Retraction: Photoactivatable Hsp47: A Tool to Regulate Collagen Secretion and Assembly.

Adv. Sci. 2019, 6, 1801982 DOI: 10.1002/advs.201801982 The above article, published online on May 3, 2019, in Wiley Online Library (https://doi.org/10.1002/advs.201801982), has been retracted by agreement between the authors, the journal Editor-in-Chief Kirsten Severing, and Wiley-VCH GmbH. The retraction has been agreed on following concerns raised by a third party and a subsequent investigation by the corresponding authors. Data depicted in Figure 4 and Figure 5 could not be reproduced in follow-up experiments. Therefore, the conclusions associated with those figures in the article are considered invalid. E.S.K. participated in the study design, performed measurements, analyzed the data, compiled the figures and participated in manuscript writing. A.d.C. and S.S. participated in the study design, research supervision, and manuscript writing. J.I.P. participated in the study design. M.K.L.H. participated in research supervision and manuscript revision. C.M. assisted with the experimental procedures and data collection.

Inhibition of Collagenase Q1 of Bacillus cereus as a Novel Antivirulence Strategy for the Treatment of Skin-Wound Infections.

Despite the progress in surgical techniques and antibiotic prophylaxis, opportunistic wound infections with Bacillus cereus remain a public health problem. Secreted toxins are one of the main factors contributing to B. cereus pathogenicity. A promising strategy to treat such infections is to target these toxins and not the bacteria. Although the exoenzymes produced by B. cereus are thoroughly investigated, little is known about the role of B. cereus collagenases in wound infections. In this report, the collagenolytic activity of secreted collagenases (Col) is characterized in the B. cereus culture supernatant (csn) and its isolated recombinantly produced ColQ1 is characterized. The data reveals that ColQ1 causes damage on dermal collagen (COL). This results in gaps in the tissue, which might facilitate the spread of bacteria. The importance of B. cereus collagenases is also demonstrated in disease promotion using two inhibitors. Compound 2 shows high efficacy in peptidolytic, gelatinolytic, and COL degradation assays. It also preserves the fibrillar COLs in skin tissue challenged with ColQ1, as well as the viability of skin cells treated with B. cereus csn. A Galleria mellonella model highlights the significance of collagenase inhibition in vivo.

Optoregulated force application to cellular receptors using molecular motors.

Progress in our understanding of mechanotransduction events requires noninvasive methods for the manipulation of forces at molecular scale in physiological environments. Inspired by cellular mechanisms for force application (i.e. motor proteins pulling on cytoskeletal fibers), we present a unique molecular machine that can apply forces at cell-matrix and cell-cell junctions using light as an energy source. The key actuator is a light-driven rotatory molecular motor linked to polymer chains, which is intercalated between a membrane receptor and an engineered biointerface. The light-driven actuation of the molecular motor is converted in mechanical twisting of the entangled polymer chains, which will in turn effectively "pull" on engaged cell membrane receptors (e.g., integrins, T cell receptors) within the illuminated area. Applied forces have physiologically-relevant magnitude and occur at time scales within the relevant ranges for mechanotransduction at cell-friendly exposure conditions, as demonstrated in force-dependent focal adhesion maturation and T cell activation experiments. Our results reveal the potential of nanomotors for the manipulation of living cells at the molecular scale and demonstrate a functionality which at the moment cannot be achieved by other technologies for force application.

Photoactivatable Hsp47: A Tool to Regulate Collagen Secretion and Assembly.

