Mechanical activation opens a lipid-lined pore in OSCA ion channels.

OSCA/TMEM63 channels are the largest known family of mechanosensitive channels1-3, playing critical roles in plant4-7 and mammalian8,9 mechanotransduction. Here we determined 44 cryogenic electron microscopy structures of OSCA/TMEM63 channels in different environments to investigate the molecular basis of OSCA/TMEM63 channel mechanosensitivity. In nanodiscs, we mimicked increased membrane tension and observed a dilated pore with membrane access in one of the OSCA1.2 subunits. In liposomes, we captured the fully open structure of OSCA1.2 in the inside-in orientation, in which the pore shows a large lateral opening to the membrane. Unusually for ion channels, structural, functional and computational evidence supports the existence of a 'proteo-lipidic pore' in which lipids act as a wall of the ion permeation pathway. In the less tension-sensitive homologue OSCA3.1, we identified an 'interlocking' lipid tightly bound in the central cleft, keeping the channel closed. Mutation of the lipid-coordinating residues induced OSCA3.1 activation, revealing a conserved open conformation of OSCA channels. Our structures provide a global picture of the OSCA channel gating cycle, uncover the importance of bound lipids and show that each subunit can open independently. This expands both our understanding of channel-mediated mechanotransduction and channel pore formation, with important mechanistic implications for the TMEM16 and TMC protein families.

In situ architecture of Opa1-dependent mitochondrial cristae remodeling.

Cristae membrane state plays a central role in regulating mitochondrial function and cellular metabolism. The protein Optic atrophy 1 (Opa1) is an important crista remodeler that exists as two forms in the mitochondrion, a membrane-anchored long form (l-Opa1) and a processed short form (s-Opa1). The mechanisms for how Opa1 influences cristae shape have remained unclear due to lack of native three-dimensional views of cristae. We perform in situ cryo-electron tomography of cryo-focused ion beam milled mouse embryonic fibroblasts with defined Opa1 states to understand how each form of Opa1 influences cristae architecture. In our tomograms, we observe a variety of cristae shapes with distinct trends dependent on s-Opa1:l-Opa1 balance. Increased l-Opa1 levels promote cristae stacking and elongated mitochondria, while increased s-Opa1 levels correlated with irregular cristae packing and round mitochondria shape. Functional assays indicate a role for l-Opa1 in wild-type apoptotic and calcium handling responses, and show a compromised respiratory function under Opa1 imbalance. In summary, we provide three-dimensional visualization of cristae architecture to reveal relationships between mitochondrial ultrastructure and cellular function dependent on Opa1-mediated membrane remodeling.

High spatial-resolved heat manipulating membrane heterogeneity alters cellular migration and signaling.

Plasma membrane heterogeneity is a key biophysical regulatory principle of membrane protein dynamics, which further influences downstream signal transduction. Although extensive biophysical and cell biology studies have proven membrane heterogeneity is essential to cell fate, the direct link between membrane heterogeneity regulation to cellular function remains unclear. Heterogeneous structures on plasma membranes, such as lipid rafts, are transiently assembled, thus hard to study via regular techniques. Indeed, it is nearly impossible to perturb membrane heterogeneity without changing plasma membrane compositions. In this study, we developed a high-spatial resolved DNA-origami-based nanoheater system with specific lipid heterogeneity targeting to manipulate the local lipid environmental temperature under near-infrared (NIR) laser illumination. Our results showed that the targeted heating of the local lipid environment influences the membrane thermodynamic properties, which further triggers an integrin-associated cell migration change. Therefore, the nanoheater system was further applied as an optimized therapeutic agent for wound healing. Our strategy provides a powerful tool to dynamically manipulate membrane heterogeneity and has the potential to explore cellular function through changes in plasma membrane biophysical properties.

Reciprocal polarization imaging of optical activity in reflection.

We present reciprocal polarization imaging for the optical activity of chiral media in reflection geometry. The method is based on the reciprocal polar decomposition of backscattering Mueller matrices accounting for the reciprocity of light waves in forward and backward scattering paths. Anisotropic depolarization is introduced to gain sensitivity to optical activity in backscattering. Experiments with glucose solutions show that while the Lu-Chipman decomposition of the backscattering Mueller matrices produces erroneous results, reciprocal polarization imaging correctly retrieves the optical activity of chiral media. The recovered optical rotation agrees with that obtained in the forward geometry and increases linearly with the concentration and thickness of the chiral media. The potential for in vivo glucose monitoring based on optical activity sensing using reciprocal polarization imaging is then discussed.

