Disulfide bridge-dependent dimerization triggers FGF2 membrane translocation into the extracellular space.

Fibroblast growth factor 2 (FGF2) exits cells by direct translocation across the plasma membrane, a type I pathway of unconventional protein secretion. This process is initiated by phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2)-dependent formation of highly dynamic FGF2 oligomers at the inner plasma membrane leaflet, inducing the formation of lipidic membrane pores. Cell surface heparan sulfate chains linked to glypican-1 (GPC1) capture FGF2 at the outer plasma membrane leaflet, completing FGF2 membrane translocation into the extracellular space. While the basic steps of this pathway are well understood, the molecular mechanism by which FGF2 oligomerizes on membrane surfaces remains unclear. In the current study, we demonstrate the initial step of this process to depend on C95-C95 disulfide-bridge-mediated FGF2 dimerization on membrane surfaces, producing the building blocks for higher FGF2 oligomers that drive the formation of membrane pores. We find FGF2 with a C95A substitution to be defective in oligomerization, pore formation, and membrane translocation. Consistently, we demonstrate a C95A variant of FGF2 to be characterized by a severe secretion phenotype. By contrast, while also important for efficient FGF2 secretion from cells, a second cysteine residue on the molecular surface of FGF2 (C77) is not involved in FGF2 oligomerization. Rather, we find C77 to be part of the interaction interface through which FGF2 binds to the α1 subunit of the Na,K-ATPase, the landing platform for FGF2 at the inner plasma membrane leaflet. Using cross-linking mass spectrometry, atomistic molecular dynamics simulations combined with a machine learning analysis and cryo-electron tomography, we propose a mechanism by which disulfide-bridged FGF2 dimers bind with high avidity to PI(4,5)P2 on membrane surfaces. We further propose a tight coupling between FGF2 secretion and the formation of ternary signaling complexes on cell surfaces, hypothesizing that C95-C95-bridged FGF2 dimers are functioning as the molecular units triggering autocrine and paracrine FGF2 signaling.

Information technology and changing role models in German libraries: The example of OPEN-CAM.

Germany has a long tradition of health libraries. From their origin in monasteries, they became centres of knowledge and education. In modern times, this tradition has been continued by the Central Library of Medicine. In addition, as a specialty in Germany, special collection areas and special libraries that focus on one topic were established. Those services were transformed to specialized information services and portals as part of the digital transformation process. One of such projects is OPEN-CAM, which provides literature on integrative medicine in a specialist library and hosts the literature database CAMbase. Based on this example, we show how digital transformation has influenced the library landscape and its structures in Germany.

Data about marine area-based management tools to assess their contribution to the UN sustainable development goals.

The dataset presented in this article contains information about marine Area-Based Management Tools (ABMTs) used to assess their contribution to the United Nations 2030 Sustainable Development Goals. Following the scope of the analysis, ABMTs were identified by scrutinizing international and regional legal sources related to ocean management in the fields of marine conservation, fisheries, deep sea bed mining, underwater natural and cultural heritage, environmental conservation, and marine spatial planning. Legal sources were screened to depict the following characteristics of individual ABMTs: i) management objectives; ii) authorities responsible for delivering such objectives; iii) the system of management and planning entailed in the ABMT including the zoning type; and iv) the specific spatial scope and domain each ABMT refer to in vertical depth and horizontal domain. Data were generated through an internal expert elicitation. Experts, initially trained in the data analysis and related protocol, contributed to the data production because of their specific knowledge and experience in ocean management. This dataset represents a unique source of information for advancing research about monitoring and assessment of the achievement of sustainable development goals that encompasses different types of ABMTs.

Reaction Time-Based Cognitive Assessments in Virtual Reality - A Feasibility Study with an Age Diverse Sample.

INTRODUCTION: While virtual reality (VR) is an emerging paradigm in a variety of research contexts, VR-based embodiment effects on behavior and performance still lack in sufficient evidence regarding to bias in cognitive performance assessment. METHODS: In this methodological observational study, we compare the VR measurement of cognitive performance with a conventional computer-based testing approach in real life (RL) in younger and older adults. The differences between VR and RL scenarios are investigated using the background of two theoretical models from cognitive psychology. Furthermore, data assessment reliability and validity are analyzed, concerning the feasibility of technological and ergonomic aspects. RESULTS: A within-group comparison showed no change in information processing speed in either one of the two age groups, i.e., both groups perform equally well in RL and in a VR testing environment. CONCLUSION: The use of lifelike VR environments for cognitive performance tests seems not to lead to any performance changes compared to RL computer-based assessments, making VR suitable for similar applications. On technical concerns, we recommend the careful use of reaction time paradigms regarding to input hardware and stimuli presentation.

IU - A Digital Application for the Graphical Examination of Interpersonal Attachment.

