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The attractive interaction in conventional BCS superconductors is provided by a bosonic mode. However, the pairing glue of most unconventional superconductors is unknown. The effect of electron-boson coupling is therefore extensively studied in these materials. A key signature is dispersion kinks that can be observed in the spectral function as abrupt changes in velocity and lifetime of quasiparticles. Here, we show the existence of two kinks in the unconventional iron-based superconductor RbFe2As2 using angle-resolved photoemission spectroscopy (ARPES) and dynamical mean field theory (DMFT). In addition, we observe the formation of a Hubbard band multiplet due to the combination of Coulomb interaction and Hund's rule coupling in this multiorbital system. We demonstrate that the two dispersion kinks are a consequence of these strong many-body interactions. This interpretation is in line with a growing number of theoretical predictions for kinks in various general models of correlated materials. Our results provide a unifying link between iron-based superconductors and different classes of correlated, unconventional superconductors such as cuprates and heavy-fermion materials.

We have investigated the charge transfer mechanism in single crystals of DTBDT-TCNQ and DTBDT-F4TCNQ (where DTBDT is dithieno[2,3-d;2',3'-d'] benzo[1,2-b;4,5-b']dithiophene) using a combination of near-edge X-ray absorption spectroscopy (NEXAFS) and density functional theory calculations (DFT) including final state effects beyond the sudden state approximation. In particular, we find that a description that considers the partial screening of the electron-hole Coulomb correlation on a static level as well as the rearrangement of electronic density shows excellent agreement with experiment and allows to uncover the details of the charge transfer mechanism in DTBDT-TCNQ and DTBDT-F4 TCNQ, as well as a reinterpretation of previous NEXAFS data on pure TCNQ. Finally, we further show that almost the same quality of agreement between theoretical results and experiment is obtained by the much faster Z+1/2 approximation, where the core hole effects are simulated by replacing N or F with atomic number Z with the neighboring atom with atomic number Z+1/2. [ABSTRACT FROM AUTHOR]

Introduction: Familial Mediterranean fever (FMF) is a genetic disease leading to recurrent episodes of inflammation. Two pathogenic variants are required for classical disease, but the disease can occur in heterozygous patients. Patients are treated continuously with colchicine to prevent amyloid A (AA) amyloidosis, including heterozygous patients who display a moderate form of FMF and rarely develop AA amyloidosis. The need for lifelong colchicine treatment in heterozygous FMF is therefore controversial. We aimed to characterise genotype-specific levels of inflammatory biomarkers, and to focus on heterozygous patients who discontinued colchicine., Methods: All patients with FMF from the European databases AIDnet and JIRcohort who received colchicine during follow-up were included. Demographics, C reactive protein (CRP), serum amyloid A (SAA), S100A8/A9 and S100A12 levels, leucocyte and neutrophil counts were extracted. Visits were classified as active, subclinical or inactive according to symptoms, CRP and SAA levels., Results: Data from 747 patients were extracted (233 homozygous, 201 compound heterozygous, 224 heterozygous patients, 49 heterozygous with one class III variant and 40 compound heterozygous with two class III variants). During active visits, all biomarker levels were higher compared with inactive visits (p<0.001). Heterozygous patients showed lower levels of CRP, SAA, S100A8/A9 and S100A12 during inactive and subclinical visits than patients with two class IV-V variants. Colchicine was discontinued in 52 heterozygous patients and reintroduced in 23 of them (44%)., Conclusion: S100A8/A9 and S100A12 proteins are biomarkers that can be used to assess disease activity. Heterozygous patients have lower levels of inflammatory biomarkers and some of them can sustainably discontinue colchicine treatment., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Introduction: Respiratory viral infections (RVIs) are a major global contributor to morbidity and mortality. The susceptibility and outcome of RVIs are strongly age-dependent and show considerable inter-population differences, pointing to genetically and/or environmentally driven developmental variability. The factors determining the age-dependency and shaping the age-related changes of human anti-RVI immunity after birth are still elusive., Methods: We are conducting a prospective birth cohort study aiming at identifying endogenous and environmental factors associated with the susceptibility to RVIs and their impact on cellular and humoral immune responses against the influenza A virus (IAV), respiratory syncytial virus (RSV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The MIAI birth cohort enrolls healthy, full-term neonates born at the University Hospital Würzburg, Germany, with follow-up at four defined time-points during the first year of life. At each study visit, clinical metadata including diet, lifestyle, sociodemographic information, and physical examinations, are collected along with extensive biomaterial sampling. Biomaterials are used to generate comprehensive, integrated multi-omics datasets including transcriptomic, epigenomic, proteomic, metabolomic and microbiomic methods., Discussion: The results are expected to capture a holistic picture of the variability of immune trajectories with a focus on cellular and humoral key players involved in the defense of RVIs and the impact of host and environmental factors thereon. Thereby, MIAI aims at providing insights that allow unraveling molecular mechanisms that can be targeted to promote the development of competent anti-RVI immunity in early life and prevent severe RVIs., Clinical Trial Registration: https://drks.de/search/de/trial/, identifier DRKS00034278., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Hartmann, Khan, Schöning, Richter, Willers, Pirr, Heckmann, Dirks, Morbach, Konrad, Fries, Winkler, Büchel, Seidenspinner, Fischer, Vollmuth, Meinhardt, Marissen, Schmolke, Haid, Pietschmann, Backes, Dölken, Löber, Keil, Heuschmann, Wöckel, Sagar, Ulas, Forslund-Startceva, Härtel and Viemann.)

