Preparation data of the bromodomains BRD3(1), BRD3(2), BRD4(1), and BRPF1B and crystallization of BRD4(1)-inhibitor complexes

AbstractThis article presents detailed purification procedures for the bromodomains BRD3(1), BRD3(2), BRD4(1), and BRPF1B. In addition we provide crystallization protocols for apo BRD4(1) and BRD4(1) in complex with numerous inhibitors. The protocols described here were successfully applied to obtain affinity data by isothermal titration calorimetry (ITC) and by differential scanning fluorimetry (DSF) as well as structural characterizations of BRD4(1) inhibitor complexes (PDB codes: PDB: 4LYI, PDB: 4LZS, PDB: 4LYW, PDB: 4LZR, PDB: 4LYS, PDB: 5D24, PDB: 5D25, PDB: 5D26, PDB: 5D3H, PDB: 5D3J, PDB: 5D3L, PDB: 5D3N, PDB: 5D3P, PDB: 5D3R, PDB: 5D3S, PDB: 5D3T). These data have been reported previously and are discussed in more detail elsewhere [1,2]

Investigation of the use of a sensor bracelet for the presymptomatic detection of changes in physiological parameters related to COVID-19: an interim analysis of a prospective cohort study (COVI-GAPP).

OBJECTIVES We investigated machinelearningbased identification of presymptomatic COVID-19 and detection of infection-related changes in physiology using a wearable device. DESIGN Interim analysis of a prospective cohort study. SETTING, PARTICIPANTS AND INTERVENTIONS Participants from a national cohort study in Liechtenstein were included. Nightly they wore the Ava-bracelet that measured respiratory rate (RR), heart rate (HR), HR variability (HRV), wrist-skin temperature (WST) and skin perfusion. SARS-CoV-2 infection was diagnosed by molecular and/or serological assays. RESULTS A total of 1.5 million hours of physiological data were recorded from 1163 participants (mean age 44±5.5 years). COVID-19 was confirmed in 127 participants of which, 66 (52%) had worn their device from baseline to symptom onset (SO) and were included in this analysis. Multi-level modelling revealed significant changes in five (RR, HR, HRV, HRV ratio and WST) device-measured physiological parameters during the incubation, presymptomatic, symptomatic and recovery periods of COVID-19 compared with baseline. The training set represented an 8-day long instance extracted from day 10 to day 2 before SO. The training set consisted of 40 days measurements from 66 participants. Based on a random split, the test set included 30% of participants and 70% were selected for the training set. The developed long short-term memory (LSTM) based recurrent neural network (RNN) algorithm had a recall (sensitivity) of 0.73 in the training set and 0.68 in the testing set when detecting COVID-19 up to 2 days prior to SO. CONCLUSION Wearable sensor technology can enable COVID-19 detection during the presymptomatic period. Our proposed RNN algorithm identified 68% of COVID-19 positive participants 2 days prior to SO and will be further trained and validated in a randomised, single-blinded, two-period, two-sequence crossover trial. Trial registration number ISRCTN51255782; Pre-results

Where are we now with European forest multi-taxon biodiversity and where can we head to?

The European biodiversity and forest strategies rely on forest sustainable management (SFM) to conserve forest biodiversity. However, current sustainability assessments hardly account for direct biodiversity indicators. We focused on forest multi-taxon biodiversity to: i) gather and map the existing information; ii) identify knowledge and research gaps; iii) discuss its research potential. We established a research network to fit data on species, standing trees, lying deadwood and sampling unit description from 34 local datasets across 3591 sampling units. A total of 8724 species were represented, with the share of common and rare species varying across taxonomic classes: some included many species with several rare ones (e.g., Insecta); others (e.g., Bryopsida) were represented by few common species. Tree-related structural attributes were sampled in a subset of sampling units (2889; 2356; 2309 and 1388 respectively for diameter, height, deadwood and microhabitats). Overall, multitaxon studies are biased towards mature forests and may underrepresent the species related to other developmental phases. European forest compositional categories were all represented, but beech forests were overrepresented as compared to thermophilous and boreal forests. Most sampling units (94%) were referred to a habitat type of conservation concern. Existing information may support European conservation and SFM strategies in: (i) methodological harmonization and coordinated monitoring; (ii) definition and testing of SFM indicators and thresholds; (iii) data-driven assessment of the effects of environmental and management drivers on multi-taxon forest biological and functional diversity, (iv) multi-scale forest monitoring integrating in-situ and remotely sensed information. Forest biodiversity Multi-taxon Sustainable management Biodiversity conservation Forest stand structurepublishedVersio

Where are we now with European forest multi-taxon biodiversity and where can we head to?

