Die Aufklärung der Tertiärstruktur des antimikrobiellen Peptids Hydramacin-1 mit Hilfe der mehrdimensionalen heteronuklearen NMR-Spektroskopie und die Untersuchung seines Wirkmechanismus

Das antimikrobielle Peptid Hydramacin-1 aus dem Süßwasserpolypen Hydra magnipapillata tötet Gram-negative und Gram-positive Bakterien im mikromolaren Konzentrationsbereich. Um den Wirkmechanismus von Hydramacin-1 zu charakterisieren, wurde in der vorliegenden Arbeit seine Tertiärstruktur mit Hilfe der mehrdimensionalen heteronuklearen NMR-Spektroskopie aufgeklärt sowie seine Interaktion mit Liposomen und Bakterien untersucht. Die Tertiärstruktur des Hydramacin-1 zeigt ein Disulfidbrücken-stabilisiertes α/β Strukturmotiv, welches für die sogenannte Knottin-Faltungstopologie charakteristisch ist. Auf Grund seiner Tertiärstruktur gehört Hydramacin-1 zur Superfamilie der Skorpiontoxin-ähnlichen Proteine in der es eine neue Familie, die Macine, begründet. Hydramacin-1 ist in der Lage sowohl Liposomen als auch Bakterien zu präzipitieren. Mit der Kenntnis der Tertiärstruktur von Hydramacin-1 konnte ein Modell formuliert werden, welches diesen Präzipitationseffekt erklärt. Ein vergleichbares Modell wurde bisher noch nicht beschrieben und erweitert somit den Kenntnisstand über die Wirkmechanismen antimikrobieller Peptide

Species-Specific Differences in the Susceptibility of Fungi to the Antifungal Protein AFP Depend on C-3 Saturation of Glycosylceramides

AFP is an antimicrobial peptide (AMP) produced by the filamentous fungus Aspergillus giganteus and is a very potent inhibitor of fungal growth that does not affect the viability of bacteria, plant, or mammalian cells. It targets chitin synthesis and causes plasma membrane permeabilization in many human- and plant-pathogenic fungi, but its exact mode of action is not known. After adoption of the “damage-response framework of microbial pathogenesis” regarding the analysis of interactions between AMPs and microorganisms, we have recently proposed that the cytotoxic capacity of a given AMP depends not only on the presence/absence of its target(s) in the host and the AMP concentration applied but also on other variables, such as microbial survival strategies. We show here using the examples of three filamentous fungi (Aspergillus niger, Aspergillus fumigatus, and Fusarium graminearum) and two yeasts (Saccharomyces cerevisiae and Pichia pastoris) that the important parameters defining the AFP susceptibilities of these fungi are (i) the presence/absence of glycosylceramides, (ii) the presence/absence of Δ3(E) desaturation of the fatty acid chain therein, and (iii) the (dis)ability of these fungi to respond to AFP inhibitory effects with the fortification of their cell walls via increased chitin and β-(1,3)-glucan synthesis. These observations support the idea of the adoption of the damage-response framework to holistically understand the outcome of AFP inhibitory effects.TU Berlin, Open-Access-Mittel - 201

OceanTEA: A Platform for Sharing Oceanographic Data and Analyses

Ocean observation systems, such as Argo floats or the modular ocean laboratory MoLab, produce an increasing amount of time series data. Both, statistical data mining techniques and manual exploration via visualization are necessary for oceanographers to extract scientific knowledge from such vast datasets. Therefore, scientists require a platform to explore and analyze data visually, supporting their collaboration and research. To deliver results and foster the impact of publications, such platform should facilitate automatic and interactive access to research results for scientists, their peers and the public. Our software platform OceanTEA (Oceanographic TimeSeries Exploration and Analysis) supports oceanographers in their research and publication efforts. The platform leverages modern web technology to support the interactive exploration and analysis of high-dimensional datasets. OceanTEA relies on a microservice architecture which can be deployed on desktops and on cloud computing infrastructure

Computational Integrative Models for Cellular Conversion: Application to Cellular Reprogramming and Disease Modeling