Natural killer (NK) cells are present in the circulation and can also be found residing in tissues, and these populations exhibit distinct developmental requirements and are thought to differ in terms of ontogeny. Here, we investigate whether circulating conventional NK (cNK) cells can develop into long-lived tissue-resident NK (trNK) cells following acute infections. We found that viral and bacterial infections of the skin triggered the recruitment of cNK cells and their differentiation into Tcf1 hi CD69 hi trNK cells that share transcriptional similarity with CD56 bright TCF1 hi NK cells in human tissues. Skin trNK cells arose from interferon (IFN)-γ-producing effector cells and required restricted expression of the transcriptional regulator Blimp1 to optimize Tcf1-dependent trNK cell formation. Upon secondary infection, trNK cells rapidly gained effector function and mediated an accelerated NK cell response. Thus, cNK cells redistribute and permanently position at sites of previous infection via a mechanism promoting tissue residency that is distinct from Hobit-dependent developmental paths of NK cells and ILC1 seeding tissues during ontogeny., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Engineered vesicular stomatitis virus (VSV) pseudotyping offers an essential method for exploring virus-cell interactions, particularly for viruses that require high biosafety levels. Although this approach has been employed effectively, the current methodologies for virus visualization and labeling can interfere with infectivity and lead to misinterpretation of results. In this study, we introduce an innovative approach combining genetic code expansion (GCE) and click chemistry with pseudotyped VSV to produce highly fluorescent and infectious pseudoviruses (clickVSVs). These clickVSVs enable robust and precise virus-cell interaction studies without compromising the biological function of the viral surface proteins. We evaluated this approach by generating VSVs bearing a unique chemical handle for click labeling and assessing the infectivity in relevant cell lines. Our results demonstrate that clickVSVs maintain their infectivity post-labeling and present an efficiency about two times higher in detecting surface proteins compared to classical immunolabeling. The utilization of clickVSVs further allowed us to visualize and track 3D virus binding and infection in living cells, offering enhanced observation of virus-host interactions. Thus, clickVSVs provide an efficient alternative for virus-associated research under the standard biosafety levels.

The angiotensin-converting enzyme 2 (ACE2) has been identified as entry receptor on cells enabling binding and infection with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) via trimeric spike (S) proteins protruding from the viral surface. It has been suggested that trimeric S proteins preferably bind to plasma membrane areas with high concentrations of possibly multimeric ACE2 receptors to achieve a higher binding and infection efficiency. Here we used direct stochastic optical reconstruction microscopy (dSTORM) in combination with different labeling approaches to visualize the distribution and quantify the expression of ACE2 on different cells. Our results reveal that endogenous ACE2 receptors are present as monomers in the plasma membrane with densities of only 1-2 receptors μm -2 . In addition, binding of trimeric S proteins does not induce the formation of ACE2 oligomers in the plasma membrane. Supported by infection studies using vesicular stomatitis virus (VSV) particles bearing S proteins our data demonstrate that a single S protein interaction per virus particle with a monomeric ACE2 receptor is sufficient for infection, which provides SARS-CoV-2 a high infectivity., (© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