Rare earth metals detection and recognition based on laser induced breakdown spectroscopy and machine learning.

The rapid detection and identification of the electronic waste (e-waste) containing rare earth (RE) elements is of great significance for the recycling of RE elements. However, the analysis of these materials is extremely challenging due to extreme similarities in appearance or chemical composition. In this research, a new system based on laser induced breakdown spectroscopy (LIBS) and machine learning algorithms is developed for identifying and classifying e-waste of rare-earth phosphors (REPs). Three different kinds of phosphors are selected and the spectra is monitored using this new developed system. The analysis of phosphor spectra shows that there are Gd, Yd, and Y RE element spectra in the phosphor. The results also verify that LIBS could be used to detect RE elements. An unsupervised learning method, principal component analysis (PCA), is used to distinguish the three phosphors and training data set is stored for further identification. Additionally, a supervised learning method, backpropagation artificial neural network (BP-ANN) algorithm is used to establish a neural network model to identify phosphors. The result show that the final phosphor recognition rate reaches 99.9%. The innovative system based on LIBS and machine learning (ML) has the potential to improve rapid in situ detection of RE elements for the classification of e-waste.

Phosphorothioated DNA Engineered Liposomes as a General Platform for Stimuli-Responsive Cell-Specific Intracellular Delivery and Genome Editing.

Intracellular protein delivery is highly desirable for protein drug-based cell therapy. Established technologies suffer from poor cell-specific cytosolic protein delivery, which hampers the targeting therapy of specific cell populations. A fusogenic liposome system enables cytosolic delivery, but its ability of cell-specific and controllable delivery is quite limited. Inspired by the kinetics of viral fusion, we designed a phosphorothioated DNA coatings-modified fusogenic liposome to mimic the function of viral hemagglutinin. The macromolecular fusion machine docks cargo-loaded liposomes at the membrane of target cells, triggers membrane fusion upon pH or UV light stimuli, and facilitates cytosolic protein delivery. Our results showed efficient cell-targeted delivery of proteins of various sizes and charges, indicating the phosphorothioated DNA plug-in unit on liposomes could be a general strategy for spatial-temporally controllable protein delivery both in vitro and in vivo.

Real-time monitoring of carbon concentration using laser-induced breakdown spectroscopy and machine learning.

The spectral analysis based on laser-induced breakdown spectroscopy (LIBS) is an effective approach to carbon concentration monitoring. In this work, a novel LIBS-based method, together with a system designed independently, was developed for carbon monitoring. The experiments were conducted in two modes: static and dynamic. In static monitoring, gases in three scenarios were selected to represent different carbon concentrations, based on which measurements of carbon concentrations were performed through a mathematical model. Then, K-nearest Neighbors (KNN) was adopted for classification, and its accuracy could reach 99.17%, which can be applied for the identification of gas composition and pollution traceability. In dynamic monitoring, respiration and fossil fuel combustion were selected because of their important roles in increasing carbon concentration. In addition, the simulation of combustion degree was performed by the radial basis function (RBF) based on the spectral information, where the accuracy reached 96.41%, which is the first time that LIBS is proposed to be used for combustion prediction. The innovative approach derived from LIBS and machine learning algorithms is fast, online, and in-situ, showing far-reaching application prospects in real-time monitoring of carbon concentrations.

Changes in Cholesterol Level Alter Integrin Sequestration in Raft-Mimicking Lipid Mixtures.

The influence of cholesterol (CHOL) level on integrin sequestration in raft-mimicking lipid mixtures forming coexisting liquid-ordered (lo) and liquid-disordered (ld) lipid domains is investigated using complementary, single-molecule-sensitive, confocal detection methods. Systematic analysis of membrane protein distribution in such a model membrane environment demonstrates that variation of CHOL level has a profound influence on lo-ld sequestration of integrins, thereby exhibiting overall ld preference in the absence of ligands and lo affinity upon vitronectin addition. Accompanying photon-counting histogram analysis of integrins in the different model membrane mixtures shows that the observed changes of integrin sequestration in response to variations of membrane CHOL level are not associated with altering integrin oligomerization states. Instead, our experiments suggest that the strong CHOL dependence of integrin sequestration can be attributed to CHOL-mediated changes of lipid packing and bilayer thickness in coexisting lo and ld domains, highlighting the significance of a biophysical mechanism of CHOL-mediated regulation of integrin sequestration. We envision that this model membrane study may help clarify the influence of CHOL in integrin functionality in plasma membranes, thus providing further insight into the role of lipid heterogeneities in membrane protein distribution and function in a cellular membrane environment.

