Mms1 is an assistant for regulating G-quadruplex DNA structures

The preservation of genome stability is fundamental for every cell. Genomic integrity is constantly challenged. Among those challenges are also non-canonical nucleic acid structures. In recent years, scientists became aware of the impact of G-quadruplex (G4) structures on genome stability. It has been shown that folded G4-DNA structures cause changes in the cell, such as transcriptional up/down-regulation, replication stalling, or enhanced genome instability. Multiple helicases have been identified to regulate G4 structures and by this preserve genome stability. Interestingly, although these helicases are mostly ubiquitous expressed, they show specificity for G4 regulation in certain cellular processes (e.g., DNA replication). To this date, it is not clear how this process and target specificity of helicases are achieved. Recently, Mms1, an ubiquitin ligase complex protein, was identified as a novel G4-DNA-binding protein that supports genome stability by aiding Pif1 helicase binding to these regions. In this perspective review, we discuss the question if G4-DNA interacting proteins are fundamental for helicase function and specificity at G4-DNA structures

Untersuchung der Wirkungsweise von Dexmedetomidin auf das adulte Rattenhirn

Das Gehirn weist eine hohe Vulnerabilität gegenüber Inflammation und oxidativem Stress auf. Beide Stressoren sind mit einer vermehrten Neurodegeneration in unterschiedlichen Bereichen des Gehirns assoziiert. Für das Medikament Dexmedetomidin (DEX) sind neben antiinflammatorischen und antioxidativen auch neuroprotektive Eigenschaften beschrieben, die komplette Wirkungsweise ist bisher jedoch nicht vollständig geklärt. In der vorliegenden Arbeit galt es, an adulten Wistar Ratten den Einfluss von DEX auf eine durch systemische Applikation von Lipopolysaccharid (LPS) induzierte Neuroinflammation zu untersuchen. Speziell wurden dazu die Zytokine IL1-beta und TNF-alpha sowie die microRNAs (miR) 124, 132, 134 und 155 zu verschiedenen Zeitpunkten im Hippocampus und Cortex mittels Real Time PCR (qPCR) analysiert. Weiterhin fand eine Analyse dieser miRNAs im Plasma statt. Es zeigte sich, dass DEX die durch LPS induzierte Expression von IL1-beta und TNF-alpha signifikant reduziert und die Expression der LPS-induzierten miR 124, 132, 134 und 155 in beiden untersuchten Hirnregionen signifikant verringert. Zusätzlich wiesen miR 132 und 134 im Plasma während der Neuroinflammation ein verändertes Expressionsmuster auf. In einem zweiten Tiermodel wurden nach Schädigung des neonatalen Rattengehirns durch Hyperoxie verschiedene DEX-Konzentrationen bezüglich ihrer möglichen Protektion untersucht. Speziell stand dabei die Begutachtung des apoptotischen Zelltods in unterschiedlichen Hirnarealen im Vordergrund. Zusätzlich wurde oxidativer Stress durch die Analyse von Glutathion und Malondialdehyd (MDA) bestimmt sowie das Zytokin IL1-beta molekularbiologisch analysiert. DEX entfaltete im Modell protektive Effekte durch Verminderung der Neurodegeneration und IL1-beta Expression sowie durch Beeinflussung der untersuchten oxidativen Stressparameter. In einer weiteren Arbeit wurde an adulten Wistar Ratten durch eine Laparotomie sowie nachfolgender LPS-Applikation eine Neuroinflammation induziert und dabei die Wirkung der Acetylcholinesterase-Inhibitoren (AChE-I) Physostigmin (Phy) und Neostigmin (Neo) untersucht. Es wurden verschiedene Zytokine im Gehirn, im Plasma und in der Milz mittels qPCR, Western Blot und Cytometrischem Bead Array (CBA) untersucht sowie der Grad an Neurodegeneration und die Aktivität der Acetylcholinesterase (AChE) im Hippocampus und Cortex bestimmt. Die Applikation von Phy und Neo reduzierte eine LPS-induzierte Neuroinflammation und - degeneration sowie die LPS-vermittelte Erhöhung der AChE-Aktivität. Ferner wurden peripher antiinflammatorische Effekte in der Milz sowie im Plasma detektiert. Zusammenfassend ergibt sich, dass unterschiedliche medikamentöse Interventionen