacology.

Phenological sensitivity to climate across taxa and trophic levels.

Differences in phenological responses to climate change among species can desynchronise ecological interactions and thereby threaten ecosystem function. To assess these threats, we must quantify the relative impact of climate change on species at different trophic levels. Here, we apply a Climate Sensitivity Profile approach to 10,003 terrestrial and aquatic phenological data sets, spatially matched to temperature and precipitation data, to quantify variation in climate sensitivity. The direction, magnitude and timing of climate sensitivity varied markedly among organisms within taxonomic and trophic groups. Despite this variability, we detected systematic variation in the direction and magnitude of phenological climate sensitivity. Secondary consumers showed consistently lower climate sensitivity than other groups. We used mid-century climate change projections to estimate that the timing of phenological events could change more for primary consumers than for species in other trophic levels (6.2 versus 2.5-2.9 days earlier on average), with substantial taxonomic variation (1.1-14.8 days earlier on average).

Assessing the Legacy of Red Mud Pollution in a Shallow Freshwater Lake: Arsenic Accumulation and Speciation in Macrophytes.

Little is known about long-term ecological responses in lakes following red mud pollution. Among red mud contaminants, arsenic (As) is of considerable concern. Determination of the species of As accumulated in aquatic organisms provides important information about the biogeochemical cycling of the element and transfer through the aquatic food-web to higher organisms. We used coupled ion chromatography and inductively coupled plasma mass spectrometry (ICP-MS) to assess As speciation in tissues of five macrophyte taxa in Kinghorn Loch, U.K., 30 years following the diversion of red mud pollution from the lake. Toxic inorganic As was the dominant species in the studied macrophytes, with As species concentrations varying with macrophyte taxon and tissue type. The highest As content measured in roots of Persicaria amphibia (L.) Gray (87.2 mg kg(-1)) greatly exceeded the 3-10 mg kg(-1) range suggested as a potential phytotoxic level. Accumulation of toxic As species by plants suggested toxicological risk to higher organisms known to utilize macrophytes as a food source.

Environmental DNA metabarcoding of lake fish communities reflects long-term data from established survey methods.

Organisms continuously release DNA into their environments via shed cells, excreta, gametes and decaying material. Analysis of this 'environmental DNA' (eDNA) is revolutionizing biodiversity monitoring. eDNA outperforms many established survey methods for targeted detection of single species, but few studies have investigated how well eDNA reflects whole communities of organisms in natural environments. We investigated whether eDNA can recover accurate qualitative and quantitative information about fish communities in large lakes, by comparison to the most comprehensive long-term gill-net data set available in the UK. Seventy-eight 2L water samples were collected along depth profile transects, gill-net sites and from the shoreline in three large, deep lakes (Windermere, Bassenthwaite Lake and Derwent Water) in the English Lake District. Water samples were assayed by eDNA metabarcoding of the mitochondrial 12S and cytochrome b regions. Fourteen of the 16 species historically recorded in Windermere were detected using eDNA, compared to four species in the most recent gill-net survey, demonstrating eDNA is extremely sensitive for detecting species. A key question for biodiversity monitoring is whether eDNA can accurately estimate abundance. To test this, we used the number of sequence reads per species and the proportion of sampling sites in which a species was detected with eDNA (i.e. site occupancy) as proxies for abundance. eDNA abundance data consistently correlated with rank abundance estimates from established surveys. These results demonstrate that eDNA metabarcoding can describe fish communities in large lakes, both qualitatively and quantitatively, and has great potential as a complementary tool to established monitoring methods.

Pathogens trigger top-down climate forcing on ecosystem dynamics.

Evaluating the effects of climate variation on ecosystems is of paramount importance for our ability to forecast and mitigate the consequences of global change. However, the ways in which complex food webs respond to climate variations remain poorly understood. Here, we use long-term time series to investigate the effects of temperature variation on the intraguild-predation (IGP) system of Windermere (UK), a lake where pike (Esox lucius, top predator) feed on small-sized perch (Perca fluviatilis) but compete with large-sized perch for the same food sources. Spectral analyses of time series reveal that pike recruitment dynamics are temperature controlled. In 1976, expansion of a size-truncating perch pathogen into the lake severely impacted large perch and favoured pike as the IGP-dominant species. This pathogen-induced regime shift to a pike-dominated IGP apparently triggered a temperature-controlled trophic cascade passing through pike down to dissolved nutrients. In simple food chains, warming is predicted to strengthen top-down control by accelerating metabolic rates in ectothermic consumers, while pathogens of top consumers are predicted to dampen this top-down control. In contrast, the local IGP structure in Windermere made warming and pathogens synergistic in their top-down effects on ecosystem functioning. More generally, our results point to top predators as major mediators of community response to global change, and show that size-selective agents (e.g. pathogens, fishers or hunters) may change the topological architecture of food webs and alter whole ecosystem sensitivity to climate variation.

Fitness consequences of early life conditions and maternal size effects in a freshwater top predator.

Conditions experienced in early life stages can be an important determinant of individual life histories. In fish, environmental conditions are known to affect early survival and growth, but recent studies have also emphasized maternal effects mediated by size or age. However, the relative sensitivity of the mean fitness (population growth rate λ) to different early life impacts remains largely unexplored. Using a female-based integral projection model (IPM) parameterized from unique long-term demographic data for pike (Esox lucius), we evaluated the relative fitness consequences of different early life impacts, including (i) maternal effects of length on egg weight, potentially affecting offspring (first year) survival, and (ii) effects of temperature on offspring growth and survival. Of the seven vital rates defining the model, offspring survival could not be directly estimated and four scenarios were defined for this rate. Elasticity analyses of the IPM were performed to calculate (i) the total contribution from different lengths to the elasticity of λ to the projection kernel, and (ii) the elasticity of λ to underlying variables of female current length, female offspring length at age 1, and temperature. These elasticities were decomposed into contributions from different vital rates across length. Egg weight increased with female length, as expected, but the effect leveled off for the largest females. However, λ was largely insensitive to this effect, even when egg weight was assumed to have a strong effect on offspring survival. In contrast, λ was sensitive to early temperature conditions through growth and survival. Among mature females, the total elasticity of λ to the projection kernel generally increased with length. The results were robust to a wide range of assumptions. These results suggest that environmental conditions experienced in early life represent a more important driver of mean population growth and fitness of pike than maternal effects of size on offspring survival. We discuss two general mechanisms underlying the weak influence of this maternal effect, suggesting that these may be general for long-lived and highly fecund fishes. This model and results are relevant for the management of long-lived top predators, including many commercially important fish species.

Discovery of Novel Adenosine Receptor Agonists That Exhibit Subtype Selectivity.

A series of N(6)-bicyclic and N(6)-(2-hydroxy)cyclopentyl derivatives of adenosine were synthesized as novel A1R agonists and their A1R/A2R selectivity assessed using a simple yeast screening platform. We observed that the most selective, high potency ligands were achieved through N(6)-adamantyl substitution in combination with 5'-N-ethylcarboxamido or 5'-hydroxymethyl groups. In addition, we determined that 5'-(2-fluoro)thiophenyl derivatives all failed to generate a signaling response despite showing an interaction with the A1R. Some selected compounds were also tested on A1R and A3R in mammalian cells revealing that four of them are entirely A1R-selective agonists. By using in silico homology modeling and ligand docking, we provide insight into their mechanisms of recognition and activation of the A1R. We believe that given the broad tissue distribution, but contrasting signaling profiles, of adenosine receptor subtypes, these compounds might have therapeutic potential.