Human attachment describes the establishment of contact between two or more people leading to a closer interpersonal relationship. For measuring attachment, the use of nonverbal assessments tools including art and drawing tasks has been shown to be an alternative to conventional assessment approaches. The present study aims at evaluating the internal criterion validity of a new digital drawing tool for measuring interpersonal attachment. 68 participants took part in this pilot study and were separated in groups of two. After completion of a 10-item subscale of Social Orientation (SO) they were asked to sit opposite to the other and to hold eye contact during a three minute drawing period. Moving the pen to the upper section near the partner stood for thoughts about the other, while moving to the lower section closest to the subject implicated thoughts about oneself. The mean distance of the resulting time series of the two subjects were calculated, using the mean Euclidean distance, and compared with the difference in the SO values via linear regression. Taking all differences together a moderate correlation of r = 0.298 was observed, which however slightly missed the level of significance (p = 0.09). We were able to find small evidence for the criterion validity of IU digital drawing tool. For future studies, other measures of similarity in the time series, i.e. the Manhattan Distance are discussed as an extension to foster the present results.

Integrated ocean management for a sustainable ocean economy.

The rapidly evolving ocean economy, driven by human needs for food, energy, transportation and recreation, has led to unprecedented pressures on the ocean that are further amplified by climate change, loss of biodiversity and pollution. The need for better governance of human activities in the ocean space has been widely recognized for years, and is now also incorporated in the United Nations Sustainable Development Goals. Even so, many challenges relating to the implementation of existing governance frameworks exist. Here, we argue that integrated ocean management (IOM) should be the key overarching approach-building upon and connecting existing sectoral governance efforts-for achieving a sustainable ocean economy. IOM is a holistic, ecosystem-based and knowledge-based approach that aims to ensure the sustainability and resilience of marine ecosystems while integrating and balancing different ocean uses to optimize the overall ocean economy. We discuss examples of IOM in practice from areas where preconditions differ substantially, and identify six universal opportunities for action that can help achieve a sustainable ocean economy.

Development and characterization of DNAzyme candidates demonstrating significant efficiency against human rhinoviruses.

BACKGROUND: Infections with human rhinoviruses (RVs) are responsible for millions of common cold episodes and the majority of asthma exacerbations, especially in childhood. No drugs specifically targeting RVs are available. OBJECTIVE: We sought to identify specific anti-RV molecules based on DNAzyme technology as candidates to a clinical study. METHODS: A total of 226 candidate DNAzymes were designed against 2 regions of RV RNA genome identified to be sufficiently highly conserved between virus strains (ie, the 5'-untranslated region and cis-acting replication element) by using 3 test strains: RVA1, RVA16, and RVA29. All DNAzymes were screened for their cleavage efficiency against in vitro-expressed viral RNA. Those showing any catalytic activity were subjected to bioinformatic analysis of their reverse complementarity to 322 published RV genomic sequences. Further molecular optimization was conducted for the most promising candidates. Cytotoxic and off-target effects were excluded in HEK293 cell-based systems. Antiviral efficiency was analyzed in infected human bronchial BEAS-2B cells and ex vivo-cultured human sinonasal tissue. RESULTS: Screening phase-generated DNAzymes characterized by either good catalytic activity or by high RV strain coverage but no single molecule represented a satisfactory combination of those 2 features. Modifications in length of the binding domains of 2 lead candidates, Dua-01(-L12R9) and Dua-02(-L10R11), improved their cleavage efficiency to an excellent level, with no loss in eminent strain coverage (about 98%). Both DNAzymes showed highly favorable cytotoxic/off-target profiles. Subsequent testing of Dua-01-L12R9 in BEAS-2B cells and sinonasal tissue demonstrated its significant antiviral efficiency. CONCLUSIONS: Effective and specific management of RV infections with Dua-01-L12R9 might be useful in preventing asthma exacerbations, which should be verified by clinical trials.

Key steps in unconventional secretion of fibroblast growth factor 2 reconstituted with purified components.

FGF2 is secreted from cells by an unconventional secretory pathway. This process is mediated by direct translocation across the plasma membrane. Here, we define the minimal molecular machinery required for FGF2 membrane translocation in a fully reconstituted inside-out vesicle system. FGF2 membrane translocation is thermodynamically driven by PI(4,5)P2-induced membrane insertion of FGF2 oligomers. The latter serve as dynamic translocation intermediates of FGF2 with a subunit number in the range of 8-12 FGF2 molecules. Vectorial translocation of FGF2 across the membrane is governed by sequential and mutually exclusive interactions with PI(4,5)P2 and heparan sulfates on opposing sides of the membrane. Based on atomistic molecular dynamics simulations, we propose a mechanism that drives PI(4,5)P2 dependent oligomerization of FGF2. Our combined findings establish a novel type of self-sustained protein translocation across membranes revealing the molecular basis of the unconventional secretory pathway of FGF2.

