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Multivalent flexible nanogels exhibit broad-spectrum antiviral activity by blocking virus entry
(2018)
The entry process of viruses into host cells is complex and involves stable but transient multivalent interactions with different cell surface receptors. The initial contact of several viruses begins with attachment to heparan sulfate (HS) proteoglycans on the cell surface, which results in a cascade of events that end up with virus entry. The development of antiviral agents based on multivalent interactions to shield virus particles and block initial interactions with cellular receptors has attracted attention in antiviral research. Here, we designed nanogels with different degrees of flexibility based on dendritic polyglycerol sulfate to mimic cellular HS. The designed nanogels are nontoxic and broad-spectrum, can multivalently interact with viral glycoproteins, shield virus surfaces, and efficiently block infection. We also visualized virus-nanogel interactions as well as the uptake of nanogels by the cells through clathrin-mediated endocytosis using confocal microscopy. As many human viruses attach to the cells through HS moieties, we introduce our flexible nanogels as robust inhibitors for these viruses.
The radula is the central foraging organ and apomorphy of the Mollusca. However, in contrast to other innovations, including the mollusk shell, genetic underpinnings of radula formation remain virtually unknown. Here, we present the first radula formative tissue transcriptome using the viviparous freshwater snail Tylomelania sarasinorum and compare it to foot tissue and the shell-building mantle of the same species. We combine differential expression, functional enrichment, and phylostratigraphic analyses to identify both specific and shared genetic underpinnings of the three tissues as well as their dominant functions and evolutionary origins. Gene expression of radula formative tissue is very distinct, but nevertheless more similar to mantle than to foot. Generally, the genetic bases of both radula and shell formation were shaped by novel orchestration of preexisting genes and continuous evolution of novel genes. A significantly increased proportion of radula-specific genes originated since the origin of stem-mollusks, indicating that novel genes were especially important for radula evolution. Genes with radula-specific expression in our study are frequently also expressed during the formation of other lophotrochozoan hard structures, like chaetae (hes1, arx), spicules (gbx), and shells of mollusks (gbx, heph) and brachiopods (heph), suggesting gene co-option for hard structure formation. Finally, a Lophotrochozoa-specific chitin synthase with a myosin motor domain (CS-MD), which is expressed during mollusk and brachiopod shell formation, had radula-specific expression in our study. CS-MD potentially facilitated the construction of complex chitinous structures and points at the potential of molecular novelties to promote the evolution of different morphological innovations.
Flowers represent a key innovation during plant evolution. Driven by reproductive optimization, evolution of flower morphology has been central in boosting species diversification. In most cases, this has happened through specialized interactions with animal pollinators and subsequent reduction of gene flow between specialized morphs. While radiation has led to an enormous variability in flower forms and sizes, recurrent evolutionary patterns can be observed. Here, we discuss the targets of selection involved in major trends of pollinator-driven flower evolution. We review recent findings on their adaptive values, developmental grounds and genetic bases, in an attempt to better understand the repeated nature of pollinator-driven flower evolution. This analysis highlights how structural innovation can provide flexibility in phenotypic evolution, adaptation and speciation. (C) 2017 Elsevier Ltd. All rights reserved.
EARLY STARVATION1 specifically affects the phosphorylation action of starch-related dikinases
(2018)
Starch phosphorylation by starch-related dikinases glucan, water dikinase (GWD) and phosphoglucan, water dikinase (PWD) is a key step in starch degradation. Little information is known about the precise structure of the glucan substrate utilized by the dikinases and about the mechanisms by which these structures may be influenced. A 50-kDa starch-binding protein named EARLY STARVATION1 (ESV1) was analyzed regarding its impact on starch phosphorylation. In various invitro assays, the influences of the recombinant protein ESV1 on the actions of GWD and PWD on the surfaces of native starch granules were analyzed. In addition, we included starches from various sources as well as truncated forms of GWD. ESV1 preferentially binds to highly ordered, -glucans, such as starch and crystalline maltodextrins. Furthermore, ESV1 specifically influences the action of GWD and PWD at the starch granule surface. Starch phosphorylation by GWD is decreased in the presence of ESV1, whereas the action of PWD increases in the presence of ESV1. The unique alterations observed in starch phosphorylation by the two dikinases are discussed in regard to altered glucan structures at the starch granule surface.
