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The live sciences currently undergo a paradigm shift to computer aided discoveries. Discoveries in the live sciences were historically made by either direct observation or as a result of chemical assays. Today we see a growing shift toward computer aided analysis and visualization. This gradual process happens in microscopy. Multidimensional laser scanning microscopy can acquire very complex multichannel data from fixed or live specimen. New probes such as visible fluorescent proteins let us observe the expression of genes and track protein localization. Ion sensitive dyes change intensity with the concentration of ions in the cell. The laser scanning confocal allows us to record these processes in three dimensions over time. This work demonstrates the application of software analysis to multidimensional microscopy data. We introduce methods for volume investigation, ion flux analysis and molecular modeling. The visualization methods are based on a multidimensional data model to accommodate complex datasets. The software uses vector processing and multiple processors to accelerate volume rendering and achieve interactive rendering. The algorithms are based on human visual perception and allow the observer a wide range of mixed render modes. The software was used to reconstruct the pituitary development in zebrafish and observe the degeneration of neurons after injury in a mouse model. Calicum indicator dyes have long been used to study calcium fluxes. We optimized the imaging method to minimize impact on the cell. Live cells were imaged continuously for 45 minutes and subjected to increasing does of a drug. We correlated the amplitude of calcium oscillations to increasing doses of a drug and obtain single cell dose response curves. Because this method is very sensitive and measures single cell responses it has potential in drug discovery and characterization. Microtubules form a dynamic cytoskeleton, which is responsible for cell shape, intracellular transport and has an integral role in mitosis. A hallmark of microtubule organization is lateral interactions. Microtubules are bundles by proteins into dense structures. To estimate the contribution of this bundling process, we created a fractal model of microtubule organization. This model demonstrates that morphology of complex microtubule arrays can be explained by bundling alone. In summary we showed that advances in software for visualization, data analysis and modeling lead to new discoveries.
The phase space for the standard model of the basic four forces for n quanta includes all possible ensemble combinations of their quantum states m, a total of n**m states. Neighbor states reach according to transition possibilities (S-matrix) with emergent time from entropic ensemble gradients.
We replace the “big bang” by a condensation event (interacting qubits become decoherent) and inflation by a crystallization event – the crystal unit cell guarantees same symmetries everywhere. Interacting qubits solidify and form a rapidly growing domain where the n**m states become separated ensemble states, rising long-range forces stop ultimately further growth. After that very early events, standard cosmology with the hot fireball model takes over. Our theory agrees well with lack of inflation traces in cosmic background measurements, large-scale structure of voids and filaments, supercluster formation, galaxy formation, dominance of matter and life-friendliness.
We prove qubit interactions to be 1,2,4 or 8 dimensional (agrees with E8 symmetry of our universe). Repulsive forces at ultrashort distances result from quantization, long-range forces limit crystal growth. Crystals come and go in the qubit ocean. This selects for the ability to lay seeds for new crystals, for self-organization and life-friendliness.
We give energy estimates for free qubits vs bound qubits, misplacements in the qubit crystal and entropy increase during qubit decoherence / crystal formation. Scalar fields for color interaction and gravity derive from the permeating qubit-interaction field. Hence, vacuum energy gets low only inside the qubit crystal. Condensed mathematics may advantageously model free / bound qubits in phase space.
Background: Patterns that arise from an ecological process can be driven as much from the landscape over which the process is run as it is by some intrinsic properties of the process itself. The disentanglement of these effects is aided if it possible to run models of the process over artificial landscapes with controllable spatial properties. A number of different methods for the generation of so-called ‘neutral landscapes’ have been developed to provide just such a tool. Of these methods, a particular class that simulate fractional Brownian motion have shown particular promise. The existing methods of simulating fractional Brownian motion suffer from a number of problems however: they are often not easily generalisable to an arbitrary number of dimensions and produce outputs that can exhibit some undesirable artefacts. Methodology: We describe here an updated algorithm for the generation of neutral landscapes by fractional Brownian motion that do not display such undesirable properties. Using Monte Carlo simulation we assess the anisotropic properties of landscapes generated using the new algorithm described in this paper and compare it against a popular benchmark algorithm. Conclusion/Significance: The results show that the existing algorithm creates landscapes with values strongly correlated in the diagonal direction and that the new algorithm presented here corrects this artefact. A number of extensions of the algorithm described here are also highlighted: we describe how the algorithm can be employed to generate landscapes that display different properties in different dimensions and how they can be combined with an environmental gradient to produce landscapes that combine environmental variation at the local and macro scales.
Melanotic melanoma (MM) of Xiphophorus (Teleostei: Poeciliidae) was studied by conventional preparations and freeze-etch preparations for electron microscopy. MM of Xiphophorus exhibits tightly packed pigment cells with prominent dendritic processes and interdigitations of their plasma membranes. The most impressive feature of MM cells is the occurrence of Iarge lobulated nuclei with numerous nuclear pores and some nuclear pockets. Abundant spheroidal or ellipsoidal melanosomes (diameter 200-650 nm) and vesicular structures are distributed throughout the cellular dendrites, whereas the perinucJear cytoplasm is free of melanosomes.