Antisense molecules: A new class of drugs.

An improved understanding of disease pathogenesis leads to identification of novel therapeutic targets. From a pharmacologic point of view, these can be addressed by small chemical compounds, so-called biologicals (eg, mAbs and recombinant proteins), or by a rather new class of molecule based on the antisense concept. Recently, a new wave of clinical studies exploring antisense strategies is evolving. In addition to cancer, they include predominantly trials on infectious and noninfectious diseases, such as chronic inflammatory and metabolic conditions. This article, based on a systematic PubMed literature search, highlights recent developments in this emerging field.

Formation of disulfide bridges drives oligomerization, membrane pore formation, and translocation of fibroblast growth factor 2 to cell surfaces.

Fibroblast growth factor 2 (FGF2) is a key signaling molecule in tumor-induced angiogenesis. FGF2 is secreted by an unconventional secretory mechanism that involves phosphatidylinositol 4,5-bisphosphate-dependent insertion of FGF2 oligomers into the plasma membrane. This process is regulated by Tec kinase-mediated tyrosine phosphorylation of FGF2. Molecular interactions driving FGF2 monomers into membrane-inserted FGF2 oligomers are unknown. Here we identify two surface cysteines that are critical for efficient unconventional secretion of FGF2. They represent unique features of FGF2 as they are absent from all signal-peptide-containing members of the FGF protein family. We show that phosphatidylinositol 4,5-bisphosphate-dependent FGF2 oligomerization concomitant with the generation of membrane pores depends on FGF2 surface cysteines as either chemical alkylation or substitution with alanines impairs these processes. We further demonstrate that the FGF2 variant forms lacking the two surface cysteines are not secreted from cells. These findings were corroborated by experiments redirecting a signal-peptide-containing FGF family member from the endoplasmic reticulum/Golgi-dependent secretory pathway into the unconventional secretory pathway of FGF2. Cis elements known to be required for unconventional secretion of FGF2, including the two surface cysteines, were transplanted into a variant form of FGF4 without signal peptide. The resulting FGF4/2 hybrid protein was secreted by unconventional means. We propose that the formation of disulfide bridges drives membrane insertion of FGF2 oligomers as intermediates in unconventional secretion of FGF2.

Micellar and biochemical properties of a propyl-ended fluorinated surfactant designed for membrane-protein study.

Our goal is to design optimised fluorinated surfactants for handling membrane proteins in solution. We report herein the self-assembling and biochemical properties of a new hemifluorinated surfactant (H3F6H3DigluM) with a branched diglucosylated polar head group and an apolar tail consisting of a perfluorohexane core decorated with a hydrogenated propyl tip. For the sake of comparison, its fluorinated analogue without propyl tip (F6H3DigluM) was also studied. Isothermal titration calorimetry and surface tension showed that the addition of a propyl tip has a significant effect on the overall hydrophobicity of the surfactant, in contrast to the behaviour described when adding an ethyl tip to a fluorinated surfactant. From dynamic light scattering, analytical ultracentrifugation and small-angle X-ray scattering, both H3F6H3DigluM and F6H3DigluM self-assemble into small globular micelles of 5-7 nm in diameter and have aggregation numbers of 62±8 and 46±2, respectively. Finally, H3F6H3DigluM was found to be the best fluorinated surfactant developed in our group to stabilise the model membrane protein bacteriorhodopsin (bR) in aqueous solution. This study demonstrates the suitability of this new propyl-ended fluorinated surfactant for biochemical and structural applications and confirms the superiority of hemifluorinated chains over fluorinated ones.

Sequence-specific dimerization of a transmembrane helix in amphipol A8-35.

As traditional detergents might destabilize or even denature membrane proteins, amphiphilic polymers have moved into the focus of membrane-protein research in recent years. Thus far, Amphipols are the best studied amphiphilic copolymers, having a hydrophilic backbone with short hydrophobic chains. However, since stabilizing as well as destabilizing effects of the Amphipol belt on the structure of membrane proteins have been described, we systematically analyze the impact of the most commonly used Amphipol A8-35 on the structure and stability of a well-defined transmembrane protein model, the glycophorin A transmembrane helix dimer. Amphipols are not able to directly extract proteins from their native membranes, and detergents are typically replaced by Amphipols only after protein extraction from membranes. As Amphipols form mixed micelles with detergents, a better understanding of Amphipol-detergent interactions is required. Therefore, we analyze the interaction of A8-35 with the anionic detergent sodium dodecyl sulfate and describe the impact of the mixed-micelle-like system on the stability of a transmembrane helix dimer. As A8-35 may highly stabilize and thereby rigidify a transmembrane protein structure, modest destabilization by controlled addition of detergents and formation of mixed micellar systems might be helpful to preserve the function of a membrane protein in Amphipol environments.