Mycosporine-like amino acids (MAAs) are UV-absorbing metabolites found in cyanobacteria. While their protective role from UV in Microcystis has been studied in a laboratory setting, a full understanding of the ecology of MAA-producing versus non-MAA-producing Microcystis in natural environments is lacking. This study presents a new tool for quantifying MAA-producing Microcystis and applies it to obtain insight into the dynamics of MAA-producing and non-MAA-producing Microcystis in Lake Erie. This study first developed a sensitive, specific TaqMan real-time PCR assay that targets MAA synthetase gene C (mysC) of Microcystis (quantitative range: 1.7 × 101 to 1.7 × 107 copies/assay). Using this assay, Microcystis was quantified with a MAA-producing genotype (mysC+) in water samples (n = 96) collected during March-November 2013 from 21 Lake Erie sites (undetectable − 8.4 × 106 copies/ml). The mysC+ genotype comprised 0.3–37.8% of the Microcystis population in Lake Erie during the study period. The proportion of the mysC+ genotype during high solar UV irradiation periods (mean = 18.8%) was significantly higher than that during lower UV periods (mean = 9.7%). Among the MAAs, shinorine (major) and porphyra (minor) were detected with HPLC-PDA-MS/MS from the Microcystis isolates and water samples. However, no significant difference in the MAA concentrations existed between higher and lower solar UV periods when the MAA concentrations were normalized with Microcystis mysC abundance. Collectively, this study’s findings suggest that the MAA-producing Microcystis are present in Lake Erie, and they may be ecologically advantageous under high UV conditions, but not to the point that they exclusively predominate over the non-MAA-producers.
Location Eurasia. Methods We generated the largest molecular dataset to date of Z. vivipara, ran phylogenetic analyses, reconstructed its evolutionary history, determined the location of glacial refuges and reconstructed ancestral biogeographic regions. Results The phylogenetic analyses revealed a complex evolutionary history, driven by expansions and contractions of the distribution due to glacials and interglacials, and the colonization of new biogeographic regions by all lineages of Z. vivipara. Many glacial refugia were detected, most were located close to the southern limit of the Last Glacial Maximum. Two subclades recolonized large areas covered by permafrost during the last glaciation: namely, Western and Northern Europe and North-Eastern Europe and Asia.
QuestionDisturbed areas offer great opportunities for restoring native biodiversity, but they are also prone to invasion by alien plants. Following the limiting similarity hypothesis, we address the question of whether or not similarity of plant functional traits helps developing seed mixtures of native communities with high resistance to invasive species at an early stage of restoration. LocationCentre of Greenhouses and Laboratories Durnast, Technische Universitat Munchen, Freising, Germany. MethodsUsing a system of linear equations, we designed native communities maximizing the similarity between the native and two invasive species according to ten functional traits. We used native grassland plants, two invasive alien species that are often problematic in disturbed areas (i.e., Ambrosia artemisiifolia and Solidago gigantea) and trait information obtained from databases. The two communities were then tested for resistance against establishment of the two invaders separately in a greenhouse experiment. We measured height of the invasive species and above-ground biomass, along with leaf area index, 4 and 8months after sowing respectively. ResultsBoth invasive species were successfully reduced by the native community designed to suppress S. gigantea dominated by small-seeded species. These results could be considered as partial support for the limiting similarity hypothesis. However, given the success of this mixture against both invasive species, suppression was better explained by a seed density effect resulting from the smaller seed mass of the native species included in this mixture. Further, the dominance of a fast-developing competitive species could also contribute to its success. ConclusionsThere was no unequivocal support for the limiting similarity hypothesis in terms of the traits selected. Instead we found that increasing seeding density of native species and selecting species with a fast vegetative development is an effective way to suppress invasive plants during early stages of restoration. If limiting similarity is used to design communities for restoration, early life-history traits should be taken into account.