A further characteristic feature of melanoma cells in fish is the occurrence of melanosome complexes (i.e., "compound melanosomes"). These melanosome complexes consist of a few to numerous melanosomes, which are enveloped by a separate rnembrane. Pinocytotic vesicles couJd be demonstrated with distinct differences in frequency and distribution patterns, indicating differences in the metabolic activities of the cells in the same melanoma. Intercellular junctions are lacking in the MM cells.
The conventional TEM technique showed clear advantages in the demonstration of intemal architecture of organelles, whereas FE bad considerable potential in respect to the visualization of membrane surface specializations.
An ordered NotI fragment map containing over 60 loci and encompassing approximately 17 Mb has been constructed for human chromosome band llpl5. Forty-two probes, including 11 NotI-linking cosmids, were subregionaUy mapped to llpl5 using a subset of the Jl-deletion hybrids. These and 23 other probes defining loci previously mapped to 11p15 were hybridized to genomic DNA digested with NotI and 5 other infrequently cleaving restriction enzymes and separated by pulsed-field gel electrophoresis. Thirty-nine distinct NotI fragments were detected encompassing approximately 85% of the estimated length of llp15. The predicted order of the gene loci used is cenMYODI- PTH-CALCA-ST5-RBTNI-HPX-HBB-RRMlTH/ INS!1GF2-H19-CTSD-MUC2-DRD4-HRAS-RNHtel. This map wiu allow higher resolution mapping of new Ilp15 markers, facilitate positional cloning of disease genes, and provide a framework for the physical mapping of llp15 in clone contigs.
Whereas the role of calcium ions (Ca\(^{2+}\)) in plant signaling is well studied, the physiological significance of pH‐changes remains largely undefined.
Here we developed CapHensor, an optimized dual‐reporter for simultaneous Ca\(^{2+}\) and pH ratio‐imaging and studied signaling events in pollen tubes (PTs), guard cells (GCs), and mesophyll cells (MCs). Monitoring spatio‐temporal relationships between membrane voltage, Ca\(^{2+}\)‐ and pH‐dynamics revealed interconnections previously not described.
In tobacco PTs, we demonstrated Ca\(^{2+}\)‐dynamics lag behind pH‐dynamics during oscillatory growth, and pH correlates more with growth than Ca\(^{2+}\). In GCs, we demonstrated abscisic acid (ABA) to initiate stomatal closure via rapid cytosolic alkalization followed by Ca2+ elevation. Preventing the alkalization blocked GC ABA‐responses and even opened stomata in the presence of ABA, disclosing an important pH‐dependent GC signaling node. In MCs, a flg22‐induced membrane depolarization preceded Ca2+‐increases and cytosolic acidification by c. 2 min, suggesting a Ca\(^{2+}\)/pH‐independent early pathogen signaling step. Imaging Ca2+ and pH resolved similar cytosol and nuclear signals and demonstrated flg22, but not ABA and hydrogen peroxide to initiate rapid membrane voltage‐, Ca\(^{2+}\)‐ and pH‐responses.
We propose close interrelation in Ca\(^{2+}\)‐ and pH‐signaling that is cell type‐ and stimulus‐specific and the pH having crucial roles in regulating PT growth and stomata movement.
Societal Impact Statement
Pollen relates to many aspects of human and environmental health, which protection and improvement are endorsed by the United Nations Sustainable Development Goals. By highlighting these connections in the frame of current challenges in monitoring and research, we discuss the need of more integrative and multidisciplinary pollen research related to societal needs, improving health of humans and our ecosystems for a sustainable future.
Summary
Pollen is at once intimately part of the reproductive cycle of seed plants and simultaneously highly relevant for the environment (pollinators, vector for nutrients, or organisms), people (food safety and health), and climate (cloud condensation nuclei and climate reconstruction). We provide an interdisciplinary perspective on the many and connected roles of pollen to foster a better integration of the currently disparate fields of pollen research, which would benefit from the sharing of general knowledge, technical advancements, or data processing solutions. We propose a more interdisciplinary and holistic research approach that encompasses total environmental pollen diversity (ePD) (wind and animal and occasionally water distributed pollen) at multiple levels of diversity (genotypic, phenotypic, physiological, chemical, and functional) across space and time. This interdisciplinary approach holds the potential to contribute to pressing human issues, including addressing United Nations Sustainable Development Goals, fostering social and political awareness of these tiny yet important and fascinating particles.
