Global patterns of allochthony in stream-riparian meta-ecosystems.

Ecosystems that are coupled by reciprocal flows of energy and nutrient subsidies can be viewed as a single "meta-ecosystem." Despite these connections, the reciprocal flow of subsidies is greatly asymmetrical and seasonally pulsed. Here, we synthesize existing literature on stream-riparian meta-ecosystems to quantify global patterns of the amount of subsidy consumption by organisms, known as "allochthony." These resource flows are important since they can comprise a large portion of consumer diets, but can be disrupted by human modification of streams and riparian zones. Despite asymmetrical subsidy flows, we found stream and riparian consumer allochthony to be equivalent. Although both fish and stream invertebrates rely on seasonally pulsed allochthonous resources, we find allochthony varies seasonally only for fish, being nearly three times greater during the summer and fall than during the winter and spring. We also find that consumer allochthony varies with feeding traits for aquatic invertebrates, fish, and terrestrial arthropods, but not for terrestrial vertebrates. Finally, we find that allochthony varies by climate for aquatic invertebrates, being nearly twice as great in arid climates than in tropical climates, but not for fish. These findings are critical to understanding the consequences of global change, as ecosystem connections are being increasingly disrupted.

Strategies for selecting and managing equipment in a light microscopy facility.

Light microscopy facilities vary in the number of imaging systems and the scope of technologies they support. Each facility must craft an identity through the selection of equipment and development of staff in order to serve the needs of its local research environment. The process of crafting a light microscopy facility can be compared to curation of an art exhibition: great care should be given to the selection and placement of each object in order to make a coherent statement. Lay Description: Light microscopy facilities vary in the number of imaging systems and the scope of technologies they support. Each facility must develop an identity through the selection of equipment and development of staff in order to serve the needs of its local research environment. The process of crafting a light microscopy facility can be compared to curation of an art exhibition: great care should be given to the selection and placement of each object in order to make a coherent statement.

Long transients and dendritic network structure affect spatial predator-prey dynamics in experimental microcosms.

Spatial dynamics can promote persistence of strongly interacting predators and prey. Theory predicts that spatial predator-prey systems are prone to long transients, meaning that the dynamics leading to persistence or extinction manifest over hundreds of generations. Furthermore, the form and duration of transients may be altered by spatial network structure. Few empirical studies have examined the importance of transients in spatial food webs, especially in a network context, due to the difficulty in collecting the large scale and long-term data required. We examined predator-prey dynamics in protist microcosms using three experimental spatial structures: isolated, river-like dendritic networks and regular lattice networks. Densities and patterns of occupancy were followed for both predators and prey over a time scale that equates to >100 predator and >500 prey generations. We found that predators persisted in dendritic and lattice networks whereas they went extinct in the isolated treatment. The dynamics leading to predator persistence played out over long transients with three distinct phases. The transient phases showed differences between dendritic and lattice structures, as did underlying patterns of occupancy. Spatial dynamics differed among organisms in different trophic positions. Predators showed higher local persistence in more connected bottles while prey showed this in more spatially isolated ones. Predictions based on spatial patterns of connectivity derived from metapopulation theory explained predator occupancy, while prey occupancy was better explained by predator occupancy. Our results strongly support the hypothesized role of spatial dynamics in promoting persistence in food webs, but that the dynamics ultimately leading to persistence may occur with long transients which in turn may be influenced by spatial network structure and trophic interactions.

PtdIns(3,4,5)P3-dependent Rac exchanger 1 (P-Rex1) promotes mammary tumor initiation and metastasis.

The Rac-GEF, P-Rex1, activates Rac1 signaling downstream of G protein-coupled receptors and PI3K. Increased P-Rex1 expression promotes melanoma progression; however, its role in breast cancer is complex, with differing reports of the effect of its expression on disease outcome. To address this we analyzed human databases, undertook gene array expression analysis, and generated unique murine models of P-Rex1 gain or loss of function. Analysis of PREX1 mRNA expression in breast cancer cDNA arrays and a METABRIC cohort revealed that higher PREX1 mRNA in ER+ve/luminal tumors was associated with poor outcome in luminal B cancers. Prex1 deletion in MMTV-neu or MMTV-PyMT mice reduced Rac1 activation in vivo and improved survival. High level MMTV-driven transgenic PREX1 expression resulted in apicobasal polarity defects and increased mammary epithelial cell proliferation associated with hyperplasia and development of de novo mammary tumors. MMTV-PREX1 expression in MMTV-neu mice increased tumor initiation and enhanced metastasis in vivo, but had no effect on primary tumor growth. Pharmacological inhibition of Rac1 or MEK1/2 reduced P-Rex1-driven tumoroid formation and cell invasion. Therefore, P-Rex1 can act as an oncogene and cooperate with HER2/neu to enhance breast cancer initiation and metastasis, despite having no effect on primary tumor growth.