Structural and functional insights into the unique CBS-CP12 fusion protein family in cyanobacteria
(2018)
Cyanobacteria are important photosynthetic organisms inhabiting a range of dynamic environments. This phylum is distinctive among photosynthetic organisms in containing genes encoding uncharacterized cystathionine beta-synthase (CBS)-chloroplast protein (CP12) fusion proteins. These consist of two domains, each recognized as stand-alone photosynthetic regulators with different functions described in cyanobacteria (CP12) and plants (CP12 and CBSX). Here we show that CBS-CP12 fusion proteins are encoded in distinct gene neighborhoods, several unrelated to photosynthesis. Most frequently, CBS-CP12 genes are in a gene cluster with thioredoxin A (TrxA), which is prevalent in bloom-forming, marine symbiotic, and benthic mat cyanobacteria. Focusing on a CBS-CP12 from Microcystis aeruginosa PCC 7806 encoded in a gene cluster with TrxA, we reveal that the domain fusion led to the formation of a hexameric protein. We show that the CP12 domain is essential for hexamerization and contains an ordered, previously structurally uncharacterized N-terminal region. We provide evidence that CBS-CP12, while combining properties of both regulatory domains, behaves different from CP12 and plant CBSX. It does not form a ternary complex with phosphoribulokinase (PRK) and glyceraldehyde-3-phosphate dehydrogenase. Instead, CBS-CP12 decreases the activity of PRK in an AMP-dependent manner. We propose that the novel domain architecture and oligomeric state of CBS-CP12 expand its regulatory function beyond those of CP12 in cyanobacteria.
1. Savanna systems exhibit a high plant functional diversity. While aridity and livestock grazing intensity have been widely discussed as drivers of savanna vegetation composition, physical soil properties have received less attention. Since savannas can show local differences in soil properties, these might act as environmental filters and affect plant diversity and ecosystem functioning at the patch scale. However, research on the link between savanna vegetation diversity and ecosystem function is widely missing. 2. In this study, we aim at understanding the impact of local heterogeneity in soil conditions on plant diversity and on ecosystem functions. For this, we used the ecohydrological savanna model EcoHyD. The model simulates the fate of multiple plant functional types and their interactions with local biotic and abiotic conditions. We applied the model to a set of different landscapes under a wide range of livestock grazing and precipitation scenarios to assess the impact of local heterogeneity in soil conditions on the composition and diversity of plant functional types and on ecosystem functions. 3. Comparisons between homogeneous and heterogeneous landscapes revealed that landscape soil heterogeneity allowed for a higher functional diversity of vegetation under conditions of high competition, i.e. scenarios of low grazing stress. However, landscape heterogeneity did not have this effect under low grazing stress in combination with high mean annual precipitation. Further, landscape heterogeneity led to a higher community biomass, especially for lower rainfall conditions, but also dependent on grazing stress. Total transpiration of the plant community decreased in heterogeneous landscapes under arid conditions. 4. This study highlights that local soil conditions interact with precipitation and grazing in driving savanna vegetation. It clearly shows that vegetation diversity and resulting ecosystem functioning can be driven by landscape heterogeneity. We therefore suggest that future research on ecosystem functioning of savanna systems should focus on the links between local environmental conditions via plant functional diversity to ecosystem functioning.
Ice-wedge polygons are common features of northeastern Siberian lowland periglacial tundra landscapes. To deduce the formation and alternation of ice-wedge polygons in the Kolyma Delta and in the Indigirka Lowland, we studied shallow cores, up to 1.3 m deep, from polygon center and rim locations. The formation of well-developed low-center polygons with elevated rims and wet centers is shown by the beginning of peat accumulation, increased organic matter contents, and changes in vegetation cover from Poaceae-, Alnus-, and Betula-dominated pollen spectra to dominating Cyperaceae and Botryoccocus presence, and Carex and Drepanocladus revolvens macro-fossils. Tecamoebae data support such a change from wetland to open-water conditions in polygon centers by changes from dominating eurybiontic and sphagnobiontic to hydrobiontic species assemblages. The peat accumulation indicates low-center polygon formation and started between 2380 +/- 30 and 1676 +/- 32 years before present (BP) in the Kolyma Delta. We recorded an opposite change from open-water to wetland conditions because of rim degradation and consecutive high-center polygon formation in the Indigirka Lowland between 2144 +/- 33 and 1632 +/- 32 years BP. The late Holocene records of polygon landscape development reveal changes in local hydrology and soil moisture.