Systems biology looks for emergent system effects from large scale assemblies of molecules and data, for instance in the human platelets. However, the computational efforts in all steps before such insights are possible can hardly be under estimated. In practice this involves numerous programming tasks, the establishment of new database systems but as well their maintenance, curation and data validation. Furthermore, network insights are only possible if strong algorithms decipher the interactions, decoding the hidden system effects. This thesis and my work are all about these challenges. To answer this requirement, an integrated platelet network, PlateletWeb, was assembled from different sources and further analyzed for signaling in a systems biological manner including multilevel data integration and visualization. PlateletWeb is an integrated network database and was established by combining the data from recent platelet proteome and transcriptome (SAGE) studies. The information on protein-protein interactions and kinase-substrate relationships extracted from bioinformatical databases as well as published literature were added to this resource. Moreover, the mass spectrometry-based platelet phosphoproteome was combined with site-specific phosphorylation/ dephosphorylation information and then enhanced with data from Phosphosite and complemented by bioinformatical sequence analysis for site-specific kinase predictions. The number of catalogued platelet proteins was increased by over 80% as compared to the previous version. The integration of annotations on kinases, protein domains, transmembrane regions, Gene Ontology, disease associations and drug targets provides ample functional tools for platelet signaling analysis. The PlateletWeb resource provides a novel systems biological workbench for the analysis of platelet signaling in the functional context of protein networks. By comprehensive exploration, over 15000 phosphorylation sites were found, out of which 2500 have the corresponding kinase associations. The network motifs were also investigated in this anucleate cell and characterize signaling modules based on integrated information on phosphorylation and protein-protein interactions. Furthermore, many algorithmic approaches have been introduced, including an exact approach (heinz) based on integer linear programming. At the same time, the concept of semantic similarities between two genes using Gene Ontology (GO) annotations has become an important basis for many analytical approaches in bioinformatics. Assuming that a higher number of semantically similar gene functional annotations reflect biologically more relevant interactions, an edge score was devised for functional network analysis. Bringing these two approaches together, the edge score, based on the GO similarity, and the node score, based on the expression of the proteins in the analyzed cell type (e.g. data from proteomic studies), the functional module as a maximum-scoring sub network in large protein-protein interaction networks was identified. This method was applied to various proteome datasets (different types of blood cells, embryonic stem cells) to identify protein modules that functionally characterize the respective cell type. This scalable method allows a smooth integration of data from various sources and retrieves biologically relevant signaling modules.
The olive tree is a venerable Mediterranean plant and often used in traditional medicine. The main aim of the present study was to evaluate the effect of Olea europaea L. cv. Arbosana leaf extract (OLE) and its encapsulation within a spanlastic dosage form on the improvement of its pro-oxidant and antiproliferative activity against HepG-2, MCF-7, and Caco-2 human cancer cell lines. The LC-HRESIMS-assisted metabolomic profile of OLE putatively annotated 20 major metabolites and showed considerable in vitro antiproliferative activity against HepG-2, MCF-7, and Caco-2 cell lines with IC\(_{50}\) values of 9.2 ± 0.8, 7.1 ± 0.9, and 6.5 ± 0.7 µg/mL, respectively. The encapsulation of OLE within a (spanlastic) nanocarrier system, using a spraying method and Span 40 and Tween 80 (4:1 molar ratio), was successfully carried out (size 41 ± 2.4 nm, zeta potential 13.6 ± 2.5, and EE 61.43 ± 2.03%). OLE showed enhanced thermal stability, and an improved in vitro antiproliferative effect against HepG-2, MCF-7, and Caco-2 (IC\(_{50}\) 3.6 ± 0.2, 2.3 ± 0.1, and 1.8 ± 0.1 µg/mL, respectively) in comparison to the unprocessed extract. Both preparations were found to exhibit pro-oxidant potential inside the cancer cells, through the potential inhibitory activity of OLE against glutathione reductase and superoxide dismutase (IC\(_{50}\) 1.18 ± 0.12 and 2.33 ± 0.19 µg/mL, respectively). These inhibitory activities were proposed via a comprehensive in silico study to be linked to the presence of certain compounds in OLE. Consequently, we assume that formulating such a herbal extract within a suitable nanocarrier would be a promising improvement of its therapeutic potential.
Background
The plant endophytic fungus Serendipita indica colonizes roots of a wide range of plant species and can enhance growth and stress resistance of these plants. Due to its ease of axenic cultivation and its broad host plant range including the model plant Arabidopsis thaliana and numerous crop plants, it is widely used as a model fungus to study beneficial fungus-root interactions. In addition, it was suggested to be utilized for commercial applications, e.g. to enhance yield in barley and other species. To produce inoculum, S. indica is mostly cultivated in a complex Hill-Kafer medium (CM medium), however, growth in this medium is slow, and yield of chlamydospores, which are often used for plant root inoculation, is relatively low.
Results
We tested and optimized a simple vegetable juice-based medium for an enhanced yield of fungal inoculum. The described vegetable juice (VJ) medium is based on commercially available vegetable juice and is easy to prepare. VJ medium was superior to the currently used CM medium with respect to biomass production in liquid medium and hyphal growth on agar plates. Using solid VJ medium supplemented with sucrose (VJS), a high amount of chlamydospores developed already after 8 days of cultivation, producing significantly more spores than on CM medium. Use of VJ medium is not restricted to S. indica, as it also supported growth of two pathogenic fungi often used in plant pathology experiments: the ascomycete Fusarium graminearum, the causal agent of Fusarium head blight disease on wheat and barley, and Verticillium longisporum, the causal agent of verticillium wilt.
Conclusions
The described VJ medium is recommended for streamlined and efficient production of inoculum for the plant endophytic fungus Serendipita indica and might prove superior for the propagation of other fungi for research purposes.