Ligand binding alters dimerization and sequestering of urokinase receptors in raft-mimicking lipid mixtures.

Lipid heterogeneities, such as lipid rafts, are widely considered to be important for the sequestering of membrane proteins in plasma membranes, thereby influencing membrane protein functionality. However, the underlying mechanisms of such sequestration processes remain elusive, in part, due to the small size and often transient nature of these functional membrane heterogeneities in cellular membranes. To overcome these challenges, here we report the sequestration behavior of urokinase receptor (uPAR), a glycosylphosphatidylinositol-anchored protein, in a planar model membrane platform with raft-mimicking lipid mixtures of well-defined compositions using a powerful optical imaging platform consisting of confocal spectroscopy XY-scans, photon counting histogram, and fluorescence correlation spectroscopy analyses. This methodology provides parallel information about receptor sequestration, oligomerization state, and lateral mobility with single molecule sensitivity. Most notably, our experiments demonstrate that moderate changes in uPAR sequestration are not only associated with modifications in uPAR dimerization levels, but may also be linked to ligand-mediated allosteric changes of these membrane receptors. Our data show that these modifications in uPAR sequestration can be induced by exposure to specific ligands (urokinase plasminogen activator, vitronectin), but not via adjustment of the cholesterol level in the planar model membrane system. Good agreement of our key findings with published results on cell membranes confirms the validity of our model membrane approach. We hypothesize that the observed mechanism of receptor translocation in the presence of raft-mimicking lipid mixtures is also applicable to other glycosylphosphatidylinositol-anchored proteins.

Magnetic ionic crystals with light controllable mobility and CO2 physisorption/desorption.

Magnetic responsive ionic liquid (MIL) demonstrated an advanced photomobility in confined narrow spaces through the doping of photoresponsive azobenzene by the interplay of supramolecular π-cations. Moreover, reversible physisorption/desorption of CO2 was achieved based on the photocontrolled solid-liquid transitions of the mixtures. Our approach opens opportunities to obtain multi-stimuli response through the coordinated supramolecular interplay of each responsive component.

Thermal Warning and Shut-down of Lithium Metal Batteries Based on Thermoresponsive Electrolytes.

The thermal runaway issue represents a long-standing obstacle that retards large-scale applications of lithium metal batteries. Various approaches to inhibit thermal runaway suffer from some intrinsic drawbacks, either being irreversible or delayed thermal protection. Herein, this work has explored thermo-responsive lower critical solution temperature (LCST) ionic liquid-based electrolytes, which provides reversible overheating protection for batteries with warning and shut-down stages, well corresponding to an initial stage of thermal runaway process. The batteries could function stably below 70 °C as a working mode, while demonstrating a warning mode above 80 °C with a noticeable reduction in specific capacitance to delay temperature increase of batteries. In terms of 110 °C as a critically dangerous temperature, a shut-down mode is designed to minimize the thermal energy releasing as the batteries are barely chargeable and dischargeable. Dynamically growing polymeric particles above LCST contributed to such an intelligent and mild control on specific capacitance. Larger size will occupy larger surfaces of electrodes and close more pores of separators, enabling a gradual suppressing of Li+ transfer and reactions. The present work demonstrated a scientific design of thermoresponsive LCST electrolytes with a superiorly precise and intelligent control of electrochemical performances to achieve self-adapted overheating protections.

SRRM2 phase separation drives assembly of nuclear speckle subcompartments.

Nuclear speckles (NSs) are nuclear biomolecular condensates that are postulated to form by macromolecular phase separation, although the detailed underlying forces driving NS formation remain elusive. SRRM2 and SON are 2 non-redundant scaffold proteins for NSs. How each individual protein governs assembly of the NS protein network and the functional relationship between SRRM2 and SON are largely unknown. Here, we uncover immiscible multiphases of SRRM2 and SON within NSs. SRRM2 and SON are functionally independent, specifically regulating alternative splicing of subsets of mRNA targets, respectively. We further show that SRRM2 forms multicomponent liquid phases in cells to drive NS subcompartmentalization, which is reliant on homotypic interaction and heterotypic non-selective protein-RNA complex coacervation-driven phase separation. SRRM2 serine/arginine-rich (RS) domains form higher-order oligomers and can be replaced by oligomerizable synthetic modules. The serine residues within the RS domains, however, play an irreplaceable role in fine-tuning the liquidity of NSs.

