Mice Expressing Low Levels of CalDAG-GEFI Exhibit Markedly Impaired Platelet Activation With Minor Impact on HemostasisHighlights

OBJECTIVE: The tight regulation of platelet adhesiveness, mediated by the αIIbβ3 integrin, is critical for hemostasis and prevention of thrombosis. We recently demonstrated that integrin affinity in platelets is controlled by the guanine nucleotide exchange factor, CalDAG-GEFI (CD-GEFI), and its target, RAP1. In this study, we investigated whether low-level expression of CD-GEFI leads to protection from thrombosis without pathological bleeding in mice. APPROACH AND RESULTS: Cdg1(low) mice were generated by knockin of human CD-GEFI cDNA into the mouse Cdg1 locus. CD-GEFI expression in platelets from Cdg1(low) mice was reduced by ≈90% when compared with controls. Activation of RAP1 and αIIbβ3 was abolished at low agonist concentrations and partially inhibited at high agonist concentrations in Cdg1(low) platelets. Consistently, the aggregation response of Cdg1(low) platelets was weaker than that of wild-type platelets, but more efficient than that observed in Cdg1(-/-) platelets. Importantly, Cdg1(low) mice were strongly protected from arterial and immune complex-mediated thrombosis, with only minimal impact on primary hemostasis. CONCLUSIONS: Together, our studies suggest the partial inhibition of CD-GEFI function as a powerful new approach to safely prevent thrombotic complications

The fibrinogen γA/γ′ isoform does not promote acute arterial thrombosis in mice

Elevated plasma fibrinogen associates with arterial thrombosis in humans and promotes thrombosis in mice by increasing fibrin formation and thrombus fibrin content. Fibrinogen is composed of six polypeptide chains: (Aα, Bβ, and γ)2. Alternative splicing of the γ chain leads to a dominant form (γA/γA) and a minor species (γA/γ’). Epidemiologic studies have detected elevated γA/γ’ fibrinogen in patients with arterial thrombosis, suggesting this isoform promotes thrombosis. However, in vitro data show that γA/γ’ is anticoagulant due to its ability to sequester thrombin, and suggest its expression is upregulated in response to inflammatory processes

Red blood cell-derived semaphorin 7A promotes thrombo-inflammation in myocardial ischemia-reperfusion injury through platelet GPIb.

Myocardial ischemia is one of the leading health problems worldwide. Therapy consists of the restitution of coronary perfusion which is followed by myocardial inflammation. Platelet-neutrophil interaction is a crucial process during inflammation, yet its consequences are not fully understood. Here, we show that platelet-neutrophil complexes (PNCs) are increased in patients with acute myocardial infarction and that this is associated with increased levels of neuronal guidance protein semaphorin 7A (SEMA7A). To investigate this further, we injected WT animals with Sema7a and found increased infarct size with increased numbers of PNCs. Experiments in genetically modified animals identify Sema7a on red blood cells to be crucial for this condition. Further studies revealed that Sema7a interacts with the platelet receptor glycoprotein Ib (GPIb). Treatment with anti-Sema7a antibody protected from myocardial tissue injury. In summary, we show that Sema7a binds to platelet GPIb and enhances platelet thrombo-inflammatory activity, aggravating post-ischemic myocardial tissue injury

Functional redundancy between RAP1 isoforms in murine platelet production and function

RAP GTPases, important regulators of cellular adhesion, are abundant signaling molecules in the platelet/megakaryocytic lineage. However, mice lacking the predominant isoform, RAP1B, display a partial platelet integrin activation defect and have a normal platelet count, suggesting the existence of a RAP1-independent pathway to integrin activation in platelets and a negligible role for RAP GTPases in megakaryocyte biology. To determine the importance of individual RAP isoforms on platelet production and on platelet activation at sites of mechanical injury or vascular leakage, we conditionally deleted Rap1a and/or Rap1b in the megakaryocytic lineage (mKO). Interestingly, Rap1a/b-mKO mice displayed a marked macrothrombocytopenia due to impaired pro- platelet formation by megakaryocytes. In platelets, RAP isoforms had both redundant and isoform-ˇspecific functions. Deletion of RAP1B, but not RAP1A, significantly reduced α- granule secretion and activation of the cytoskeleton regulator RAC1. Both isoforms significantly contributed to thromboxane A2 generation and the inside-out activation of platelet integrins. Combined deficiency of RAP1A and RAP1B markedly impaired platelet aggregation, spreading and clot retraction. Consistently, thrombus formation in physiological flow conditions was abolished in Rap1a/b-mKO, but not Rap1a-mKO or Rap1b-mKO platelets. Rap1a/b-mKO mice were strongly protected from experimental thrombosis and exhibited a severe defect in hemostasis after mechanical injury. Surprisingly, Rap1a/b-mKO platelets were indistinguishable from controls in their ability to prevent blood-lymphatic mixing during development and hemorrhage at sites of inflammation. In summary, our studies demonstrate an essential role for RAP1 signaling in platelet integrin activation and a critical role in platelet production. While important for hemostatic/thrombotic plug formation, platelet RAP1 signaling is dispensable for vascular integrity during development and inflammation

