Uncovering the forces between nucleosomes using DNA origami

Revealing the energy landscape for nucleosome association may contribute to the understanding of higher-order chromatin structures and their impact on genome regulation. We accomplish this in a direct measurement by integrating two nucleosomes into a DNA origami-based force spectrometer, which enabled subnanometer-resolution measurements of nucleosome-nucleosome distance frequencies via single-particle electron microscopy imaging. From the data, we derived the Boltzmann-weighted distance-dependent energy landscape for nucleosome pair interactions. We find a shallow but long-range (similar to 6 nm) attractive nucleosome pair potential with a minimum of -1.6 kcal/mol close to direct contact distances. The relative nucleosome orientation had little influence, but histone H4 acetylation or removal of histone tails drastically decreased the interaction strength. Because of the weak and shallow pair potential, high-erorder nucleosome assemblies will be compliant and experience dynamic shape fluctuations in the absence of additional cofactors. Our results contribute to a more accurate description of chromatin and our force spectrometer provides a powerful tool for the direct and high-resolution study of molecular interactions using imaging techniques

Uncovering the forces between nucleosomes using DNA origami

Revealing the energy landscape for nucleosome association may contribute to the understanding of higher-order chromatin structures and their impact on genome regulation. We accomplish this in a direct measurement by integrating two nucleosomes into a DNA origami-based force spectrometer, which enabled subnanometer-resolution measurements of nucleosome-nucleosome distance frequencies via single-particle electron microscopy imaging. From the data, we derived the Boltzmann-weighted distance-dependent energy landscape for nucleosome pair interactions. We find a shallow but long-range (similar to 6 nm) attractive nucleosome pair potential with a minimum of -1.6 kcal/mol close to direct contact distances. The relative nucleosome orientation had little influence, but histone H4 acetylation or removal of histone tails drastically decreased the interaction strength. Because of the weak and shallow pair potential, high-erorder nucleosome assemblies will be compliant and experience dynamic shape fluctuations in the absence of additional cofactors. Our results contribute to a more accurate description of chromatin and our force spectrometer provides a powerful tool for the direct and high-resolution study of molecular interactions using imaging techniques

The Changing Dynamics Of A Japanese Company Amid The Globalizing Business Environment: A Case Study Of Hitachi Construction Machinery Company

Japan’s mature market combined with its decreasing population has left little room for domestic market expansion. Because of this, many Japanese companies have been required to explore overseas markets in order to maintain and expand their sales revenue. While numerous Japanese companies have struggled to adapt to globalization, Hitachi Ltd., a Japanese multinational conglomerate, has become a frontrunner in its organizational transformation. A recent example is its move from the traditional, seniority and lifelong employment system to a merit-based salary system, which aims to attract talented, high-level workers overseas. Hitachi Construction Machinery (HCM), one of Hitachi’s eleven businesses, is an excellent case study of how Japanese companies are changing in response to globalization. Three research groups investigated this on-going, momentous transformation of a representative of Japan Inc., HCM, through first-hand experiences and semi-structured interviews with company representatives throughout Japan. The research included in-depth investigations of how globalization has affected HCM in three areas: language, cultural, and unification training processes; human resource management; and reaction to global environmental issues through its product lines

Development and Presentation of an Ethical Framework for Health and Medical Apps

The ubiquity of mobile applications makes it difficult to weigh up opportunities and potential risks. One significant aspect of an ethical assessment regarding the development and use of health and medical apps is the lack of clear and consistent guidelines and measures of value. Aim: The aim of this paper was to identify the main ethical determinants associated in the context of health and medical apps and develop a framework for ethical reflection. Methods: A systematic literature search in the database PubMed was undertaken and the journal Biomed Central Medical Ethics was hand searched and 32 met the review criteria. On this basis of the review an interview guide was developed and five purposively selected experts on the field of payers, science, consumers, medical ethics and society were interviewed regarding health and medical apps. Results: There is a lack of consistent and clear guidelines regarding the use of health and medical apps in literature. Indeed, findings refer to established biomedical principles. The interviews identified that several insecurities related to legal and regulatory issues are common. On the other hand, many advantages of the use of health and medical apps have been pointed out by experts. Conclusion: Debates on ethics and the use of health and medical apps should play a significant role in public discussion. Ethical endpoints cannot be considered isolated from each other, and must be regarded as a complex network of aspects connected with each other. Further research should concentrate on the development of consistent guidelines and a framework for reflection of occurring ethical concerns

The role of apoptosis repressor with a CARD domain (ARC) in the therapeutic resistance of renal cell carcinoma (RCC): the crucial role of ARC in the inhibition of extrinsic and intrinsic apoptotic signalling