Collagen is the most abundant structural protein in mammals and is crucial for the mechanical integrity of tissues. Hsp47, an endoplasmic reticulum resident collagen-specific chaperone, is involved in collagen biosynthesis and plays a fundamental role in the folding, stability, and intracellular transport of procollagen triple helices. This work reports on a photoactivatable derivative of Hsp47 that allows regulation of collagen biosynthesis within mammalian cells using light. Photoactivatable Hsp47 contains a non-natural light-responsive tyrosine (o-nitro benzyl tyrosine (ONBY)) at Tyr383 position of the protein sequence. This mutation renders Hsp47 inactive toward collagen binding. The inactive, photoactivatable protein is easily uptaken by cells within a few minutes of incubation, and accumulated at the endoplasmic reticulum via retrograde KDEL receptor-mediated uptake. Upon light exposure, the photoactivatable Hsp47 turns into functional Hsp47 in situ. The increased intracellular concentration of Hsp47 results in stimulated secretion of collagen. The ability to promote collagen synthesis on demand, with spatiotemporal resolution, and in diseased state cells is demonstrated in vitro. It is envisioned that photoactivatable Hsp47 allows unprecedented fundamental studies of collagen biosynthesis, matrix biology, and inspires new therapeutic concepts in biomedicine and tissue regeneration.

The microstructural evolution of water ice in the solar system through sintering.

Ice sintering is a form of metamorphism that drives the microstructural evolution of an aggregate of grains through surface and volume diffusion. This leads to an increase in the grain-to-grain contact area ("neck") and density of the aggregate over time, resulting in the evolution of its strength, porosity, thermal conductivity, and other properties. This process plays an important role in the evolution of icy planetary surfaces, though its rate and nature are not well constrained. In this study, we explore the model of Swinkels and Ashby (1981), and assess the extent to which it can be used to quantify sintering timescales for water ice. We compare predicted neck growth rates to new and historical observations of ice sintering, and find agreement to some studies at the order of magnitude level. First-order estimates of neck growth timescales on planetary surfaces show that ice may undergo significant modification over geologic timescales, even in the outer solar system. Densification occurs over much longer timescales, suggesting some surfaces may develop cohesive, but porous, crusts. Sintering rates are extremely sensitive to temperature and grain size, occurring faster in warmer aggregates of smaller grains. This suggests that the microstructural evolution of ices may vary not only throughout the solar system, but also spatially across the surface and in the near-surface of a given body. Our experimental observations of complex grain growth and mass redistribution in ice aggregates point to components of the model that may benefit from improvement, and areas where additional laboratory studies are needed.

Zygotic vinculin is not essential for embryonic development in zebrafish.

The formation of multicellular tissues during development is governed by mechanical forces that drive cell shape and tissue architecture. Protein complexes at sites of adhesion to the extracellular matrix (ECM) and cell neighbors, not only transmit these mechanical forces, but also allow cells to respond to changes in force by inducing biochemical feedback pathways. Such force-induced signaling processes are termed mechanotransduction. Vinculin is a central protein in mechanotransduction that in both integrin-mediated cell-ECM and cadherin-mediated cell-cell adhesions mediates force-induced cytoskeletal remodeling and adhesion strengthening. Vinculin was found to be important for the integrity and remodeling of epithelial tissues in cell culture models and could therefore be expected to be of broad importance in epithelial morphogenesis in vivo. Besides a function in mouse heart development, however, the importance of vinculin in morphogenesis of other vertebrate tissues has remained unclear. To investigate this further, we knocked out vinculin functioning in zebrafish, which contain two fully functional isoforms designated as vinculin A and vinculin B that both show high sequence conservation with higher vertebrates. Using TALEN and CRISPR-Cas gene editing technology we generated vinculin-deficient zebrafish. While single vinculin A mutants are viable and able to reproduce, additional loss of zygotic vinculin B was lethal after embryonic stages. Remarkably, vinculin-deficient embryos do not show major developmental defects, apart from mild pericardial edemas. These results lead to the conclusion that vinculin is not of broad importance for the development and morphogenesis of zebrafish tissues.

Resolving the cadherin-F-actin connection.

Cadherin adhesion complexes have recently emerged as sensors of tissue tension that regulate key developmental processes. Super-resolution microscopy experiments now unravel the spatial organization of the interface between cadherins and the actin cytoskeleton and reveal how vinculin, a central component in cadherin mechanotransduction, is regulated by mechanical and biochemical signals.

αE-catenin-dependent mechanotransduction is essential for proper convergent extension in zebrafish.