protektive Eigenschaften entfalten, um potentiellen Hyperoxie-Schäden bzw. LPS-induzierten Schäden entgegen zu wirken. Besonders die durch DEX vermittelte Beeinflussung der miRNA Expression eröffnet neue Erkenntnisse, um die komplexe Wirkungsweise dieses Medikaments besser verstehen zu können.The brain is highly vulnerable to inflammation and oxidative stress. Both stressors are associated with increased neurodegeneration in different areas of the brain. The drug dexmedetomidine (DEX) exerts neuroprotective as well as anti-inflammatory and antioxidative properties. However, the complete molecular mechanisms are not fully understood. In the present work, adult Wistar rats were used to investigate the influence of DEX on neuroinflammation induced by the systemic application of lipopolysaccharide (LPS). The cytokines IL1-beta and TNF-alpha, as well as the microRNAs (miR) 124, 132, 134 and 155 were analyzed in the hippocampus and cortex using real-time PCR (qPCR). Furthermore, these miRNAs were studied in plasma. It was found that DEX significantly reduced the LPS-induced expression of IL1-beta and TNF-alpha and significantly decreased the expression of LPS-induced miR 124, 132, 134, and 155 in both brain regions. In addition, miR 132 and 134 in plasma showed an altered expression pattern during neuroinflammation. In a second animal model various DEX concentrations were tested in the neonatal rat brain exposed to hyperoxia. In particular, the focus was on the evaluation of apoptotic cell death in different areas of the brain. Moreover, oxidative stress was determined by the analysis of glutathione and malondialdehyde (MDA) and the cytokine IL1-beta was investigated. DEX showed protective effects in the neonatal brain by reducing hypoxia-induced neurodegeneration and IL1-beta expression. Furthermore, the drug influenced the oxidative stress parameters in a positive way. In a further work, neuroinflammation was induced in adult Wistar rats by laparotomy and subsequent LPS administration to investigate the effect of the acetylcholinesterase-inhibitors (AChE-I) physostigmine (Phy) and neostigmine (Neo). Different cytokines in the brain, plasma, and spleen were investigated using qPCR, western blot, and cytometric bead array (CBA). Furthermore, the degree of neurodegeneration and the activity of acetylcholinesterase (AChE) were evaluated in the hippocampus and cortex. The application of Phy and Neo reduced LPS-induced neuroinflammation and -degeneration as well as the LPS-mediated increase of AChE activity in both investigated brain regions. Peripheral anti-inflammatory effects were also observed in the spleen and plasma. In summary, different drug interventions show protective properties in order to reduce potential hyperoxic- or LPS-induced damage. The influence of DEX on miRNA expression, in particular, opens up new insights into the complex mode of action of this drug

Hrq1, a Homolog of the Human RecQ4 Helicase, Acts Catalytically and Structurally to Promote Genome Integrity

SummaryHuman RecQ4 (hRecQ4) affects cancer and aging but is difficult to study because it is a fusion between a helicase and an essential replication factor. Budding yeast Hrq1 is homologous to the disease-linked helicase domain of RecQ4 and, like hRecQ4, is a robust 3′-5′ helicase. Additionally, Hrq1 has the unusual property of forming heptameric rings. Cells lacking Hrq1 exhibited two DNA damage phenotypes: hypersensitivity to DNA interstrand crosslinks (ICLs) and telomere addition to DNA breaks. Both activities are rare; their coexistence in a single protein is unprecedented. Resistance to ICLs requires helicase activity, but suppression of telomere addition does not. Hrq1 also affects telomere length by a noncatalytic mechanism, as well as telomerase-independent telomere maintenance. Because Hrq1 binds telomeres in vivo, it probably affects them directly. Thus, the tumor-suppressing activity of RecQ4 could be due to a role in ICL repair and/or suppression of de novo telomere addition

