The stoichiometry of P2X2/6 receptor heteromers depends on relative subunit expression levels

Fast synaptic transmission involves the operation of ionotropic receptors, which are often composed of at least two types of subunit. We have developed a method, based on atomic force microscopy imaging to determine the stoichiometry and subunit arrangement within ionotropic receptors. We showed recently that the P2X(2) receptor for ATP is expressed as a trimer but that the P2X(6) subunit is unable to oligomerize. In this study we addressed the subunit stoichiometry of heteromers containing both P2X(2) and P2X(6) subunits. We transfected tsA 201 cells with both P2X(2) and P2X(6) subunits, bearing different epitope tags. We manipulated the transfection conditions so that either P2X(2) or P2X(6) was the predominant subunit expressed. By atomic force microscopy imaging of isolated receptors decorated with antiepitope antibodies, we demonstrate that when expression of the P2X(2) subunit predominates, the receptors contain primarily 2 x P2X(2) subunits and 1 x P2X(6) subunit. In contrast, when the P2X(6) subunit predominates, the subunit stoichiometry of the receptors is reversed. Our results show that the composition of P2X receptor heteromers is plastic and dependent on the relative subunit expression levels. We suggest that this property of receptor assembly might introduce an additional layer of subtlety into P2X receptor signaling

Alpha- and Gammaproteobacterial Methanotrophs Codominate the Active Methane-Oxidizing Communities in an Acidic Boreal Peat Bog

The objective of this study was to characterize metabolically active, aerobic methanotrophs in an ombrotrophic peatland in the Marcell Experimental Forest, Minnesota, USA. Methanotrophs were investigated in the field and in laboratory incubations using DNA-stable isotope probing, expression studies on particulate methane monooxygenase (pmoA) genes, and amplicon sequencing of 16S rRNA genes. Potential rates of oxidation ranged from 14-17 μmol CH4 g dry wt soil-1 d-1. Within DNA-SIP incubations, the relative abundance of methanotrophs increased from 4% in situ to 25-36% after 8 -14 days. Phylogenetic analysis of the 13C-enriched DNA fractions revealed the active methanotrophs were dominated by the genera Methylocystis (Type II; Alphaproteobacteria), Methylomonas, and Methylovulum (Type I; Gammaproteobacteria). In field samples, a transcript-to-gene ratio of 1 to 2 was observed for pmoA in surface peat layers which attenuated rapidly with depth, indicating the highest methane consumption was associated with the 0-10 cm depth interval. Metagenomes and sequencing of cDNA pmoA amplicons from field samples confirmed the dominant active methanotrophs were Methylocystis and Methylomonas. Although Type II methanotrophs have long been shown to mediate methane consumption in peatlands, our results indicate members of the genera Methylomonas and Methylovulum (Type I) can significantly contribute to aerobic methane oxidation in these ecosystems

Interplay between substrate rigidity and tissue fluidity regulates cell monolayer spreading

Coordinated and cooperative motion of cells is essential for embryonic development, tissue morphogenesis, wound healing and cancer invasion. A predictive understanding of the emergent mechanical behaviors in collective cell motion is challenging due to the complex interplay between cell-cell interactions, cell-matrix adhesions and active cell behaviors. To overcome this challenge, we develop a predictive cellular vertex model that can delineate the relative roles of substrate rigidity, tissue mechanics and active cell properties on the movement of cell collectives. We apply the model to the specific case of collective motion in cell aggregates as they spread into a two-dimensional cell monolayer adherent to a soft elastic matrix. Consistent with recent experiments, we find that substrate stiffness regulates the driving forces for the spreading of cellular monolayer, which can be pressure-driven or crawling-based depending on substrate rigidity. On soft substrates, cell monolayer spreading is driven by an active pressure due to the influx of cells coming from the aggregate, whereas on stiff substrates, cell spreading is driven primarily by active crawling forces. Our model predicts that cooperation of cell crawling and tissue pressure drives faster spreading, while the spreading rate is sensitive to the mechanical properties of the tissue. We find that solid tissues spread faster on stiff substrates, with spreading rate increasing with tissue tension. By contrast, the spreading of fluid tissues is independent of substrate stiffness and is slower than solid tissues. We compare our theoretical results with experimental results on traction force generation and spreading kinetics of cell monolayers, and provide new predictions on the role of tissue fluidity and substrate rigidity on collective cell motion.Comment: revised paper title, more references adde

Ecological theory predicts ecosystem stressor interactions in freshwater communities, but highlights the strengths and weaknesses of the additive null model