The groundbreaking identification of only four transcription factors that are able to induce pluripotency in any somatic cell upon perturbation stimulated the discovery of copious amounts of instructive factors triggering different cellular conversions. Such conversions are highly significant to regenerative medicine with its ultimate goal of replacing or regenerating damaged and lost cells. Precise directed conversion of damaged cells into healthy cells offers the tantalizing prospect of promoting regeneration in situ. In the advent of high-throughput sequencing technologies, the distinct transcriptional and accessible chromatin landscapes of several cell types have been characterized. This characterization provided clear evidences for the existence of cell type specific gene regulatory networks determined by their distinct epigenetic landscapes that control cellular phenotypes. Further, these networks are known to dynamically change during the ectopic expression of genes initiating cellular conversions and stabilize again to represent the desired phenotype. Over the years, several computational approaches have been developed to leverage the large amounts of high-throughput datasets for a systematic prediction of instructive factors that can potentially induce desired cellular conversions. To date, the most promising approaches rely on the reconstruction of gene regulatory networks for a panel of well-studied cell types relying predominantly on transcriptional data alone. Though useful, these methods are not designed for newly identified cell types as their frameworks are restricted only to the panel of cell types originally incorporated. More importantly, these approaches rely majorly on gene expression data and cannot account for the cell type specific regulations modulated by the interplay of the transcriptional and epigenetic landscape. In this thesis, a computational method for reconstructing cell type specific gene regulatory networks is proposed that aims at addressing the aforementioned limitations of current approaches. This method integrates transcriptomics, chromatin accessibility assays and available prior knowledge about gene regulatory interactions for predicting instructive factors that can potentially induce desired cellular conversions. Its application to the prioritization of drugs for reverting pathologic phenotypes and the identification of instructive factors for inducing the cellular conversion of adipocytes into osteoblasts underlines the potential to assist in the discovery of novel therapeutic interventions

Capella: A Space-only High-frequency Radio VLBI Network Formed by a Constellation of Small Satellites

Very long baseline radio interferometry (VLBI) with ground-based observatories is limited by the size of Earth, the geographic distribution of antennas, and the transparency of the atmosphere. In this whitepaper, we present Capella, a tentative design of a space-only VLBI system. Using four small (<500 kg) satellites on two orthogonal polar low-Earth orbits, and single-band heterodyne receivers operating at frequencies around 690 GHz, the interferometer is able to achieve angular resolutions of approximately 7 microarcsec. Within a total observing time of three days, a near-complete uv plane coverage can be reached, with a 1-sigma point source sensitivity as good as about 6~mJy for an instantaneous bandwidth of 1 GHz. The required downlink data rates of >10 Gbps can be reached through near-infrared laser communication; depending on the actual downlink speed, one or multiple ground communication stations are necessary. We note that all key technologies required for the Capella system are already available, some of them off-the-shelf. Data can be correlated using dedicated versions of existing Fourier transform (FX) software correlators; dedicated routines will be needed to handle the effects of orbital motion, including relativistic corrections. With the specifications assumed in this whitepaper, Capella will be able to address a range of science cases, including: photon rings around supermassive black holes; the acceleration and collimation zones of plasma jets emitted from the vicinity of supermassive black holes; the chemical composition of accretion flows into active galactic nuclei through observations of molecular absorption lines; mapping supermassive binary black holes; the magnetic activity of stars; and nova eruptions of symbiotic binary stars - and, like any substantially new observing technique, has the potential for unexpected discoveries.Comment: 18 pages, 2 figures, 1 table. Whitepaper version 1.0. Living document, will be updated when necessar

Smarte Sirenen im Rahmen kommunaler Klimaanpassungskonzepte im Spannungsfeld von Stadtklimatologie und Stadtplanung