What is the structure of our infrastructure? A review of UK light microscopy facilities.

Core Facilities and Technology Platforms are increasingly important components of the science research landscape. However, data on facility operations and staff careers are lacking to inform their development. Here we have surveyed 114 people working in 46 light microscopy (LM) facilities within the United Kingdom. Our survey explores issues around career progression, facility operations and funding. The data show that facilities are substantial repositories of equipment and knowledge which adapt to meet the needs of their local environments. Our report highlights the challenges faced by facility managers, institutions and funders in evaluating facility performance and devising strategies to maximise the return on research funding investment.

HIRA orchestrates a dynamic chromatin landscape in senescence and is required for suppression of neoplasia.

Cellular senescence is a stable proliferation arrest that suppresses tumorigenesis. Cellular senescence and associated tumor suppression depend on control of chromatin. Histone chaperone HIRA deposits variant histone H3.3 and histone H4 into chromatin in a DNA replication-independent manner. Appropriately for a DNA replication-independent chaperone, HIRA is involved in control of chromatin in nonproliferating senescent cells, although its role is poorly defined. Here, we show that nonproliferating senescent cells express and incorporate histone H3.3 and other canonical core histones into a dynamic chromatin landscape. Expression of canonical histones is linked to alternative mRNA splicing to eliminate signals that confer mRNA instability in nonproliferating cells. Deposition of newly synthesized histones H3.3 and H4 into chromatin of senescent cells depends on HIRA. HIRA and newly deposited H3.3 colocalize at promoters of expressed genes, partially redistributing between proliferating and senescent cells to parallel changes in expression. In senescent cells, but not proliferating cells, promoters of active genes are exceptionally enriched in H4K16ac, and HIRA is required for retention of H4K16ac. HIRA is also required for retention of H4K16ac in vivo and suppression of oncogene-induced neoplasia. These results show that HIRA controls a specialized, dynamic H4K16ac-decorated chromatin landscape in senescent cells and enforces tumor suppression.

Label-Free Multiphoton Microscopy: Much More Than Fancy Images.

Multiphoton microscopy has recently passed the milestone of its first 30 years of activity in biomedical research. The growing interest around this approach has led to a variety of applications from basic research to clinical practice. Moreover, this technique offers the advantage of label-free multiphoton imaging to analyze samples without staining processes and the need for a dedicated system. Here, we review the state of the art of label-free techniques; then, we focus on two-photon autofluorescence as well as second and third harmonic generation, describing physical and technical characteristics. We summarize some successful applications to a plethora of biomedical research fields and samples, underlying the versatility of this technique. A paragraph is dedicated to an overview of sample preparation, which is a crucial step in every microscopy experiment. Afterwards, we provide a detailed review analysis of the main quantitative methods to extract important information and parameters from acquired images using second harmonic generation. Lastly, we discuss advantages, limitations, and future perspectives in label-free multiphoton microscopy.

p53 status determines the role of autophagy in pancreatic tumour development.

Macroautophagy (hereafter referred to as autophagy) is a process in which organelles termed autophagosomes deliver cytoplasmic constituents to lysosomes for degradation. Autophagy has a major role in cellular homeostasis and has been implicated in various forms of human disease. The role of autophagy in cancer seems to be complex, with reports indicating both pro-tumorigenic and tumour-suppressive roles. Here we show, in a humanized genetically-modified mouse model of pancreatic ductal adenocarcinoma (PDAC), that autophagy's role in tumour development is intrinsically connected to the status of the tumour suppressor p53. Mice with pancreases containing an activated oncogenic allele of Kras (also called Ki-Ras)--the most common mutational event in PDAC--develop a small number of pre-cancerous lesions that stochastically develop into PDAC over time. However, mice also lacking the essential autophagy genes Atg5 or Atg7 accumulate low-grade, pre-malignant pancreatic intraepithelial neoplasia lesions, but progression to high-grade pancreatic intraepithelial neoplasias and PDAC is blocked. In marked contrast, in mice containing oncogenic Kras and lacking p53, loss of autophagy no longer blocks tumour progression, but actually accelerates tumour onset, with metabolic analysis revealing enhanced glucose uptake and enrichment of anabolic pathways, which can fuel tumour growth. These findings provide considerable insight into the role of autophagy in cancer and have important implications for autophagy inhibition in cancer therapy. In this regard, we also show that treatment of mice with the autophagy inhibitor hydroxychloroquine, which is currently being used in several clinical trials, significantly accelerates tumour formation in mice containing oncogenic Kras but lacking p53.