The herpes simplex virus 1 (HSV-1) virion host shut-off (vhs) protein cleaves both cellular and viral mRNAs by a translation-initiation-dependent mechanism, which should spare circular RNAs (circRNAs). Here, we show that vhs-mediated degradation of linear mRNAs leads to an enrichment of circRNAs relative to linear mRNAs during HSV-1 infection. This was also observed in influenza A virus (IAV) infection, likely due to degradation of linear host mRNAs mediated by the IAV PA-X protein and cap-snatching RNA-dependent RNA polymerase. For most circRNAs, enrichment was not due to increased circRNA synthesis but due to a general loss of linear RNAs. In contrast, biogenesis of a circRNA originating from the long isoform (NEAT1&#95;2) of the nuclear paraspeckle assembly transcript 1 (NEAT1) was induced both in HSV-1 infection-in a vhs-independent manner-and in IAV infection. This was associated with induction of novel linear splicing of NEAT1&#95;2 both within and downstream of the circRNA. NEAT1&#95;2 forms a scaffold for paraspeckles, nuclear bodies located in the interchromatin space, must likely remain unspliced for paraspeckle assembly and is up-regulated in HSV-1 and IAV infection. We show that NEAT1&#95;2 splicing and up-regulation can be induced by ectopic co-expression of the HSV-1 immediate-early proteins ICP22 and ICP27, potentially linking increased expression and splicing of NEAT1&#95;2. To identify other conditions with NEAT1&#95;2 splicing, we performed a large-scale screen of published RNA-seq data. This uncovered both induction of NEAT1&#95;2 splicing and poly(A) read-through similar to HSV-1 and IAV infection in cancer cells upon inhibition or knockdown of CDK7 or the MED1 subunit of the Mediator complex phosphorylated by CDK7. In summary, our study reveals induction of novel circular and linear NEAT1&#95;2 splicing isoforms as a common characteristic of HSV-1 and IAV infection and highlights a potential role of CDK7 in HSV-1 or IAV infection., Competing Interests: The authors have declared that no competing interests exist.

Unhoused people have higher COVID-19 mortality and lower vaccine uptake than housed community members. Understanding vaccine hesitancy among unhoused people is key for developing programs that address their unique needs. A three-round, rapid, field-based survey was conducted to describe attitudes toward COVID-19 vaccination. Round 1 assessed vaccine brand preference, round 2 assessed intention to accept a financial incentive for vaccination, and round 3 measured vaccine uptake and assessed reasons for vaccine readiness during implementation of a financial incentive program. A total of 5177 individuals were approached at COVID-19 vaccination events for unhoused people in Los Angeles County from May through November 2021. Analyses included 4949 individuals: 3636 (73.5%) unsheltered and 1313 (26.5%) sheltered. Per self-report, 2008 (40.6%) were already vaccinated, 1732 (35%) wanted to get vaccinated, 359 (7.3%) were not yet ready, and 850 (17.2%) did not want to get vaccinated. Brand preference was evenly split among participants (Moderna 31.0%, J&J 35.5%, either 33.5%, p = 0.74). Interest in a financial incentive differed between those who were not yet ready and those who did not want to get vaccinated (43.2% vs. 16.2%, p < 0.01). After implementing a financial incentive program, 97.4% of participants who indicated interest in vaccination were vaccinated that day; the financial incentive was the most cited reason for vaccine readiness (n = 731, 56%). This study demonstrated the utility of an iterative, field-based assessment for program implementation during the rapidly evolving pandemic. Personal engagement, a variety of brand choices, and financial incentives could be important for improving vaccine uptake among unhoused people., (© 2022. The Author(s).)