Fluorescence fluctuation spectroscopy has become a popular toolbox for non-disruptive analysis of molecular interactions in living cells. The quantification of protein oligomerization in the native cellular environment is highly relevant for a detailed understanding of complex biological processes. An important parameter in this context is the molecular brightness, which serves as a direct measure of oligomerization and can be easily extracted from temporal or spatial fluorescence fluctuations. However, fluorescent proteins (FPs) typically used in such studies suffer from complex photophysical transitions and limited maturation, inducing non-fluorescent states. Here, we show how these processes strongly affect molecular brightness measurements. We perform a systematic characterization of non-fluorescent states for commonly used FPs and provide a simple guideline for accurate, unbiased oligomerization measurements in living cells. Further, we focus on novel red FPs and demonstrate that mCherry2, an mCherry variant, possesses superior properties with regards to precise quantification of oligomerization.
Endothelial cells (ECs) are involved in a variety of cellular responses. As multifunctional components of vascular structures, endothelial (progenitor) cells have been utilized in cellular therapies and are required as an important cellular component of engineered tissue constructs and in vitro disease models. Although primary ECs from different sources are readily isolated and expanded, cell quantity and quality in terms of functionality and karyotype stability is limited. ECs derived from human induced pluripotent stem cells (hiPSCs) represent an alternative and potentially superior cell source, but traditional culture approaches and 2D differentiation protocols hardly allow for production of large cell numbers. Aiming at the production of ECs, we have developed a robust approach for efficient endothelial differentiation of hiPSCs in scalable suspension culture. The established protocol results in relevant numbers of ECs for regenerative approaches and industrial applications that show in vitro proliferation capacity and a high degree of chromosomal stability.
Cardiomyocyte proliferation is crucial for cardiac growth, patterning and regeneration; however, few studies have investigated the behavior of dividing cardiomyocytes in vivo. Here, we use time-lapse imaging of beating hearts in combination with the FUCCI system to monitor the behavior of proliferating cardiomyocytes in developing zebrafish. Confirming in vitro observations, sarcomere disassembly, as well as changes in cell shape and volume, precede cardiomyocyte cytokinesis. Notably, cardiomyocytes in zebrafish embryos and young larvae mostly divide parallel to the myocardial wall in both the compact and trabecular layers, and cardiomyocyte proliferation is more frequent in the trabecular layer. While analyzing known regulators of cardiomyocyte proliferation, we observed that the Nrg/ErbB2 and TGF beta signaling pathways differentially affect compact and trabecular layer cardiomyocytes, indicating that distinct mechanisms drive proliferation in these two layers. In summary, our data indicate that, in zebrafish, cardiomyocyte proliferation is essential for trabecular growth, but not initiation, and set the stage to further investigate the cellular and molecular mechanisms driving cardiomyocyte proliferation in vivo.