Background: Renal cell carcinomas (RCCs) display broad resistance against conventional radio- and chemotherapies, which is due at least in part to impairments in both extrinsic and intrinsic apoptotic pathways. One important anti-apoptotic factor that is strongly overexpressed in RCCs and known to inhibit both apoptotic pathways is ARC (apoptosis repressor with a CARD domain). Methods: Expression and subcellular distribution of ARC in RCC tissue samples and RCC cell lines were determined by immunohistochemistry and fluorescent immunohistochemistry, respectively. Extrinsic and intrinsic apoptosis signalling were induced by TRAIL (TNF-related apoptosis-inducing ligand), ABT-263 or topotecan. ARC knock-down was performed in clearCa-12 cells using lentiviral transduction of pGIPZ. shRNAmir constructs. Extrinsic respectively intrinsic apoptosis were induced by TRAIL (TNF-related apoptosis-inducing ligand), ABT263 or topotecan. Potential synergistic effects were tested by pre-treatment with topotecan and subsequent treatment with ABT263. Activation of different caspases and mitochondrial depolarisation (JC-1 staining) were analysed by flow cytometry. Protein expression of Bcl-2 family members and ARC in RCC cell lines was measured by Western blotting. Statistical analysis was performed by Student’s t-test. Results: Regarding the extrinsic pathway, ARC knockdown strongly enhanced TRAIL-induced apoptosis by increasing the activation level of caspase-8. Regarding the intrinsic pathway, ARC, which was only weakly expressed in the nuclei of RCCs in vivo, exerted its anti-apoptotic effect by impairing mitochondrial activation rather than inhibiting p53. Topotecan- and ABT-263-induced apoptosis was strongly enhanced following ARC knockdown in RCC cell lines. In addition, topotecan pre-treatment enhanced ABT-263-induced apoptosis and this effect was amplified in ARC-knockdown cells. Conclusion: Taken together, our results are the first to demonstrate the importance of ARC protein in the inhibition of both the extrinsic and intrinsic pathways of apoptosis in RCCs. In this context, ARC cooperates with anti-apoptotic Bcl-2 family members to exert its strong anti-apoptotic effects and is therefore an important factor not only in the therapeutic resistance but also in future therapy strategies (i.e., Bcl-2 inhibitors) in RCC. In sum, targeting of ARC may enhance the therapeutic response in combination therapy protocols

Das NetzDG in der praktischen Anwendung

What are the practical effects of the Network Enforcement Act, which came into force in 2017 and was controversial from the start? Which compliance structures have the big three social networks Facebook, YouTube and Twitter implemented with regard to the removal of illegal user content? In addition to a presentation and evaluation of previous studies, monitoring reports and NetzDG reports from the social networks, possible criteria for “over-blocking” are identified and subsumed in this third volume of the series. On the occasion of the NetzDG evaluation commissioned by the federal government, the study was carried out additionally and independently of this. It is not based on any commissioning by public or private bodies.Welche Auswirkungen hat das 2017 in Kraft getretene, von Beginn an umstrittene Netzwerkdurchsetzungsgesetz in der praktischen Anwendung? Welche Compliance-Strukturen haben die großen drei Sozialen Netzwerke Facebook, YouTube und Twitter in Bezug auf die Entfernung rechtswidriger Nutzerinhalte umgesetzt? Neben einer Darstellung und Bewertung bisheriger Studien, Monitoring-Berichte und NetzDG-Reports der Sozialen Netzwerke werden im vorliegenden dritten Band der Schriftenreihe auch mögliche Kriterien für ein “Overblocking” eruiert und subsumiert. Die Studie ist aus Anlass der von der Bundesregierung beauftragten NetzDG-Evaluation zusätzlich und unabhängig hiervon durchgeführt worden. Ihr liegt keine Beauftragung durch öffentliche oder private Stellen zugrunde

The spike gene is a major determinant for the SARS-CoV-2 Omicron-BA.1 phenotype.

Variant of concern (VOC) Omicron-BA.1 has achieved global predominance in early 2022. Therefore, surveillance and comprehensive characterization of Omicron-BA.1 in advanced primary cell culture systems and animal models are urgently needed. Here, we characterize Omicron-BA.1 and recombinant Omicron-BA.1 spike gene mutants in comparison with VOC Delta in well-differentiated primary human nasal and bronchial epithelial cells in vitro, followed by in vivo fitness characterization in hamsters, ferrets and hACE2-expressing mice, and immunized hACE2-mice. We demonstrate a spike-mediated enhancement of early replication of Omicron-BA.1 in nasal epithelial cultures, but limited replication in bronchial epithelial cultures. In hamsters, Delta shows dominance over Omicron-BA.1, and in ferrets Omicron-BA.1 infection is abortive. In hACE2-knock-in mice, Delta and a Delta spike clone also show dominance over Omicron-BA.1 and an Omicron-BA.1 spike clone, respectively. Interestingly, in naïve K18-hACE2 mice, we observe Delta spike-mediated increased replication and pathogenicity and Omicron-BA.1 spike-mediated reduced replication and pathogenicity, suggesting that the spike gene is a major determinant of replication and pathogenicity. Finally, the Omicron-BA.1 spike clone is less well-controlled by mRNA-vaccination in K18-hACE2-mice and becomes more competitive compared to the progenitor and Delta spike clones, suggesting that spike gene-mediated immune evasion is another important factor that led to Omicron-BA.1 dominance