LCST ion gels fabricating "all-in-one" smart windows: thermotropic, electrochromic and power-generating.

Smart windows always respond to single stimuli, which cannot satisfy various needs in practical applications. Smart windows that integrate thermotropic, electrochromic and power-generating functions in one device is highly challenging yet important in satisfying on-demand light modulation and energy efficiency in practical applications. Herein, a thermoresponsive lower critical solution temperature (LCST) ion gel was fabricated via a facile in situ polymerization of butyl acrylate in a conventional ionic liquid to explore "all in one" smart windows. The ion gel-assembled smart windows are thermotropic and electrochromic with a reliable adjustment of light transparency as well as power-generating, enabled by the ionic Soret effect of ionic liquids. Additionally, the ion gels demonstrated self-defensive robust mechanical properties, thermal insulating and antifogging properties. With such an interdisciplinary and comprehensive study of the ion gels, the LCST ion gels could fulfil the requirements of genius windows with high energy-saving potential and exceptional climate adaptability, such as shut-down of light transmission in summer, daily solar energy collection, and colour changes on demand. It conceptually updates smart windows from an energy saving to an energy supplier in buildings. It is the first time to explore the "all in one" smart windows based on integrated multifunctional ionic liquids, which could greatly bridge the gap between the materials and buildings to accelerate practical applications of smart windows.

RNA nanotherapeutics with fibrosis overexpression and retention for MASH treatment.

Metabolic dysfunction-associated steatohepatitis (MASH) poses challenges for targeted delivery and retention of therapeutic proteins due to excess extracellular matrix (ECM). Here we present a new approach to treat MASH, termed "Fibrosis overexpression and retention (FORT)". In this strategy, we design (1) retinoid-derivative lipid nanoparticle (LNP) to enable enhanced mRNA overexpression in fibrotic regions, and (2) mRNA modifications which facilitate anchoring of therapeutic proteins in ECM. LNPs containing carboxyl-retinoids, rather than alcohol- or ester-retinoids, effectively deliver mRNA with over 10-fold enhancement of protein expression in fibrotic livers. The carboxyl-retinoid rearrangement on the LNP surface improves protein binding and membrane fusion. Therapeutic proteins are then engineered with an endogenous collagen-binding domain. These fusion proteins exhibit increased retention in fibrotic lesions and reduced systemic toxicity. In vivo, fibrosis-targeting LNPs encoding fusion proteins demonstrate superior therapeutic efficacy in three clinically relevant male-animal MASH models. This approach holds promise in fibrotic diseases unsuited for protein injection.

Bilayer asymmetry influences integrin sequestering in raft-mimicking lipid mixtures.

There is growing recognition that lipid heterogeneities in cellular membranes play an important role in the distribution and functionality of membrane proteins. However, the detection and characterization of such heterogeneities at the cellular level remains challenging. Here we report on the poorly understood relationship between lipid bilayer asymmetry and membrane protein sequestering in raft-mimicking model membrane mixtures using a powerful experimental platform comprised of confocal spectroscopy XY-scan and photon-counting histogram analyses. This experimental approach is utilized to probe the domain-specific sequestering and oligomerization state of αvß3 and α5ß1 integrins in bilayers, which contain coexisting liquid-disordered/liquid-ordered (ld/lo) phase regions exclusively in the top leaflet of the bilayer (bottom leaflet contains ld phase). Comparison with previously reported integrin sequestering data in bilayer-spanning lo-ld phase separations demonstrates that bilayer asymmetry has a profound influence on αvß3 and α5ß1 sequestering behavior. For example, both integrins sequester preferentially to the lo phase in asymmetric bilayers, but to the ld phase in their symmetric counterparts. Furthermore, our data show that bilayer asymmetry significantly influences the role of native ligands in integrin sequestering.

Chitosan Lactate Particles for Non-Compression Hemostasis on Hepatic Resection.