Dexmedetomidine Prevents Lipopolysaccharide-Induced MicroRNA Expression in the Adult Rat Brain

During surgery or infection, peripheral inflammation can lead to neuroinflammation, which is associated with cognitive impairment, neurodegeneration, and several neurodegenerative diseases. Dexmedetomidine, an α-2-adrenoceptor agonist, is known to exert anti-inflammatory and neuroprotective properties and reduces the incidence of postoperative cognitive impairments. However, on the whole the molecular mechanisms are poorly understood. This study aims to explore whether dexmedetomidine influences microRNAs (miRNAs) in a rat model of lipopolysaccharide (LPS)-induced neuroinflammation. Adult Wistar rats were injected with 1 mg/kg LPS intraperitoneal (i.p.) in the presence or absence of 5 µg/kg dexmedetomidine. After 6 h, 24 h, and 7 days, gene expressions of interleukin 1-β (IL1-β), tumor necrosis factor-α (TNF-α), and microRNA expressions of miR 124, 132, 134, and 155 were measured in the hippocampus, cortex, and plasma. Dexmedetomidine decreased the LPS-induced neuroinflammation in the hippocampus and cortex via significant reduction of the IL1-β and TNF-α gene expressions after 24 h. Moreover, the LPS-mediated increased expressions of miR 124, 132, 134, and 155 were significantly decreased after dexmedetomidine treatment in both brain regions. In plasma, dexmedetomidine significantly reduced LPS- induced miR 155 after 6 h. Furthermore, there is evidence that miR 132 and 134 may be suitable as potential biomarkers for the detection of neuroinflammation. View Full-Tex

Mgs1 function at G-quadruplex structures during DNA replication

The coordinated action of DNA polymerases and DNA helicases is essential at genomic sites that are hard to replicate. Among these are sites that harbour G-quadruplex DNA structures (G4). G4s are stable alternative DNA structures, which have been implicated to be involved in important cellular processes like the regulation of gene expression or telomere maintenance. G4 structures were shown to hinder replication fork progression and cause genomic deletions, mutations and recombination events. Many helicases unwind G4 structures and preserve genome stability, but a detailed understanding of G4 replication and the re-start of stalled replication forks around formed G4 structures is not clear, yet. In our recent study, we identified that Mgs1 preferentially binds to G4 DNA structures in vitro and is associated with putative G4-forming chromosomal regions in vivo. Mgs1 binding to G4 motifs in vivo is partially dependent on the helicase Pif1. Pif1 is the major G4-unwinding helicase in S. cerevisiae. In the absence of Mgs1, we determined elevated gross chromosomal rearrangement (GCR) rates in yeast, similar to Pif1 deletion. Here, we highlight the recent findings and set these into context with a new mechanistic model. We propose that Mgs1's functions support DNA replication at G4-forming regions

Classification of Microglial Morphological Phenotypes Using Machine Learning

Microglia are the brain’s immunocompetent macrophages with a unique feature that allows surveillance of the surrounding microenvironment and subsequent reactions to tissue damage, infection, or homeostatic perturbations. Thereby, microglia’s striking morphological plasticity is one of their prominent characteristics and the categorization of microglial cell function based on morphology is well established. Frequently, automated classification of microglial morphological phenotypes is performed by using quantitative parameters. As this process is typically limited to a few and especially manually chosen criteria, a relevant selection bias may compromise the resulting classifications. In our study, we describe a novel microglial classification method by morphological evaluation using a convolutional neuronal network on the basis of manually selected cells in addition to classical morphological parameters. We focused on four microglial morphologies, ramified, rod-like, activated and amoeboid microglia within the murine hippocampus and cortex. The developed method for the classification was confirmed in a mouse model of ischemic stroke which is already known to result in microglial activation within affected brain regions. In conclusion, our classification of microglial morphological phenotypes using machine learning can serve as a time-saving and objective method for post-mortem characterization of microglial changes in healthy and disease mouse models, and might also represent a useful tool for human brain autopsy samples