Cadherin complexes mediate cell-cell adhesion and are crucial for embryonic development. Besides their structural function, cadherin complexes also transduce tension across the junction-actomyosin axis into proportional biochemical responses. Central to this mechanotransduction is the stretching of the cadherin-F-actin-linker α-catenin, which opens its central domain for binding to effectors such as vinculin. Mechanical unfolding of α-catenin leads to force-dependent reinforcement of cadherin-based junctions as studied in cell culture. The importance of cadherin mechanotransduction for embryonic development has not been studied yet. Here we used TALEN-mediated gene disruption to perturb endogenous αE-catenin in zebrafish development. Zygotic α-catenin mutants fail to maintain their epithelial barrier, resulting in tissue rupturing. We then specifically disrupted mechanotransduction, while maintaining cadherin adhesion, by expressing an αE-catenin construct in which the mechanosensitive domain was perturbed. Expression of either wild-type or mechano-defective α-catenin fully rescues barrier function in α-catenin mutants; however, expression of mechano-defective α-catenin also induces convergence and extension defects. Specifically, the polarization of cadherin-dependent, lamellipodia-driven cell migration of the lateral mesoderm was lost. These results indicate that cadherin mechanotransduction is crucial for proper zebrafish morphogenesis, and uncover one of the essential processes affected by its perturbation.

Converging and Unique Mechanisms of Mechanotransduction at Adhesion Sites.

The molecular mechanisms by which physical forces control tissue development are beginning to be elucidated. Sites of adhesion between both cells and the extracellular environment [extracellular matrix (ECM) or neighboring cells] contain protein complexes capable of sensing fluctuations in tensile forces. Tension-dependent changes in the dynamics and composition of these complexes mark the transformation of physical input into biochemical signals that defines mechanotransduction. It is becoming apparent that, although the core constituents of these different adhesions are distinct, principles and proteins involved in mechanotransduction are conserved. Here, we discuss the current knowledge of overlapping and distinct aspects of mechanotransduction between integrin and cadherin adhesion complexes.

Mapping the cellular response to small molecules using chemogenomic fitness signatures.

Genome-wide characterization of the in vivo cellular response to perturbation is fundamental to understanding how cells survive stress. Identifying the proteins and pathways perturbed by small molecules affects biology and medicine by revealing the mechanisms of drug action. We used a yeast chemogenomics platform that quantifies the requirement for each gene for resistance to a compound in vivo to profile 3250 small molecules in a systematic and unbiased manner. We identified 317 compounds that specifically perturb the function of 121 genes and characterized the mechanism of specific compounds. Global analysis revealed that the cellular response to small molecules is limited and described by a network of 45 major chemogenomic signatures. Our results provide a resource for the discovery of functional interactions among genes, chemicals, and biological processes.

Ubiquitination of CXCR7 controls receptor trafficking.

The chemokine receptor CXCR7 binds CXCL11 and CXCL12 with high affinity, chemokines that were previously thought to bind exclusively to CXCR4 and CXCR3, respectively. Expression of CXCR7 has been associated with cardiac development as well as with tumor growth and progression. Despite having all the canonical features of G protein-coupled receptors (GPCRs), the signalling pathways following CXCR7 activation remain controversial, since unlike typical chemokine receptors, CXCR7 fails to activate Gα(i)-proteins. CXCR7 has recently been shown to interact with ß-arrestins and such interaction has been suggested to be responsible for G protein-independent signals through ERK-1/2 phosphorylation. Signal transduction by CXCR7 is controlled at the membrane by the process of GPCR trafficking. In the present study we investigated the regulatory processes triggered by CXCR7 activation as well as the molecular interactions that participate in such processes. We show that, CXCR7 internalizes and recycles back to the cell surface after agonist exposure, and that internalization is not only ß-arrestin-mediated but also dependent on the Serine/Threonine residues at the C-terminus of the receptor. Furthermore we describe, for the first time, the constitutive ubiquitination of CXCR7. Such ubiquitination is a key modification responsible for the correct trafficking of CXCR7 from and to the plasma membrane. Moreover, we found that CXCR7 is reversibly de-ubiquitinated upon treatment with CXCL12. Finally, we have also identified the Lysine residues at the C-terminus of CXCR7 to be essential for receptor cell surface delivery. Together these data demonstrate the differential regulation of CXCR7 compared to the related CXCR3 and CXCR4 receptors, and highlight the importance of understanding the molecular determinants responsible for this process.