Pharmaceuticals are found in freshwater ecosystems where even low concentrations in the range of ng L−1 may affect aquatic organisms. In the current study, we investigated the effects of chronic exposure to three pharmaceuticals on two microalgae, a potential modulation of the effects by additional inorganic phosphorus (Pi) limitation, and a potential propagation of the pharmaceuticals’ effect across a trophic interaction. The latter considers that pharmaceuticals are bioaccumulated by algae, potentially metabolized into more (or less) toxic derivates and consequently consumed by zooplankton. We cultured Acutodesmus obliquus and Nannochloropsis limnetica in Pi-replete and Pi-limited medium contaminated with one of three commonly human used pharmaceuticals: fluoxetine, ibuprofen, and propranolol. Secondly, we tested to what extent first level consumers (Daphnia magna) were affected when fed with pharmaceutical-grown algae. Chronic exposure, covering 30 generations, led to (i) decreased cell numbers of A. obliquus in the presence of fluoxetine (under Pi-replete conditions) (ii) increased carotenoid to chlorophyll ratios in N. limnetica (under Pi-limited conditions), and (iii) increased photosynthetic yields in A. obliquus (in both Pi-conditions). In addition, ibuprofen affected both algae and their consumer: Feeding ibuprofen-contaminated algae to Pi-stressed D. magna improved their survival. We demonstrate, that even very low concentrations of pharmaceuticals present in freshwater ecosystems can significantly affect aquatic organisms when chronically exposed. Our study indicates that pharmaceutical effects can cross trophic levels and travel up the food chain.
Plants can be primed by a stress cue to mount a faster or stronger activation of defense mechanisms upon subsequent stress. A crucial component of such stress priming is the modified reactivation of genes upon recurring stress; however, the underlying mechanisms of this are poorly understood. Here, we report that dozens of Arabidopsis thaliana genes display transcriptional memory, i.e. stronger upregulation after a recurring heat stress, that lasts for at least 3 days. We define a set of transcription factors involved in this memory response and show that the transcriptional memory results in enhanced transcriptional activation within minutes of the onset of a heat stress cue. Further, we show that the transcriptional memory is active in all tissues. It may last for up to a week, and is associated during this time with histone H3 lysine 4 hypermethylation. This transcriptional memory is cis-encoded, as we identify a promoter fragment that confers memory onto a heterologous gene. In summary, heat-induced transcriptional memory is a widespread and sustained response, and our study provides a framework for future mechanistic studies of somatic stress memory in higher plants.
Calcineurin is involved in development and cell differentiation of the social amoeba Dictyostelium discoideum. However, since knockouts of the calcineurin-encoding genes are not possible in D. discoideum it is assumed that the phosphatase also plays a crucial role during vegetative growth of the amoebae. Therefore, we investigated the role of calcineurin during vegetative growth in D. discoideum. RNAi-silenced calcineurin mutants showed cellular alterations with an abnormal morphology of mitochondria and had increased content of mitochondrial DNA (mtDNA). In contrast, mitochondria showed no substantial functional impairment. Calcineurin-silencing led to altered expression of calcium-regulated genes as well as mitochondrially-encoded genes. Furthermore, genes related to oxidative stress were higher expressed in the mutants, which correlated to an increased resistance towards reactive oxygen species (ROS). Most of the changes observed during vegetative growth were not seen after starvation of the calcineurin mutants. We show that impairment of calcineurin led to many subtle, but in the sum crucial cellular alterations in vegetative D. discoideum cells. As these alterations were not observed after starvation we propose a dual role for calcineurin during growth and development. Our results imply that calcineurin is one player in the mutual interplay between mitochondria and ROS during vegetative growth.
Caffeine protects against anticonvulsant-induced impaired neurogenesis in the developing rat brain
(2018)
In preterm infants, phenobarbital is the first-line antiepileptic drug for neonatal seizures while caffeine is used for the treatment of apnea. Data from experimental animals suggest that phenobarbital and other anticonvulsants are toxic for the developing brain, while neuroprotective effects have been reported for caffeine both in newborn rodents and preterm human infants. To characterize the interaction of phenobarbital and caffeine in the hippocampus of the developing rodent brain, we examined the effects of both drugs given separately or together on postnatal neurogenesis after administration to neonatal rats throughout postnatal day (P) 4 to P6. Phenobarbital treatment (50 mg/kg) resulted in a significant decrease of proliferative capacity in the dentate gyrus. Phenobarbital also reduced expression of neuronal markers (doublecortin (DCX), calretinin, NeuN), neuronal transcription factors (Pax6, Sox2, Tbr1/2, Prox1), and neurotrophins (NGF, BDNF, NT-3) up to 24 h after the last administration. The phenobarbital-mediated impairment of neurogenesis was largely ameliorated by preconditioning with caffeine (10 mg/kg). In contrast, caffeine alone reduced proliferative capacity and expression of the neuronal markers DCX and NeuN at 6 h, but increased expression of neurotrophins and neuronal transcription factors at 6 and 12 h. These results indicate that administration of phenobarbital during the vulnerable phase of brain development negatively interferes with neuronal development, which can be prevented in part by co-administration of caffeine.