Strukturdynamik- und Maschinendiagnose an einem Cherenkov-Teleskop

Das Deutsche Elektronen-Synchrotron (DESY) in Zeuthen entwickelt zurzeit die 12m-Teleskope im Rahmen des internationalen Cherenkov-Telescope-Array (CTA)-Konsortiums. Es ist geplant, die Teleskope über einen Zeitraum von mindestens 30 Jahren kostengünstig zu betreiben. Dabei führt der autarke Betrieb von Teleskopen an abgelegenen Standorten zu speziellen Anforderungen an die Ausfallsicherheit. Diese Anforderungen gelten sowohl für die Tragstruktur als auch für die Antriebskomponenten. Zustandsüberwachung und Verschleißanalysen sind in diesem Anwendungsfall mit klassischen Methoden der Maschinen- und Strukturüberwachung nur eingeschränkt möglich. Methoden der Mustererkennung aus dem Bereich der Psychoakustik konnten erfolgreich verwendet werden, um den Langsamlauf zu analysieren. In diesem Beitrag werden Ergebnisse von Schwingungsuntersuchungen an einem 1:1-Prototyp des Teleskops vorgestellt. Zusätzlich wird die Frage beantwortet, wie ein kostengünstiges Überwachungssystem mit minimaler Sensoranzahl realisiert werden kann.The Deutsche Elektronen-Synchrotron (DESY) in Zeuthen currently develops telescopes with 12 m diameter in the framework of the international Cherenkov Telescope Array (CTA) consortium. These telescopes are supposed to work cost-efficiently for a period of at least 30 years. Their operation at remote locations leads to special demands for the telescope’s support structure and drive components system stability. In this case, traditional methods of condition monitoring and structural health monitoring reach their limits. Psychoacoustical methods could be successfully applied in order to analyse low revolution speed regime. In this paper, we present results of a structural dynamic analysis of a real-sized telescope prototype. Additionally, the question of how to realise a monitoring system with a minimum amount of sensors is addressed

Dexmedetomidine Restores Autophagic Flux, Modulates Associated microRNAs and the Cholinergic Anti-inflammatory Pathway upon LPS-Treatment in Rats

Infections and perioperative stress can lead to neuroinflammation, which in turn is linked to cognitive impairments such as postoperative delirium or postoperative cognitive dysfunctions. The alpha 2-adrenoceptor agonist dexmedetomidine (DEX) prevents cognitive impairments and has organo-protective and anti-inflammatory properties. Macroautophagy (autophagy) regulates many biological processes, but its role in DEX-mediated anti-inflammation and the underlying mechanism of DEX remains largely unclear. We were interested how a pretreatment with DEX protects against lipopolysaccharide (LPS)-induced inflammation in adult male Wistar rats. We used Western blot and activity assays to study how DEX modulated autophagy- and apoptosis-associated proteins as well as molecules of the cholinergic anti-inflammatory pathway, and qPCR to analyse the expression of autophagy and inflammation-associated microRNAs (miRNA) in the spleen, cortex and hippocampus at different time points (6 h, 24 h, 7 d). We showed that a DEX pretreatment prevents LPS-induced impairments in autophagic flux and attenuates the LPS-induced increase in the apoptosis-associated protein cleaved poly(ADP-ribose)-polymerase (PARP) in the spleen. Both, DEX and LPS altered miRNA expression and molecules of the cholinergic anti-inflammatory pathway in the spleen and brain. While only a certain set of miRNAs was up- and/or downregulated by LPS in each tissue, which was prevented or attenuated by a DEX pretreatment in the spleen and hippocampus, all miRNAs were up- and/or downregulated by DEX itself - independent of whether or not they were altered by LPS. Our results indicate that the organo-protective effect of DEX may be mediated by autophagy, possibly by acting on associated miRNAs, and the cholinergic anti-inflammatory pathway

Author Correction to: Telomerase subunit Est2 marks internal sites that are prone to accumulate DNA damage

An amendment to this paper has been published and can be accessed via the original article.</p