Der urbane Raum wird voraussichtlich besonders stark von Klimaveränderungen betroffen sein. Dies gilt es gegenwärtig zu berücksichtigen, um auf zukünftige Herausforderungen entsprechend vorbereitet und angepasst zu sein. Dabei gilt es vielfältige, oftmals widersprüchliche Anforderungen zu bewältigen, die mit unterschiedlichen Aspekten der Planung einhergehen. Mittels einer soliden und vorausschauenden Planung gilt es darauf Rücksicht zu nehmen, dass die Bevölkerung infolge von Stadtentwicklungs-, Stadterneuerungs- und Stadtumbaumaßnahmen in ihrer Wohn- und Lebensqualität nicht negativ beeinflusst wird (z. B. durch die Veränderungen des Lokalklimas, wenn Fußgänger im Sommer unnötigem Hitzestress ausgesetzt werden, da möglicherweise die vormals schattenspendende Vegetation einer Umstrukturierung zum Opfer gefallen ist). Eine der großen Herausforderungen der kommenden Jahre auf der Ebene der örtlichen Planung wird es sein, sich auf wetterbedingte Extreme wie Starkregenereignisse oder Hitzewellen vorzubereiten. Im Rahmen solcher rezenter Extremereignisse zeigt sich, dass die Folgen für die gesellschaftlichen und räumlichen Strukturen mitunter deutlich ausfallen können. Städte sind aber vielfach noch nicht ausreichend auf solche Szenarien vorbereitet. Auch wenn es gegenwärtig bereits eine große Anzahl an Klimaanpassungskonzepten gibt, ist die Klimaanpassung ein Arbeitsfeld, das weiterer Forschung bedarf, beispielweise hinsichtlich der Bewertung der Effektivität geplanter bzw. bereits umgesetzter Maßnahmen. Am Beispiel der knapp 100.000 Einwohner zählenden Stadt Kaiserslautern wird im Rahmen eines Forschungsprojektes ein Klimaanpassungskonzept entwickelt, das aufzeigt, wie die Siedlungs- und Freiraumentwicklung der Stadt klimaoptimiert auf Hitze- und Starkregenereignisse reagieren kann. Ein erster, wichtiger Baustein eines solchen Konzeptes ist eine zielführende stadtklimatologische/ siedlungsökologische Kartierung der Vor-Ort-Situation, um potentielle Risikogebiete für z. B. Starkregenereignisse im Bestand zu identifizieren. Darauf aufbauend sollen strategische Maßnahmenkonzepte für betroffene Quartiere erarbeitet werden. Als neues Konzept der Klimaanpassung soll dabei der Aufbau eines akustischen Warnsystems für die Bevölkerung untersucht und technisch ausgelegt werden. Extremereignisse mit Gefahrenpotential für die Bevölkerung, insbesondere Starkniederschläge, Gewitter und Stürme, erreichen die Städte oft mit sehr kurzer Vorwarnzeit. Nach dem weitgehenden Abbau der Zivilschutzsirenen bieten „smarte“ Sirenensysteme, die dezentral mit verhältnismäßig geringer Schallleistung installiert werden und situationsangepasste Sprachdurchsagen verteilen können, einen neuen Ansatz zur Schadenminimierung. Die Menschen sollen mithilfe eines optimierten Einsatzes von „Smarten Sirenen“ vor dem Eintreffen von Extremereignissen gewarnt werden, und dies unter Berücksichtigung der urbanen Gebäudestruktur, mit dem Ziel die betroffenen Standorte vorab zu informieren, um notwendige Maßnahmen, z. B. die Räumung von Tiefgaragen und Kellern, die Betätigung von Absperrventilen oder das Aufsuchen sicherer Orte, rechtzeitig treffen zu können

Structure–Activity Predictions From Computational Mining of Protein Databases to Assist Modular Design of Antimicrobial Peptides

Antimicrobial peptides (AMPs) are naturally produced by pro- and eukaryotes and are promising alternatives to antibiotics to fight multidrug-resistant microorganisms. However, despite thousands of AMP entries in respective databases, predictions about their structure–activity relationships are still limited. Similarly, common or dissimilar properties of AMPs that have evolved in different taxonomic groups are nearly unknown. We leveraged data entries for 10,987 peptides currently listed in the three antimicrobial peptide databases APD, DRAMP and DBAASP to aid structure–activity predictions. However, this number reduced to 3,828 AMPs that we could use for computational analyses, due to our stringent quality control criteria. The analysis uncovered a strong bias towards AMPs isolated from amphibians (1,391), whereas only 35 AMPs originate from fungi (0.9%), hindering evolutionary analyses on the origin and phylogenetic relationship of AMPs. The majority (62%) of the 3,828 AMPs consists of less than 40 amino acids but with a molecular weight higher than 2.5 kDa, has a net positive charge and shares a hydrophobic character. They are enriched in glycine, lysine and cysteine but are depleted in glutamate, aspartate and methionine when compared with a peptide set of the same size randomly selected from the UniProt database. The AMPs that deviate from this pattern (38%) can be found in different taxonomic groups, in particular in Gram-negative bacteria. Remarkably, the γ-core motif claimed so far as a unifying structural signature in cysteine-stabilised AMPs is absent in nearly 90% of the peptides, questioning its relevance as a prerequisite for antimicrobial activity. The disclosure of AMPs pattern and their variation in producing organism groups extends our knowledge of the structural diversity of AMPs and will assist future peptide screens in unexplored microorganisms. Structural design of peptide antibiotic drugs will benefit using natural AMPs as lead compounds. However, a reliable and statistically balanced database is missing which leads to a large knowledge gap in the AMP field. Thus, thorough evaluation of the available data, mitigation of biases and standardised experimental setups need to be implemented to leverage the full potential of AMPs for drug development programmes in the clinics and agriculture.DFG, 392923329, GRK 2473: Bioaktive Peptide - Innovative Aspekte zur Synthese und Biosynthes