The liver is the most complex vascular anatomy of all human organs, with extremely rich blood flow and fragile texture. Massive liver bleeding usually occurs after traumatic liver injury, causing severe systematic issues. Thus, bleeding control is critical in hindering mortality rates and complications in patients. In this study, non-compression hemostasis materials based on chitosan lactate particles (CLP) were developed for handling liver bleeding after injuries. CLP showed good blood biocompatibility and antibacterial performance against S. aureus. Taking advantage of the vital capacity of CLP to promote red blood cell and platelet adhesion, CLP exhibited in vivo homeostasis properties as non-compression hemostasis materials for traumatic liver injury, both in SD rats, New Zealand rabbits, or in beagles. Whereas CLP has better hemostasis than the commercial hemostatic agent Celox™.

Time-resolved live-cell spectroscopy reveals EphA2 multimeric assembly.

Ephrin type-A receptor 2 (EphA2) is a receptor tyrosine kinase that initiates both ligand-dependent tumor-suppressive and ligand-independent oncogenic signaling. We used time-resolved, live-cell fluorescence spectroscopy to show that the ligand-free EphA2 assembles into multimers driven by two types of intermolecular interactions in the ectodomain. The first type entails extended symmetric interactions required for ligand-induced receptor clustering and tumor-suppressive signaling that inhibits activity of the oncogenic extracellular signal-regulated kinase (ERK) and protein kinase B (AKT) protein kinases and suppresses cell migration. The second type is an asymmetric interaction between the amino terminus and the membrane proximal domain of the neighboring receptors, which supports oncogenic signaling and promotes migration in vitro and tumor invasiveness in vivo. Our results identify the molecular interactions that drive the formation of the EphA2 multimeric signaling clusters and reveal the pivotal role of EphA2 assembly in dictating its opposing functions in oncogenesis.

In situ architecture of Opa1-dependent mitochondrial cristae remodeling.

Cristae membrane state plays a central role in regulating mitochondrial function and cellular metabolism. The protein Optic atrophy 1 (Opa1) is an important crista remodeler that exists as two forms in the mitochondrion, a membrane-anchored long form (l-Opa1) and a processed short form (s-Opa1). The mechanisms for how Opa1 influences cristae shape have remained unclear due to lack of native three-dimensional views of cristae. We perform in situ cryo-electron tomography of cryo-focused ion beam milled mouse embryonic fibroblasts with defined Opa1 states to understand how each form of Opa1 influences cristae architecture. In our tomograms, we observe a variety of cristae shapes with distinct trends dependent on s-Opa1:l-Opa1 balance. Increased l-Opa1 levels promote cristae stacking and elongated mitochondria while increased s-Opa1 levels correlated with irregular cristae packing and round mitochondria shape. Functional assays indicate a role for l-Opa1 in wild-type apoptotic and calcium handling responses, and compromised respiratory function under Opa1 imbalance. In summary, we provide three-dimensional visualization of cristae architecture to reveal relationships between mitochondrial ultrastructure and cellular function dependent on Opa1-mediated membrane remodeling.

Distilling Knowledge by Mimicking Features.

Knowledge distillation (KD) is a popular method to train efficient networks ("student") with the help of high-capacity networks ("teacher"). Traditional methods use the teacher's soft logits as extra supervision to train the student network. In this paper, we argue that it is more advantageous to make the student mimic the teacher's features in the penultimate layer. Not only the student can directly learn more effective information from the teacher feature, feature mimicking can also be applied for teachers trained without a softmax layer. Experiments show that it can achieve higher accuracy than traditional KD. To further facilitate feature mimicking, we decompose a feature vector into the magnitude and the direction. We argue that the teacher should give more freedom to the student feature's magnitude, and let the student pay more attention on mimicking the feature direction. To meet this requirement, we propose a loss term based on locality-sensitive hashing (LSH). With the help of this new loss, our method indeed mimics feature directions more accurately, relaxes constraints on feature magnitudes, and achieves state-of-the-art distillation accuracy. We provide theoretical analyses of how LSH facilitates feature direction mimicking, and further extend feature mimicking to multi-label recognition and object detection.

Corrigendum to "Polydopamine/poly(sulfobetaine methacrylate) Co-deposition coatings triggered by CuSO4/H2O2 on implants for improved surface hemocompatibility and antibacterial activity" [ Bioactive materials 6 (2021) 2546-2556].

[This corrects the article DOI: 10.1016/j.bioactmat.2021.01.025.].