Non-vascular vegetation has been shown to capture considerable quantities of rainfall, which may affect the hydrological cycle and climate at continental scales. However, direct measurements of rainfall interception by non-vascular vegetation are confined to the local scale, which makes extrapolation to the global effects difficult. Here we use a process-based numerical simulation model to show that non-vascular vegetation contributes substantially to global rainfall interception. Inferred average global water storage capacity including non-vascular vegetation was 2.7 mm, which is consistent with field observations and markedly exceeds the values used in land surface models, which average around 0.4 mm. Consequently, we find that the total evaporation of free water from the forest canopy and soil surface increases by 61% when non-vascular vegetation is included, resulting in a global rainfall interception flux that is 22% of the terrestrial evaporative flux (compared with only 12% for simulations where interception excludes non-vascular vegetation). We thus conclude that non-vascular vegetation is likely to significantly influence global rainfall interception and evaporation with consequences for regional-to continental-scale hydrologic cycling and climate.
Laboratory mice are the most commonly used animal model for Staphylococcus aureus infection studies. We have previously shown that laboratory mice from global vendors are frequently colonized with S. aureus. Laboratory mice originate from wild house mice. Hence, we investigated whether wild rodents, including house mice, as well as shrews are naturally colonized with S. aureus and whether S. aureus adapts to the wild animal host. 295 animals of ten different species were caught in different locations over four years (2012-2015) in Germany, France and the Czech Republic. 45 animals were positive for S. aureus (15.3%). Three animals were co-colonized with two different isolates, resulting in 48 S. aureus isolates in total. Positive animals were found in Germany and the Czech Republic in each studied year. The S. aureus isolates belonged to ten different spa types, which grouped into six lineages (clonal complex (CC) 49, CC88, CC130, CC1956, sequence type (ST) 890, ST3033). CC49 isolates were most abundant (17/48, 35.4%), followed by CC1956 (14/48, 29.2%) and ST890 (9/48, 18.8%). The wild animal isolates lacked certain properties that are common among human isolates, e.g., a phage-encoded immune evasion cluster, superantigen genes on mobile genetic elements and antibiotic resistance genes, which suggests long-term adaptation to the wild animal host. One CC130 isolate contained the mecC gene, implying wild rodents might be both reservoir and vector for methicillin-resistant. In conclusion, we demonstrated that wild rodents and shrews are naturally colonized with S. aureus, and that those S. aureus isolates show signs of host adaptation.
Cobras are among the most widely known venomous snakes, and yet their taxonomy remains incompletely understood, particularly in Africa. Here, we use a combination of mitochondrial and nuclear gene sequences and morphological data to diagnose species limits within the African forest cobra, Naja (Boulengerina) melanoleuca. Mitochondrial DNA sequences reveal deep divergences within this taxon. Congruent patterns of variation in mtDNA, nuclear genes and morphology support the recognition of five separate species, confirming the species status of N. subfulva and N. peroescobari, and revealing two previously unnamed West African species, which are described as new: Naja (Boulengerina) guineensis sp. nov. Broadley, Trape, Chirio, Ineich & Wuster, from the Upper Guinea forest of West Africa, and Naja (Boulengerina) savannula sp. nov. Broadley, Trape, Chirio & Wuster, a banded form from the savanna-forest mosaic of the Guinea and Sudanian savannas of West Africa. The discovery of cryptic diversity in this iconic group highlights our limited understanding of tropical African biodiversity, hindering our ability to conserve it effectively.