Neutralizing antibodies against SARS-CoV-2 are important to protect against infection and/or disease. Using an assay to detect antibodies directed against the receptor binding domain (RBD) of SARS-CoV-2 Spike, we identified individuals with SARS-CoV-2 infection after an outbreak at a local health institution. All but one COVID-19 patient developed detectable anti-RBD antibodies and 77% had virus neutralizing antibody titers of >1:25. Antibody levels declined slightly over time. However, we still detected virus neutralizing antibody titers in 64% of the COVID-19 patients at >300 days after infection, demonstrating durability of neutralizing antibody levels after infection. Importantly, full COVID-19 vaccination of these individuals resulted in higher antibody titers compared to fully vaccinated individuals in the absence of prior infection. These data demonstrate long-lived antibody-mediated immunity after SARS-CoV-2 infection, and a clear benefit of two vaccine doses for recovered individuals., (© 2022 The Authors. European Journal of Immunology published by Wiley-VCH GmbH.)

Learning and assessment are increasingly mediated by digital technologies. Thus, learners' experiences with these digital technologies are growing in importance, as they might affect learning and assessment. The present paper explores the impact of user experience on digital concept mapping. It builds on user experience theory to explain variance in the intention to use digital concept mapping tools and in concept map-based assessment scores. Furthermore, it identifies fulfillment of psychological needs as an important driver of positive experiences. In a field study in three schools and a university (N = 71), we tested two concept mapping prototypes on computers and tablets. We found that user experience is a significant factor explaining variance in intention to use. User experience also explained variance in three out of four concept mapping scores on tablets, potentially related to the lower pragmatic quality of the tablet prototypes. Fulfillment of psychological needs strongly affected perceptions of different qualities of user experience with digital concept mapping. These results indicate that user experience needs to be considered in digital concept mapping to provide a positive and successful environment for learning and assessment. Finally, we discuss implications for designers of digital learning and assessment tools., Competing Interests: The authors declare no conflict of interest., (© 2022 The Author(s).)

Modern fluoroscopes used for image guidance have become quite complex. Adding to this complexity are the many regulatory and accreditation requirements that must be fulfilled during acceptance testing of a new unit. Further, some of these acceptance tests have pass/fail criteria, whereas others do not, making acceptance testing a subjective and time-consuming task. The AAPM Task Group 272 Report spells out the details of tests that are required and gives visibility to some of the tests that while not yet required are recommended as good practice. The organization of the report begins with the most complicated fluoroscopes used in interventional radiology or cardiology and continues with general fluoroscopy and mobile C-arms. Finally, the appendices of the report provide useful information, an example report form and topics that needed their own section due to the level of detail., (© 2022 American Association of Physicists in Medicine.)

The functional and structural characterization of macromolecular complexes requires protocols for their native isolation. Here, we describe a protocol for this task based on the recombinant poxvirus Vaccinia expressing tagged proteins of interest in infected cells. Tagged proteins and their interactors can then be isolated via affinity chromatography. The procedure is illustrated for the Vaccinia virus encoded multi-subunit RNA polymerase. Our protocol also allows the expression and isolation of heterologous proteins and hence is suitable for a broader application. For complete details on the use and execution of this profile, please refer to Grimm et al. (2019)., Competing Interests: The authors declare no competing interests., (© 2021 The Authors.)

SPT6 is a histone chaperone that tightly binds RNA polymerase II (RNAPII) during transcription elongation. However, its primary role in transcription is uncertain. We used targeted protein degradation to rapidly deplete SPT6 in human cells and analyzed defects in RNAPII behavior by a multi-omics approach and mathematical modeling. Our data indicate that SPT6 is a crucial factor for RNAPII processivity and is therefore required for the productive transcription of protein-coding genes. Unexpectedly, SPT6 also has a vital role in RNAPII termination, as acute depletion induced readthrough transcription for thousands of genes. Long-term depletion of SPT6 induced cryptic intragenic transcription, as observed earlier in yeast. However, this phenotype was not observed upon acute SPT6 depletion and therefore can be attributed to accumulated epigenetic perturbations in the prolonged absence of SPT6. In conclusion, targeted degradation of SPT6 allowed the temporal discrimination of its function as an epigenetic safeguard and RNAPII elongation factor., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)