Predicting Pattern Formation in Multilayer Networks.

We investigate how the structure of interactions between coupled oscillators influences the formation of asynchronous patterns in a multilayer network by formulating a simple, general multilayer oscillator model. We demonstrate the analysis of this model in three-oscillator systems, illustrating the role of interactions among oscillators in sustaining differences in both the phase and amplitude of oscillations leading to the formation of asynchronous patterns. Finally, we demonstrate the generalizability of our model's predictions through comparison with a more realistic multilayer model. Overall, our model provides a useful approach for predicting the types of asynchronous patterns that multilayer networks of coupled oscillators which cannot be achieved by the existing methods which focus on characterizing the synchronous state.

Effects of uncertainty and variability on population declines and IUCN Red List classifications.

The International Union for Conservation of Nature (IUCN) Red List Categories and Criteria is a quantitative framework for classifying species according to extinction risk. Population models may be used to estimate extinction risk or population declines. Uncertainty and variability arise in threat classifications through measurement and process error in empirical data and uncertainty in the models used to estimate extinction risk and population declines. Furthermore, species traits are known to affect extinction risk. We investigated the effects of measurement and process error, model type, population growth rate, and age at first reproduction on the reliability of risk classifications based on projected population declines on IUCN Red List classifications. We used an age-structured population model to simulate true population trajectories with different growth rates, reproductive ages and levels of variation, and subjected them to measurement error. We evaluated the ability of scalar and matrix models parameterized with these simulated time series to accurately capture the IUCN Red List classification generated with true population declines. Under all levels of measurement error tested and low process error, classifications were reasonably accurate; scalar and matrix models yielded roughly the same rate of misclassifications, but the distribution of errors differed; matrix models led to greater overestimation of extinction risk than underestimations; process error tended to contribute to misclassifications to a greater extent than measurement error; and more misclassifications occurred for fast, rather than slow, life histories. These results indicate that classifications of highly threatened taxa (i.e., taxa with low growth rates) under criterion A are more likely to be reliable than for less threatened taxa when assessed with population models. Greater scrutiny needs to be placed on data used to parameterize population models for species with high growth rates, particularly when available evidence indicates a potential transition to higher risk categories.

Implications of different population model structures for management of threatened plants.

Population viability analysis (PVA) is a reliable tool for ranking management options for a range of species despite parameter uncertainty. No one has yet investigated whether this holds true for model uncertainty for species with complex life histories and for responses to multiple threats. We tested whether a range of model structures yielded similar rankings of management and threat scenarios for 2 plant species with complex postfire responses. We examined 2 contrasting species from different plant functional types: an obligate seeding shrub and a facultative resprouting shrub. We exposed each to altered fire regimes and an additional, species-specific threat. Long-term demographic data sets were used to construct an individual-based model (IBM), a complex stage-based model, and a simple matrix model that subsumes all life stages into 2 or 3 stages. Agreement across models was good under some scenarios and poor under others. Results from the simple and complex matrix models were more similar to each other than to the IBM. Results were robust across models when dominant threats are considered but were less so for smaller effects. Robustness also broke down as the scenarios deviated from baseline conditions, likely the result of a number of factors related to the complexity of the species' life history and how it was represented in a model. Although PVA can be an invaluable tool for integrating data and understanding species' responses to threats and management strategies, this is best achieved in the context of decision support for adaptive management alongside multiple lines of evidence and expert critique of model construction and output.

Proteomics-based metabolic modeling reveals that fatty acid oxidation (FAO) controls endothelial cell (EC) permeability.