Disturbance indicator values for European plants

Motivation Indicator values are numerical values used to characterize the ecological niches of species and to estimate their occurrence along gradients. Indicator values on climatic and edaphic niches of plant species have received considerable attention in ecological research, whereas data on the optimal positioning of species along disturbance gradients are less developed. Here, we present a new data set of disturbance indicator values identifying optima along gradients of natural and anthropogenic disturbance for 6382 vascular plant species based on the analysis of 736,366 European vegetation plots and using expert-based characterization of disturbance regimes in 236 habitat types. The indicator values presented here are crucial for integrating disturbance niche optima into large-scale vegetation analyses and macroecological studies. Main types of variables contained We set up five main continuous indicator values for European vascular plants: disturbance severity, disturbance frequency, mowing frequency, grazing pressure and soil disturbance. The first two indicators are provided separately for the whole community and for the herb layer. We calculated the values as the average of expert-based estimates of disturbance values in all habitat types where a species occurs, weighted by the number of plots in which the species occurs within a given habitat type. Spatial location and grain Europe. Vegetation plots ranging in size from 1 to 1000 m(2). Time period and grain Vegetation plots mostly sampled between 1956 and 2013 (= 5th and 95th quantiles of the sampling year, respectively). Major taxa and level of measurement Species-level indicator values for vascular plants. Software format csv file

Chemoproteomic Discovery of AADACL1 as a Regulator of Human Platelet Activation

A comprehensive knowledge of the platelet proteome is necessary for understanding thrombosis and for conceiving novel antiplatelet therapies. To discover new biochemical pathways in human platelets, we screened platelets with a carbamate library designed to interrogate the serine hydrolase subproteome and used competitive activity-based protein profiling to map the targets of active carbamates. We identified an inhibitor that targets arylacetamide deacetylase-like 1 (AADACL1), a lipid deacetylase originally identified in invasive cancers. Using this compound, along with highly selective second-generation inhibitors of AADACL1, metabolomics and RNA interference, we show that AADACL1 regulates platelet aggregation, thrombus growth, RAP1 and PKC activation, lipid metabolism and fibrinogen binding to platelets and megakaryocytes. These data provide the first evidence that AADACL1 regulates platelet and megakaryocyte activation and highlight the value of this chemoproteomic strategy for target discovery in platelets

Long-term land-cover/use change in a traditional farming landscape in Romania inferred from pollen data, historical maps and satellite images

Traditional farming landscapes in the temperate zone that have persisted for millennia can be exceptionally species-rich and are therefore key conservation targets. In contrast to Europe’s West, Eastern Europe harbours widespread traditional farming landscapes, but drastic socio-economic and political changes in the twentieth century are likely to have impacted these landscapes profoundly. We reconstructed long-term land-use/cover and biodiversity changes over the last 150 years in a traditional farming landscape of outstanding species diversity in Transylvania. We used the Regional Estimates of Vegetation Abundance from Large Sites model applied to a pollen record from the Transylvanian Plain and a suite of historical and satellite-based maps. We documented widespread changes in the extent and location of grassland and cropland, a loss of wood pastures as well as a gradual increase in forest extent. Land management in the socialist period (1947–1989) led to grassland expansion, but grassland diversity decreased due to intensive production. Land-use intensity has declined since the collapse of socialism in 1989, resulting in widespread cropland abandonment and conversion to grassland. However, these trends may be temporary due to both ongoing woody encroachment as well as grassland management intensification in productive areas. Remarkably, only 8% of all grasslands existed throughout the entire time period (1860–2010), highlighting the importance of land-use history when identifying target areas for conservation, given that old-growth grasslands are most valuable in terms of biodiversity. Combining datasets from different disciplines can yield important additional insights into dynamic landscape and biodiversity changes, informing conservation actions to maintain these species-rich landscapes in the longer term

Dimensions of invasiveness: Links between local abundance, geographic range size, and habitat breadth in Europe's alien and native floras

Understanding drivers of success for alien species can inform on potential future invasions. Recent conceptual advances highlight that species may achieve invasiveness via performance along at least three distinct dimensions: 1) local abundance, 2) geographic range size, and 3) habitat breadth in naturalized distributions. Associations among these dimensions and the factors that determine success in each have yet to be assessed at large geographic scales. Here, we combine data from over one million vegetation plots covering the extent of Europe and its habitat diversity with databases on species' distributions, traits, and historical origins to provide a comprehensive assessment of invasiveness dimensions for the European alien seed plant flora. Invasiveness dimensions are linked in alien distributions, leading to a continuum from overall poor invaders to super invaders - abundant, widespread aliens that invade diverse habitats. This pattern echoes relationships among analogous dimensions measured for native European species. Success along invasiveness dimensions was associated with details of alien species' introduction histories: earlier introduction dates were positively associated with all three dimensions, and consistent with theory-based expectations, species originating from other continents, particularly acquisitive growth strategists, were among the most successful invaders in Europe. Despite general correlations among invasiveness dimensions, we identified habitats and traits associated with atypical patterns of success in only one or two dimensions - for example, the role of disturbed habitats in facilitating widespread specialists. We conclude that considering invasiveness within a multidimensional framework can provide insights into invasion processes while also informing general understanding of the dynamics of species distributions.Deutsche Forschungsgemeinschaft (264740629) Grantová Agentura České Republiky (19-28491X) Grantová Agentura České Republiky (19-28807X) Grantová Agentura České Republiky (RVO 67985939) Austrian Science Fund (I 2086 - B29) Bundesministerium für Bildung und Forschung (01LC1807A) Eusko Jaurlaritza (IT299-10) National Research Foundation of Korea (2018R1C1B6005351) University of Latvia (AAp2016/B041//Zd2016/AZ03) Villum Fonden (16549