For the control of immunity in COVID-19 survivors and vaccinated subjects, there is an urgent need for reliable and rapid serological assays. Based on samples from 63 COVID-19 survivors up to 7 months after symptom onset, and on 50 serum samples taken before the beginning of the pandemic, we compared the performances of three commercial immunoassays for the detection of SARS-CoV-2 IgA and IgG antibodies (Euroimmun SARS-COV-2 IgA/IgG, Mikrogen recom Well SARS-CoV-2 IgA/IgG, and Serion ELISA agile SARS-CoV-2 IgA/IgG) and three rapid lateral flow (immunochromatographic) tests (Abbott PanBio COVID-19 IgG/IgM, Nadal COVID-19 IgG/IgM, and Cleartest Corona 2019-nCOV IgG/IgM) with a 50% plaque-reduction neutralization test (PRNT50) representing the gold standard. Fifty-seven out of 63 PCR-confirmed COVID-19 patients (90%) showed neutralizing antibodies. The sensitivity of the seven assays ranged from 7.0% to 98.3%, and the specificity ranged from 86.0% to 100.0%. Only one commercial immunoassay showed a sensitivity and specificity of greater than 98%.

Most mitochondrial proteins are synthesized as precursors in the cytosol and post-translationally transported into mitochondria. The mitochondrial surface protein Tom70 acts at the interface of the cytosol and mitochondria. In vitro import experiments identified Tom70 as targeting receptor, particularly for hydrophobic carriers. Using in vivo methods and high-content screens, we revisit the question of Tom70 function and considerably expand the set of Tom70-dependent mitochondrial proteins. We demonstrate that the crucial activity of Tom70 is its ability to recruit cytosolic chaperones to the outer membrane. Indeed, tethering an unrelated chaperone-binding domain onto the mitochondrial surface complements most of the defects caused by Tom70 deletion. Tom70-mediated chaperone recruitment reduces the proteotoxicity of mitochondrial precursor proteins, particularly of hydrophobic inner membrane proteins. Thus, our work suggests that the predominant function of Tom70 is to tether cytosolic chaperones to the outer mitochondrial membrane, rather than to serve as a mitochondrion-specifying targeting receptor., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)

To slow down the proliferation of Covid-19, governments virtually shut down public life, temporarily closed schools, and forced teaching to be done exclusively on a remote basis. These measures offer an opportunity to reexamine conventional teaching and learning arrangements, test new digital and analogue concepts, and provide essential inspiration for curriculum making in the twenty-first century. This article addresses the historical development of schooling in the classroom as differentiated from "homeschooling". On one hand, the question of how school closures and digitally supported teaching settings may affect an increase in educational inequalities is investigated using an international comparison. On the other hand, the pedagogical and didactical implications of distance learning and a digital teaching culture, which constitute the foundation for digital curriculum making, are examined., (© The Author(s) 2021.)

We investigate the role of nonlocal correlations in LiFeAs by exploring an ab initio-derived multiorbital Hubbard model for LiFeAs via the two-particle self-consistent (TPSC) approach. The multiorbital formulation of TPSC approximates the irreducible interaction vertex to be an orbital-dependent constant, which is self-consistently determined from local spin and charge sum rules. Within this approach, we disentangle the contribution of local and nonlocal correlations in LiFeAs and show that in the local approximation one recovers the dynamical mean field theory result. The comparison of our theoretical results to most recent angular-resolved photoemission spectroscopy and de Haas-van Alphen data shows that nonlocal correlations in LiFeAs are decisive to describe the measured spectral function A(k[over →],ω), Fermi surface, and scattering rates. These findings underline the importance of nonlocal correlations and benchmark different theoretical approaches for iron-based superconductors.