Endothelial cells (ECs) play a key role to maintain the functionality of blood vessels. Altered EC permeability causes severe impairment in vessel stability and is a hallmark of pathologies such as cancer and thrombosis. Integrating label-free quantitative proteomics data into genome-wide metabolic modeling, we built up a model that predicts the metabolic fluxes in ECs when cultured on a tridimensional matrix and organize into a vascular-like network. We discovered how fatty acid oxidation increases when ECs are assembled into a fully formed network that can be disrupted by inhibiting CPT1A, the fatty acid oxidation rate-limiting enzyme. Acute CPT1A inhibition reduces cellular ATP levels and oxygen consumption, which are restored by replenishing the tricarboxylic acid cycle. Remarkably, global phosphoproteomic changes measured upon acute CPT1A inhibition pinpointed altered calcium signaling. Indeed, CPT1A inhibition increases intracellular calcium oscillations. Finally, inhibiting CPT1A induces hyperpermeability in vitro and leakage of blood vessel in vivo, which were restored blocking calcium influx or replenishing the tricarboxylic acid cycle. Fatty acid oxidation emerges as central regulator of endothelial functions and blood vessel stability and druggable pathway to control pathological vascular permeability.

Geometric indicators of population persistence in branching continuous-space networks.

We study population persistence in branching tree networks emulating systems such as river basins, cave systems, organisms on vegetation surfaces, and vascular networks. Population dynamics are modeled using a reaction-diffusion-advection equation on a metric graph which provides a continuous, spatially explicit model of network habitat. A metric graph, in contrast to a standard graph, allows for population dynamics to occur within edges rather than just at graph vertices, subsequently adding a significant level of realism. Within this framework, we stochastically generate branching tree networks with a variety of geometric features and explore the effects of network geometry on the persistence of a population which advects toward a lethal outflow boundary. We identify a metric (CM), the distance from the lethal outflow point at which half of the habitable volume of the network lies upstream of, as a promising indicator of population persistence. This metric outperforms other metrics such as the maximum and minimum distances from the lethal outflow to an upstream boundary and the total habitable volume of the network. The strength of CM as a predictor of persistence suggests that it is a proper "system length" for the branching networks we examine here that generalizes the concept of habitat length in the classical linear space models.

In-Depth Proteomics Identifies a Role for Autophagy in Controlling Reactive Oxygen Species Mediated Endothelial Permeability.

Endothelial cells (ECs) form the inner layer of blood vessels and physically separate the blood from the surrounding tissue. To support tissues with nutrients and oxygen, the endothelial monolayer is semipermeable. When EC permeability is altered, blood vessels are not functional, and this is associated with disease. A comprehensive knowledge of the mechanisms regulating EC permeability is key in developing strategies to target this mechanism in pathologies. Here we have used an in vitro model of human umbilical vein endothelial cells mimicking the formation of a physiologically permeable vessel and performed time-resolved in-depth molecular profiling using stable isotope labeling by amino acids in cell culture mass spectrometry (MS)-proteomics. Autophagy is induced when ECs are assembled into a physiologically permeable monolayer. By using siRNA and drug treatment to block autophagy in combination with functional assays and MS proteomics, we show that ECs require autophagy flux to maintain intracellular reactive oxygen species levels, and this is required to maintain the physiological permeability of the cells.

Colonisation rate and adaptive foraging control the emergence of trophic cascades.

Ecological communities are assembled and sustained by colonisation. At the same time, predators make foraging decisions based on the local availabilities of potential resources, which reflects colonisation. We combined field and laboratory experiments with mathematical models to demonstrate that a feedback between these two processes determines emergent patterns in community structure. Namely, our results show that prey colonisation rate determines the strength of trophic cascades - a feature of virtually all ecosystems - by prompting behavioural shifts in adaptively foraging omnivorous fish predators. Communities experiencing higher colonisation rates were characterised by higher invertebrate prey and lower producer biomasses. Consequently, fish functioned as predators when colonisation rate was high, but as herbivores when colonisation rate was low. Human land use is changing habitat connectivity worldwide. A deeper quantitative understanding of how spatial processes modify individual behaviour, and how this scales to the community level, will be required to predict ecosystem responses to these changes.

Can inducible resistance in plants cause herbivore aggregations? Spatial patterns in an inducible plant/herbivore model.