Compound prioritization methods increase rates of chemical probe discovery in model organisms.

Preselection of compounds that are more likely to induce a phenotype can increase the efficiency and reduce the costs for model organism screening. To identify such molecules, we screened ~81,000 compounds in Saccharomyces cerevisiae and identified ~7500 that inhibit cell growth. Screening these growth-inhibitory molecules across a diverse panel of model organisms resulted in an increased phenotypic hit-rate. These data were used to build a model to predict compounds that inhibit yeast growth. Empirical and in silico application of the model enriched the discovery of bioactive compounds in diverse model organisms. To demonstrate the potential of these molecules as lead chemical probes, we used chemogenomic profiling in yeast and identified specific inhibitors of lanosterol synthase and of stearoyl-CoA 9-desaturase. As community resources, the ~7500 growth-inhibitory molecules have been made commercially available and the computational model and filter used are provided.

A comparison of five approaches to measurement of anatomic knee alignment from radiographs.

OBJECTIVE: The recent recognition of the correlation of the hip-knee-ankle angle (HKA) with femur-tibia angle (FTA) on a standard knee radiograph has led to the increasing inclusion of FTA assessments in OA studies due to its clinical relevance, cost effectiveness and minimal radiation exposure. Our goal was to investigate the performance metrics of currently used methods of FTA measurement to determine whether a specific protocol could be recommended based on these results. METHODS: Inter- and intra-rater reliability of FTA measurements were determined by intraclass correlation coefficient (ICC) of two independent analysts. Minimal detectable differences were determined and the correlation of FTA and HKA was analyzed by linear regression. Differences among methods of measuring HKA were assessed by ANOVA. RESULTS: All five methods of FTA measurement demonstrated high precision by inter- and intra-rater reproducibility (ICCs>or=0.93). All five methods displayed good accuracy, but after correction for the offset of FTA from HKA, the femoral notch landmark method was the least accurate. However, the methods differed according to their minimal detectable differences; the FTA methods utilizing the center of the base of the tibial spines or the center of the tibial plateau as knee center landmarks yielded the smallest minimal detectable differences (1.25 degrees and 1.72 degrees, respectively). CONCLUSION: All methods of FTA were highly reproducible, but varied in their accuracy and sensitivity to detect meaningful differences. Based on these parameters we recommend standardizing measurement angles with vertices at the base of the tibial spines or the center of the tibia and comparing single-point and two-point methods in larger studies.

The lakes of Titan.

The surface of Saturn's haze-shrouded moon Titan has long been proposed to have oceans or lakes, on the basis of the stability of liquid methane at the surface. Initial visible and radar imaging failed to find any evidence of an ocean, although abundant evidence was found that flowing liquids have existed on the surface. Here we provide definitive evidence for the presence of lakes on the surface of Titan, obtained during the Cassini Radar flyby of Titan on 22 July 2006 (T16). The radar imaging polewards of 70 degrees north shows more than 75 circular to irregular radar-dark patches, in a region where liquid methane and ethane are expected to be abundant and stable on the surface. The radar-dark patches are interpreted as lakes on the basis of their very low radar reflectivity and morphological similarities to lakes, including associated channels and location in topographic depressions. Some of the lakes do not completely fill the depressions in which they lie, and apparently dry depressions are present. We interpret this to indicate that lakes are present in a number of states, including partly dry and liquid-filled. These northern-hemisphere lakes constitute the strongest evidence yet that a condensable-liquid hydrological cycle is active in Titan's surface and atmosphere, in which the lakes are filled through rainfall and/or intersection with the subsurface 'liquid methane' table.