The European biodiversity and forest strategies rely on forest sustainable management (SFM) to conserve forest biodiversity. However, current sustainability assessments hardly account for direct biodiversity indicators. We focused on forest multi-taxon biodiversity to: i) gather and map the existing information; ii) identify knowledge and research gaps; iii) discuss its research potential. We established a research network to fit data on species, standing trees, lying deadwood and sampling unit description from 34 local datasets across 3591 sampling units. A total of 8724 species were represented, with the share of common and rare species varying across taxonomic classes: some included many species with several rare ones (e.g., Insecta); others (e.g., Bryopsida) were represented by few common species. Tree-related structural attributes were sampled in a subset of sampling units (2889; 2356; 2309 and 1388 respectively for diameter, height, deadwood and microhabitats). Overall, multi-taxon studies are biased towards mature forests and may underrepresent the species related to other developmental phases. European forest compositional categories were all represented, but beech forests were over-represented as compared to thermophilous and boreal forests. Most sampling units (94%) were referred to a habitat type of conservation concern. Existing information may support European conservation and SFM strategies in: (i) methodological harmonization and coordinated monitoring; (ii) definition and testing of SFM indicators and thresholds; (iii) data-driven assessment of the effects of environmental and management drivers on multi-taxon forest biological and functional diversity, (iv) multi-scale forest monitoring integrating in-situ and remotely sensed information

Mitochondrial physiology

As the knowledge base and importance of mitochondrial physiology to evolution, health and disease expands, the necessity for harmonizing the terminology concerning mitochondrial respiratory states and rates has become increasingly apparent. The chemiosmotic theory establishes the mechanism of energy transformation and coupling in oxidative phosphorylation. The unifying concept of the protonmotive force provides the framework for developing a consistent theoretical foundation of mitochondrial physiology and bioenergetics. We follow the latest SI guidelines and those of the International Union of Pure and Applied Chemistry (IUPAC) on terminology in physical chemistry, extended by considerations of open systems and thermodynamics of irreversible processes. The concept-driven constructive terminology incorporates the meaning of each quantity and aligns concepts and symbols with the nomenclature of classical bioenergetics. We endeavour to provide a balanced view of mitochondrial respiratory control and a critical discussion on reporting data of mitochondrial respiration in terms of metabolic flows and fluxes. Uniform standards for evaluation of respiratory states and rates will ultimately contribute to reproducibility between laboratories and thus support the development of data repositories of mitochondrial respiratory function in species, tissues, and cells. Clarity of concept and consistency of nomenclature facilitate effective transdisciplinary communication, education, and ultimately further discovery

Mitochondrial physiology

As the knowledge base and importance of mitochondrial physiology to evolution, health and disease expands, the necessity for harmonizing the terminology concerning mitochondrial respiratory states and rates has become increasingly apparent. The chemiosmotic theory establishes the mechanism of energy transformation and coupling in oxidative phosphorylation. The unifying concept of the protonmotive force provides the framework for developing a consistent theoretical foundation of mitochondrial physiology and bioenergetics. We follow the latest SI guidelines and those of the International Union of Pure and Applied Chemistry (IUPAC) on terminology in physical chemistry, extended by considerations of open systems and thermodynamics of irreversible processes. The concept-driven constructive terminology incorporates the meaning of each quantity and aligns concepts and symbols with the nomenclature of classical bioenergetics. We endeavour to provide a balanced view of mitochondrial respiratory control and a critical discussion on reporting data of mitochondrial respiration in terms of metabolic flows and fluxes. Uniform standards for evaluation of respiratory states and rates will ultimately contribute to reproducibility between laboratories and thus support the development of data repositories of mitochondrial respiratory function in species, tissues, and cells. Clarity of concept and consistency of nomenclature facilitate effective transdisciplinary communication, education, and ultimately further discovery

Thermodynamical and structural characterisation of importinβ dependent nuclear import processes