Light-activatable TET-dioxygenases reveal dynamics of 5-Methylcytosine oxidation and transcriptome reorganization

Ten-eleven-translocation (TET) dioxygenases catalyze the oxidation of 5-methylcytosine (5mC), the central epigenetic regulator of mammalian DNA. This activity dy- namically reshapes epigenome and transcriptome by deposit- ing oxidized 5mC derivatives, and initiating active DNA de- methylation. However, studying this dynamic is hampered by the inability to selectively activate individual TETs with tem- poral control in cells. We report activation of TETs in mam- malian cells by incorporation of genetically encoded 4,5- dimethoxy-2-nitrobenzyl-L-serine as transient active site block, and its subsequent deprotection with light. Our ap- proach enables precise insights into the impact of cancer- associated TET2 mutations on the kinetics of TET2 catalysis in vivo, and allows time-resolved monitoring of target gene activation and transcriptome reorganization. This sets a basis for dissecting the order and kinetics of chromatin-associated events triggered by TET catalysis, ranging from DNA de- methylation to chromatin and transcription regulation

Light-Activatable TET-Dioxygenases Reveal Dynamics of 5-Methylcytosine Oxidation and Transcriptome Reorganization

Ten-eleven-translocation (TET) dioxygenases catalyze the oxidation of 5-methylcytosine (5mC), the central epigenetic regulator of mammalian DNA. This activity dynamically reshapes the epigenome and transcriptome by depositing oxidized 5mC derivatives and initiating active DNA demethylation. However, studying this dynamic is hampered by the inability to selectively activate individual TETs with temporal control in cells. We report activation of TETs in mammalian cells by incorporation of genetically encoded 4,5-dimethoxy-2-nitrobenzyl-l-serine as a transient active-site block, and its subsequent deprotection with light. Our approach enables precise insights into the impact of cancer-associated TET2 mutations on the kinetics of TET2 catalysis in vivo, and allows time-resolved monitoring of target gene activation and transcriptome reorganization. This sets a basis for dissecting the order and kinetics of chromatin-associated events triggered by TET catalysis, ranging from DNA demethylation to chromatin and transcription regulation

Capillary leak and endothelial permeability in critically ill patients: a current overview

Capillary leak syndrome (CLS) represents a phenotype of increased fluid extravasation, resulting in intravascular hypovolemia, extravascular edema formation and ultimately hypoperfusion. While endothelial permeability is an evolutionary preserved physiological process needed to sustain life, excessive fluid leak—often caused by systemic inflammation—can have detrimental effects on patients’ outcomes. This article delves into the current understanding of CLS pathophysiology, diagnosis and potential treatments. Systemic inflammation leading to a compromise of endothelial cell interactions through various signaling cues (e.g., the angiopoietin–Tie2 pathway), and shedding of the glycocalyx collectively contribute to the manifestation of CLS. Capillary permeability subsequently leads to the seepage of protein-rich fluid into the interstitial space. Recent insights into the importance of the sub-glycocalyx space and preserving lymphatic flow are highlighted for an in-depth understanding. While no established diagnostic criteria exist and CLS is frequently diagnosed by clinical characteristics only, we highlight more objective serological and (non)-invasive measurements that hint towards a CLS phenotype. While currently available treatment options are limited, we further review understanding of fluid resuscitation and experimental approaches to target endothelial permeability. Despite the improved understanding of CLS pathophysiology, efforts are needed to develop uniform diagnostic criteria, associate clinical consequences to these criteria, and delineate treatment options. Graphical Abstrac