Mitochondria are unique organelles harboring two distinct membranes, the mitochondrial inner and outer membrane (MIM and MOM, respectively). Mitochondria comprise only a subset of metabolic pathways for the synthesis of membrane lipids; therefore most lipid species and their precursors have to be imported from other cellular compartments. One such import process is mediated by the ER mitochondria encounter structure (ERMES) complex. Both mitochondrial membranes surround the hydrophilic intermembrane space (IMS). Therefore, additional systems are required that shuttle lipids between the MIM and MOM. Recently, we identified the IMS protein Mcp2 as a high-copy suppressor for cells that lack a functional ERMES complex. To understand better how mitochondria facilitate transport and biogenesis of lipids, we searched for genetic interactions of this suppressor. We found that MCP2 has a negative genetic interaction with the gene TGL2 encoding a neutral lipid hydrolase. We show that this lipase is located in the intermembrane space of the mitochondrion and is imported via the Mia40 disulfide relay system. Furthermore, we show a positive genetic interaction of double deletion of MCP2 and PSD1 , the gene encoding the enzyme that synthesizes the major amount of cellular phosphatidylethanolamine. Finally, we demonstrate that the nucleotide-binding motifs of the predicted atypical kinase Mcp2 are required for its proper function. Taken together, our data suggest that Mcp2 is involved in mitochondrial lipid metabolism and an increase of this involvement by overexpression suppresses loss of ERMES.

Class II hydrophobins are amphiphilic proteins produced by filamentous fungi. One of their typical features is the tendency to accumulate at the interface between an aqueous phase and a hydrophobic phase, such as the air-water interface. The kinetics of the interfacial self-assembly of wild-type hydrophobins HFBI and HFBII and some of their engineered variants at the air-water interface were measured by monitoring the accumulated mass at the interface via nondestructive ellipsometry measurements. The resulting mass vs time curves revealed unusual kinetics for a monolayer formation that did not follow a typical Langmuir-type of behavior but had a rather coverage-independent rate instead. Typically, the full surface coverage was obtained at masses corresponding to a monolayer. The formation of multilayers was not observed. Atomic force microscopy revealed formation and growth of non-fusing protein clusters at the interface. The mechanism of the adsorption was studied by varying the structure or charges of the protein or the ionic strength of the subphase, revealing that the lateral interactions between the hydrophobins play a role in their interfacial assembly. Additionally, a theoretical model was introduced to identify the underlying mechanism of the unconventional adsorption kinetics.

The mitochondrial intermembrane space evolved from the bacterial periplasm. Presumably as a consequence of their common origin, most proteins of these compartments are stabilized by structural disulfide bonds. The molecular machineries that mediate oxidative protein folding in bacteria and mitochondria, however, appear to share no common ancestry. Here we tested whether the enzymes Erv1 and Mia40 of the yeast mitochondrial disulfide relay could be functionally replaced by corresponding components of other compartments. We found that the sulfhydryl oxidase Erv1 could be replaced by the Ero1 oxidase or the protein disulfide isomerase from the endoplasmic reticulum, however at the cost of respiration deficiency. In contrast to Erv1, the mitochondrial oxidoreductase Mia40 proved to be indispensable and could not be replaced by thioredoxin-like enzymes, including the cytoplasmic reductase thioredoxin, the periplasmic dithiol oxidase DsbA, and Pdi1. From our studies we conclude that the profound inertness against glutathione, its slow oxidation kinetics and its high affinity to substrates renders Mia40 a unique and essential component of mitochondrial biogenesis. Evidently, the development of a specific mitochondrial disulfide relay system represented a crucial step in the evolution of the eukaryotic cell., (© The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The aim of this study is to tailor the inner structure of positively charged poly-( N-isopropylacrylamid- co-allylamine) (P(NIPAM- co-AA)) microgels for a better control of the distribution of negatively charged magnetic cobaltferrite (CoFe 2 O 4 @CA) nanoparticles (MNPs) within the microgels. Therefore, two different strategies are followed for the microgel synthesis: the (one pot) batch method which leads to a higher cross-linker density in the microgel core and the feeding method which compensates different reaction kinetics of the cross-linker and the monomers. The latter one is expected to result in a homogeneous cross-linker distribution. Information about the cross-linker distribution is indirectly gained by measuring the elastic modulus via indentation experiments with an atomic force microscope. While the batch method results in a higher elastic modulus in the center of the microgel indicating a core/shell structure, the feeding method leads to a constant elastic modulus over the whole microgel. The loading with MNPs and their distribution are studied with transmission electron microscopy (TEM). The TEM images show a large difference in the MNP distribution which is correlated to the cross-linker distribution of both types of microgels. The batch method microgel has a low MNP concentration in the core. The feeding method microgel shows a much more homogeneous distribution of MNPs across the microgel. The latter one also shows a stronger charge reversal which is a hint for a higher loading of the feeding method microgel. Dynamic light scattering and electrophoretic mobility measurements demonstrate that for both types of microgels, the temperature sensitivity is preserved after loading with MNPs.