Many theories regarding the evolution of inducible resistance in plants have an implicit spatial component, but most relevant population dynamic studies ignore spatial dynamics. We examined a spatially explicit model of plant inducible resistance and herbivore population dynamics to explore how realistic features of resistance and herbivore responses influence spatial patterning. Both transient and persistent spatial patterns developed in all models examined, where patterns manifested as wave-like aggregations of herbivores and variation in induction levels. Patterns arose when herbivores moved away from highly induced plants, there was a lag between damage and deployment of induced resistance, and the relationship between herbivore density and strength of the induction response had a sigmoid shape. These mechanisms influenced pattern formation regardless of the assumed functional relationship between resistance and herbivore recruitment and mortality. However, in models where induction affected herbivore mortality, large-scale herbivore population cycles driven by the mortality response often co-occurred with smaller scale spatial patterns driven by herbivore movement. When the mortality effect dominated, however, spatial pattern formation was completely replaced by spatially synchronized herbivore population cycles. Our results present a new type of ecological pattern formation driven by induced trait variation, consumer behavior, and time delays that has broad implications for the community and evolutionary ecology of plant defenses.

Targeting mTOR dependency in pancreatic cancer.

OBJECTIVE: Pancreatic cancer is a leading cause of cancer-related death in the Western world. Current chemotherapy regimens have modest survival benefit. Thus, novel, effective therapies are required for treatment of this disease. DESIGN: Activating KRAS mutation almost always drives pancreatic tumour initiation, however, deregulation of other potentially druggable pathways promotes tumour progression. PTEN loss leads to acceleration of Kras(G12D)-driven pancreatic ductal adenocarcinoma (PDAC) in mice and these tumours have high levels of mammalian target of rapamycin (mTOR) signalling. To test whether these KRAS PTEN pancreatic tumours show mTOR dependence, we compared response to mTOR inhibition in this model, to the response in another established model of pancreatic cancer, KRAS P53. We also assessed whether there was a subset of pancreatic cancer patients who may respond to mTOR inhibition. RESULTS: We found that tumours in KRAS PTEN mice exhibit a remarkable dependence on mTOR signalling. In these tumours, mTOR inhibition leads to proliferative arrest and even tumour regression. Further, we could measure response using clinically applicable positron emission tomography imaging. Importantly, pancreatic tumours driven by activated KRAS and mutant p53 did not respond to treatment. In human tumours, approximately 20% of cases demonstrated low PTEN expression and a gene expression signature that overlaps with murine KRAS PTEN tumours. CONCLUSIONS: KRAS PTEN tumours are uniquely responsive to mTOR inhibition. Targeted anti-mTOR therapies may offer clinical benefit in subsets of human PDAC selected based on genotype, that are dependent on mTOR signalling. Thus, the genetic signatures of human tumours could be used to direct pancreatic cancer treatment in the future.

Synthesis of migrastatin and its macroketone analogue and in vivo FRAP analysis of the macroketone on E-cadherin dynamics.

An efficient and scalable synthesis of a key acyclic intermediate used for the preparation of migrastatin and its macroketone analogue is described; Brown alkoxyallylation is the key step for this synthesis. The macroketone was prepared on 100 mg scale by this route. Treatment of invasive pancreatic cancer cells grown on a cell-derived matrix or as subcutaneous tumours in nude mice with the macroketone inhibited E-cadherin dynamics in a manner consistent with increased cell adhesion and reduced invasive potential.

Astroglial-axonal interactions during early stages of myelination in mixed cultures using in vitro and ex vivo imaging techniques.

BACKGROUND: [corrected] Myelination is a very complex process that requires the cross talk between various neural cell types. Previously, using cytosolic or membrane associated GFP tagged neurospheres, we followed the interaction of oligodendrocytes with axons using time-lapse imaging in vitro and ex vivo and demonstrated dynamic changes in cell morphology. In this study we focus on GFP tagged astrocytes differentiated from neurospheres and their interactions with axons. RESULTS: We show the close interaction of astrocyte processes with axons and with oligodendrocytes in mixed mouse spinal cord cultures with formation of membrane blebs as previously seen for oligodendrocytes in the same cultures. When GFP-tagged neurospheres were transplanted into the spinal cord of the dysmyelinated shiverer mouse, confirmation of dynamic changes in cell morphology was provided and a prevalence for astrocyte differentiation compared with oligodendroglial differentiation around the injection site. Furthermore, we were able to image GFP tagged neural cells in vivo after transplantation and the cells exhibited similar membrane changes as cells visualised in vitro and ex vivo. CONCLUSION: These data show that astrocytes exhibit dynamic cell process movement and changes in their membrane topography as they interact with axons and oligodendrocytes during the process of myelination, with the first demonstration of bleb formation in astrocytes.