Der aktive Transport von cytoplasmatischen Substraten in den Nucleus wird von Transportrezeptoren vermittelt, welche auch Importine genannt werden. Unter ihnen ist der Kernimportrezeptor Importinβ (Impβ) der wohl vielseitigste: Impβ ist sowohl als einzelner Rezeptor aktiv, als auch im Verbund mit anderen Rezeptoren oder Adaptern. Beispiele für letztere sind der Kernimport von Substraten mit klassischem Kernlokalisationssignal (cNLS-Substrate) gemeinsam mit dem Adapterprotein Importinα (Impα), der Kernimport von H1 Linker Histonen mithilfe des Co-Rezeptors Importin7 (Imp7) und der Kernimport spleißosomaler Untereinheiten, der U snRNPs, im Verbund mit Snurportin1 (SPN1). Diese außergewöhnliche Variabilität von Impβ stand im Mittelpunkt der vorliegenden Arbeit und ihre Ursachen sollten thermodynamisch und strukturell ergründet werden, mit besonderer Fokussierung auf den H1-Kernimport und den Kernimports von U snRNPs. Beim Kernimport von H1 Linker Histonen binden zwar sowohl Impβ als auch Imp7 an das Substrat, jedoch nur als Heterodimer können sie die Translokation von H1 durch die Kernpore bewältigen. In dieser Arbeit konnte die H1-Bindungsstelle von Imp7 ermittelt werden, welche zwei saure Bereiche nahe dem C-Terminus umfaßt, die als mögliche Schleifen identifiziert wurden. Die thermodynamische Analyse der Bildung des H1-Importkomplexes aus Impβ, Imp7 und H1 und seiner Dissoziation durch RanGTP mittels isothermer Titrationskalorimetrie (ITC) zeigte, daß beide Prozesse allosterisch reguliert sind. Dabei konnte dargelegt werden, daß die Bildung des Impβ/Imp7-Heterodimers in vitro enthalpiegetrieben ist, wohingegen die nachfolgende H1-Bindung an das Heterodimer entropiegetrieben ist. Der erforderliche Entropiegewinn wird dabei durch die Freisetzung von Salzionen von der H1-Oberfläche durch die komplementär zu H1 geladene, gemeinsame Oberfläche von Impβ und Imp7 bereitgestellt. Hieraus ist ersichtlich, daß die Energiebarriere bei der H1-Bindung durch den Einsatz eines Rezeptordimers überwunden wird. Außerdem konnte nachgewiesen werden, daß der H1-Bindung eine allosterische Aktivierung von Imp7 durch Impβ vorausgeht, bei welcher die Impβ-Bindungsdomäne von Imp7 eine Schlüsselrolle spielt. Im Kernimport von U snRNPs dagegen bedient sich Impβ nicht eines Co-Rezeptors, sondern eines Adapters für das Substrat, nämlich SPN1. Im Gegensatz zu allen anderen bislang untersuchten Impβ-abhängigen Kernimportprozessen bedarf der Impβ/SPN1/U snRNP-Importkomplex für die Freisetzung vom nuclear basket nach dem Import jedoch nicht des Schalterproteins RanGTP. Die hier vorgestellte Kristallstruktur von Impβ_127-876 im Komplex mit der Impβ-Bindungs-Domäne von SPN1 (IBBSPN1, AS 1-65) offenbart eine weit geöffnete Konformation von Impβ, wie sie einzigartig unter den funktionellen Impβ/Substrat-Komplexen ist. Überraschenderweise gleicht sie vielmehr der Konformation von Impβ/RanGTP. Da die Bindung von RanGTP an Impβ üblicherweise die Freisetzung von Importkomplexen vom nuclear basket auslöst, kann die Schlußfolgerung gezogen werden, daß die freie Dissoziation von Impβ/SPN1 vom nuclear basket durch die Mimikry des RanGTP-gebundenen Zustands ermöglicht wird. Mittels ITC konnte überdies eine Korrelation zwischen der offenen Konformation von Impβ im Komplex mit IBBSPN1 und seiner hohen Molekülentropie belegt werden. Dies steht im Gegensatz zur geschlossenen und daher entropiearmen Konformation von Impβ im Komplex mit Impα. Die hier vorgestellten Ergebnisse verdeutlichen also die zentrale Rolle von molekularen Modulationen im Impβ-abhängigen Kernimport und geben neue Einblicke in Strategien, die Energiebarrieren im Kernimport zu überwinden