The review addresses nanomechanics and nanorheology of stimuli responsive microgels adsorbed at an interface. In order to measure the mechanical properties on a local scale, an atomic force microscope is used. The tip presents an indenter with a radius of curvature of a few 10 s of nm. Static indentation experiments and dynamic studies with an excited cantilever are presented. The effect of several internal and external parameters on the mechanical properties is reviewed. The focus is on the correlation between the swelling abilities of the gels and their mechanical properties. Several results are surprising and show that the relationship is not as simple as one might expect.

The biogenesis of mitochondria depends on the import of hundreds of preproteins. N-terminal matrix-targeting signals (MTSs) direct preproteins to the surface receptors Tom20, Tom22, and Tom70. In this study, we show that many preproteins contain additional internal MTS-like signals (iMTS-Ls) in their mature region that share the characteristic properties of presequences. These features allow the in silico prediction of iMTS-Ls. Using Atp1 as model substrate, we show that iMTS-Ls mediate the binding to Tom70 and have the potential to target the protein to mitochondria if they are presented at its N terminus. The import of preproteins with high iMTS-L content is significantly impaired in the absence of Tom70, whereas preproteins with low iMTS-L scores are less dependent on Tom70. We propose a stepping stone model according to which the Tom70-mediated interaction with internal binding sites improves the import competence of preproteins and increases the efficiency of their translocation into the mitochondrial matrix., (© 2018 Backes et al.)

The present study addresses the multiresponsive behavior of poly(N-isopropylacrylamide) (PNIPAM) microgels adsorbed to interfaces. The microgels react to changes in temperature by shrinking in aqueous solution above their volume phase transition temperature (VPTT). Additionally, they shrink in mixtures of water and ethanol, although both individual liquids are good solvents for PNIPAM. The combination of this so-called cononsolvency effect and the temperature response of adsorbed microgels is studied by atomic force microscopy (AFM). Adsorbed microgels are of special interest because they are compressed considerably compared to those in bulk solution. It is shown that the impact of adsorption on swelling depends on the specific surface details, as well as the sample preparation. Thereby, the microgels are deposited on two different kinds of surfaces: on gold surface and on polycation (PAH) coating which show different interactions with the microgels in terms of electrostatic interaction and wettability. In addition, the microgels were deposited from different solvent mixtures. This influences the microgel structure and thereby the swelling properties. Nanorheology studies by dynamic AFM measurements lead to surprising results which are explained by the fact that not only polymer density but a subtle interaction between polymer and solvent might dominate the rheological properties. This work supports the view that preferential adsorption of ethanol at PNIPAM drives cononsolvency, while the shrinking at T > VPTT is caused by general breaking of hydrogen bonds between solvents and PNIPAM.

Mitochondria contain two aqueous subcompartments, the matrix and the intermembrane space (IMS). The matrix is enclosed by both the inner and outer mitochondrial membranes, whilst the IMS is sandwiched between the two. Proteins of the matrix are synthesized in the cytosol as preproteins, which contain amino-terminal matrix targeting sequences that mediate their translocation through translocases embedded in the outer and inner membrane. For these proteins, the translocation reaction is driven by the import motor which is part of the inner membrane translocase. The import motor employs matrix Hsp70 molecules and ATP hydrolysis to ratchet proteins into the mitochondrial matrix. Most IMS proteins lack presequences and instead utilize the IMS receptor Mia40, which facilitates their translocation across the outer membrane in a reaction that is coupled to the formation of disulfide bonds within the protein. This process requires neither ATP nor the mitochondrial membrane potential. Mia40 fulfills two roles: First, it acts as a holdase, which is crucial in the import of IMS proteins and second, it functions as a foldase, introducing disulfide bonds into newly imported proteins, which induces and stabilizes their natively folded state. For several Mia40 substrates, oxidative folding is an essential prerequisite for their assembly into oligomeric complexes. Interestingly, recent studies have shown that the two functions of Mia40 can be experimentally separated from each other by the use of specific mutants, hence providing a powerful new way to dissect the different physiological roles of Mia40. In this review we summarize the current knowledge relating to the mitochondrial matrix-targeting and the IMS-targeting/Mia40 pathway. Moreover, we discuss the mechanistic properties by which the mitochondrial import motor on the one hand and Mia40 on the other, drive the translocation of their substrates into the organelle. We propose that the lateral diffusion of Mia40 in the inner membrane and the oxidation-mediated folding of incoming polypeptides supports IMS import.