Understanding and predicting how multiple co-occurring environmental stressors combine to affect biodiversity and ecosystem services is an on-going grand challenge for ecology. So far progress has been made through accumulating large numbers of smaller-scale individual studies that are then investigated by meta-analyses to look for general patterns. In particular there has been an interest in checking for so-called ecological surprises where stressors interact in a synergistic manner. Recent reviews suggest that such synergisms do not dominate, but few other generalities have emerged. This lack of general prediction and understanding may be due in part to a dearth of ecological theory that can generate clear hypotheses and predictions to tested against empirical data. Here we close this gap by analysing food web models based upon classical ecological theory and comparing their predictions to a large (546 interactions) dataset for the effects of pairs of stressors on freshwater communities, using trophic- and population-level metrics of abundance, density, and biomass as responses. We find excellent overall agreement between the stochastic version of our models and the experimental data, and both conclude additive stressor interactions are the most frequent, but that meta-analyses report antagonistic summary interaction classes. Additionally, we show that the statistical tests used to classify the interactions are very sensitive to sampling variation. It is therefore likely that current weak sampling and low sample sizes are masking many non-additive stressor interactions, which our theory predicts to dominate when sampling variation is removed. This leads us to suspect ecological surprises may be more common than currently reported. Our results highlight the value of developing theory in tandem with empirical tests, and the need to examine the robustness of statistical machinery, especially the widely-used null models, before we can draw strong conclusions about how environmental drivers combine

Stribild: A Review of Component Characteristics and Combination Drug Efficacy

BACKGROUND: Numerous methods have been devised to combat human immunodeficiency virus (HIV) replication and disease progression. Composed of an integrase strand transfer inhibitor, a pharmacoenhancer, and two reverse transcriptase inhibitors, Stribild is a relatively new combination HIV drug formulated for once-a-day dosing. METHODS: Relevant information, original research articles and reviews, were gathered primarily through the use of the PubMed database. The search was conducted without date restrictions in order to collect both historical and recent information concerning HIV, individual drugs, and combinations for a thorough overview. RESULTS: Stribild, when taken with food, provides therapeutic drug concentrations as seen through comparison with the respective individual or boosted individual drugs. Stribild non-inferiority has been shown when compared to other HIV drug combinations, ritonavir-boosted atazanavir or efavirenz each with a tenofovir disoproxil fumarate (TDF) and emtricitabine (FTC) backbone. The co-formulation also retained high viral suppression in patients switching from other regimens, such as efavirenz/TDF/FTC, raltegravir/TDF/FTC, or various ritonavir-boosted protease inhibitors with TDF/FTC. The elvitegravir and cobicistat combination was unaffected by moderate hepatic impairment; however, hepatic and renal function along with changes in bone mineral density should be monitored closely. Stribild presented with relatively few side effect occurrences, but drug interactions may pose a larger problem for continuous therapy. CONCLUSIONS: Stribild provides viral suppression, comparable to other combination HIV drugs through review of non-inferiority and regimen simplification studies, with minimal adverse effects. Although the breadth of Stribild effectiveness has begun to unfold, studies are lacking in older patients as well as adolescents

IL-21 receptor expression in human tendinopathy

The pathogenetic mechanisms underlying tendinopathy remain unclear, with much debate as to whether inflammation or degradation has the prominent role. Increasing evidence points toward and early inflammatory infiltrate and associated inflammatory cytokine production in human and animal models of tendon disease. The IL-21/IL-21R axis is a proinflammatory cytokine complex that has been associated with chronic inflammatory diseases including rheumatoid arthritis and inflammatory bowel disease. This project aimed to investigate the role and expression of the cytokine/receptor pair IL-21/IL-21R in human tendinopathy. We found significantly elevated expression of IL-21 receptor message and protein in human tendon samples but found no convincing evidence of the presence of IL-21 at message or protein level. The level of expression of IL-21R message/protein in human tenocytes was significantly up regulated by proinflammatory cytokines (TNFα/IL-1β) in vitro. These findings demonstrate that IL-21R is present in early human tendinopathy mainly expressed by tenocytes and macrophages. Despite a lack of IL-21 expression these data again suggest that early tendinopathy has an inflammatory/cytokine phenotype, which may provide novel translational targets in the treatment of tendinopathy

Spectral estimation for spatial point patterns

This article determines how to implement spatial spectral analysis of point processes (in two dimensions or more), by establishing the moments of raw spectral summaries of point processes. We establish the first moments of raw direct spectral estimates such as the discrete Fourier transform of a point pattern. These have a number of surprising features that departs from the properties of raw spectral estimates of random fields and time series. As for random fields, the special case of isotropic processes warrants special attention, which we discuss. For time series and random fields white noise plays a special role, mirrored by the Poisson processes in the case of the point process. For random fields bilinear estimators are prevalent in spectral analysis. We discuss how to smooth any bilinear spectral estimator for a point process. We also determine how to taper this bilinear spectral estimator, how to calculate the periodogram, sample the wavenumbers and discuss the correlation of the periodogram. In parts this corresponds to recommending suitable separable as well as isotropic tapers in d dimensions. This, in aggregation, establishes the foundations for spectral analysis of point processes.Comment: 29 pages + 23 pages of supplements, 6 figure