Background: Many proteins of the mitochondrial intermembrane space (IMS) contain structural disulfide bonds formed by the mitochondrial disulfide relay. In fungi and animals, the sulfhydryl oxidase Erv1 'generates' disulfide bonds that are passed on to the oxidoreductase Mia40, which oxidizes substrate proteins. A different structural organization of plant Erv1 proteins compared to that of animal and fungal orthologs was proposed to explain its inability to complement the corresponding yeast mutant., Results: Herein, we have revisited the biochemical and functional properties of Arabidopsis thaliana Erv1 by both in vitro reconstituted activity assays and complementation of erv1 and mia40 yeast mutants. These mutants were viable, however, they showed severe defects in the biogenesis of IMS proteins. The plant Erv1 was unable to oxidize yeast Mia40 and rather even blocked its activity. Nevertheless, it was able to mediate the import and folding of mitochondrial proteins., Conclusions: We observed that plant Erv1, unlike its homologs in fungi and animals, can promote protein import and oxidative protein folding in the IMS independently of the oxidoreductase Mia40. In accordance to the absence of Mia40 in many protists, our study suggests that the mitochondrial disulfide relay evolved in a stepwise reaction from an Erv1-only system to which Mia40 was added in order to improve substrate specificity. Graphical Abstract The mitochondrial disulfide relay evolved in a step-wise manner from an Erv1-only system.

Poly( N -isopropylacrylamide) (PNIPAM) microgel is a smart polymer that shows a volume phase transition temperature (VPTT) at around 32 °C in aqueous solutions, above which it collapses. In this work, combining experiments and molecular simulations, it is shown that PNIPAM microgels do not always exhibit a collapsed structure above the VPTT. Instead, PNIPAM in aqueous alcohol mixtures shows a two-step conformational transition, i.e., a collapse at low temperatures ( T < 32 °C) and a reswelling when T > 50 °C. The present analysis indicates that delicate microscopic interaction details, together with the bulk solution properties, play a key role in dictating the reswelling behavior. Even when PNIPAM microgels swell with increasing T , this is not a standard upper critical solution behavior.

Biomimetic multilayers based on layer-by-layer (LbL) assembly were prepared as functional films with compact structure by incorporating the mussel-inspired catechol cross-linking. Dopamine-modified poly(acrylic acid) (PAADopa) was synthesized as a polyanion to offer electrostatic interaction with the prelayer polyethylenimine (PEI) and consecutively cross-linked by zinc to generate compact multilayers with tunable physicochemical properties. In situ layer-by-layer growth and cross-linking were monitored by a quartz crystal microbalance with dissipation (QCM-D) to reveal the kinetics of the process and the influence of Dopa chemistry. Addition of Dopa enhanced the mass adsorption and led to the formation of a more compact structure. An increase of ionic strength induced an increase in mass adsorption in the Dopa-cross-linked multilayers. This is a universal approach for coating of various surfaces such as Au, SiO2, Ti, and Al2O3. Roughness observed by AFM in both wet and dry conditions was compared to confirm the compact morphology of Dopa-cross-linked multilayers. Because of the pH sensitivity of Dopa moiety, metal-chelated Dopa groups can be turned into softer structure at higher pH as revealed by reduction of Young's modulus determined by MFP-3D AFM. A deeper insight into the growth and mechanical properties of Dopa-cross-linked polyelectrolyte multilayers was addressed in the present study. This allows a better control of these systems for bioapplications.