New geographical records and molecular investigation of the ciliate Spirostomum.

Spirostomum is a genus of large ciliates, and its species are distributed worldwide. However, there has been limited research conducted on their geographical distribution and genomics. We obtained nine samples of ciliates from eight regions in Liaoning Province, China, and conducted a study on their geographical distribution and characteristics. Morphological and second-generation high-throughput sequencing methods were applied to identify the species, and a phylogenetic tree was established to gain a deeper understanding of the geographical distribution and evolutionary relationships of Spirostomum in Northeast China. The results identified Spirostomum yagiui and Spirostomum subtilis as a newly recorded species in Northeast China region. There are now five species of Spirostomum that have been recorded in China, and new details on the genomic characteristics of Spirostomum yagiui were provided. In addition, this study also identified the main branches of Spirostomum teres and Spirostomum minus in northern China, and provided a theoretical basis for the existence of hidden species. Spirostomum yagiui is the first species in the family Spirostomidae to have undergone mitochondrial genome sequencing.

GOLEM: Automated and Robust Cryo-EM-Guided Ligand Docking with Explicit Water Molecules.

A detailed understanding of ligand-protein interaction is essential for developing rational drug-design strategies. In recent years, technological advances in cryo-electron microscopy (cryo-EM) brought a new era to the structural determination of biological macromolecules and assemblies at high resolution, marking cryo-EM as a promising tool for studying ligand-protein interactions. However, even in high-resolution cryo-EM results, the densities for the bound small-molecule ligands are often of lower quality due to their relatively dynamic and flexible nature, frustrating their accurate coordinate assignment. To address the challenge of ligand modeling in cryo-EM maps, here we report the development of GOLEM (Genetic Optimization of Ligands in Experimental Maps), an automated and robust ligand docking method that predicts a ligand's pose and conformation in cryo-EM maps. GOLEM employs a Lamarckian genetic algorithm to perform a hybrid global/local search for exploring the ligand's conformational, orientational, and positional space. As an important feature, GOLEM explicitly considers water molecules and places them at optimal positions and orientations. GOLEM takes into account both molecular energetics and the correlation with the cryo-EM maps in its scoring function to optimally place the ligand. We have validated GOLEM against multiple cryo-EM structures with a wide range of map resolutions and ligand types, returning ligand poses in excellent agreement with the densities. As a VMD plugin, GOLEM is free of charge and accessible to the community. With these features, GOLEM will provide a valuable tool for ligand modeling in cryo-EM efforts toward drug discovery.

In-situ direct seawater electrolysis using floating platform in ocean with uncontrollable wave motion.

Direct hydrogen production from inexhaustible seawater using abundant offshore wind power offers a promising pathway for achieving a sustainable energy industry and fuel economy. Various direct seawater electrolysis methods have been demonstrated to be effective at the laboratory scale. However, larger-scale in situ demonstrations that are completely free of corrosion and side reactions in fluctuating oceans are lacking. Here, fluctuating conditions of the ocean were considered for the first time, and seawater electrolysis in wave motion environment was achieved. We present the successful scaling of a floating seawater electrolysis system that employed wind power in Xinghua Bay and the integration of a 1.2 Nm3 h-1-scale pilot system. Stable electrolysis operation was achieved for over 240 h with an electrolytic energy consumption of 5 kWh Nm-3 H2 and a high purity (>99.9%) of hydrogen under fluctuating ocean conditions (0~0.9 m wave height, 0~15 m s-1 wind speed), which is comparable to that during onshore water electrolysis. The concentration of impurity ions in the electrolyte was low and stable over a long period of time under complex and changing scenarios. We identified the technological challenges and performances of the key system components and examined the future outlook for this emerging technology.

Autophagy counters inflammation-driven glycolytic impairment in aging hematopoietic stem cells.

Autophagy is central to the benefits of longevity signaling programs and to hematopoietic stem cell (HSC) response to nutrient stress. With age, a subset of HSCs increases autophagy flux and preserves regenerative capacity, but the signals triggering autophagy and maintaining the functionality of autophagy-activated old HSCs (oHSCs) remain unknown. Here, we demonstrate that autophagy is an adaptive cytoprotective response to chronic inflammation in the aging murine bone marrow (BM) niche. We find that inflammation impairs glucose uptake and suppresses glycolysis in oHSCs through Socs3-mediated inhibition of AKT/FoxO-dependent signaling, with inflammation-mediated autophagy engagement preserving functional quiescence by enabling metabolic adaptation to glycolytic impairment. Moreover, we show that transient autophagy induction via a short-term fasting/refeeding paradigm normalizes glycolytic flux and significantly boosts oHSC regenerative potential. Our results identify inflammation-driven glucose hypometabolism as a key driver of HSC dysfunction with age and establish autophagy as a targetable node to reset oHSC regenerative capacity.

From bottom-up to cell surface proteomics: detergents or no detergents, that is the question.

Measuring the expression levels of membrane proteins (MPs) is crucial for understanding cell differentiation and tissue specificity, defining disease characteristics, identifying biomarkers, and developing therapeutics. While bottom-up proteomics addresses the need for accurately surveying the membrane proteome, the lower abundance and hydrophobic nature of MPs pose challenges in sample preparation. As MPs normally reside in the lipid bilayer, conventional extraction methods rely on detergents, introducing here a paradox - detergents prevent aggregation and facilitate protein processing, but themselves become contaminants that interfere with downstream analytical applications. Various detergent removal methods exist to mitigate this issue, including filter-aided sample preparation, SP3, suspension trapping, and membrane mimetics. This review delves into the fundamentals of each strategy, applications, merits, and limitations, providing insights into their effectiveness in MP research.

Capture of endogenous lipids in peptidiscs and effect on protein stability and activity.

Compared to protein-protein and protein-nucleic acid interactions, our knowledge of protein-lipid interactions remains limited. This is primarily due to the inherent insolubility of membrane proteins (MPs) in aqueous solution. The traditional use of detergents to overcome the solubility barrier destabilizes MPs and strips away certain lipids that are increasingly recognized as crucial for protein function. Recently, membrane mimetics have been developed to circumvent the limitations. In this study, using the peptidisc, we find that MPs in different lipid states can be isolated based on protein purification and reconstitution methods, leading to observable effects on MP activity and stability. Peptidisc also enables re-incorporating specific lipids to fine-tune the protein microenvironment and assess the impact on downstream protein associations. This study offers a first look at the illusive protein-lipid interaction specificity, laying the path for a systematic evaluation of lipid identity and contributions to membrane protein function.

Ascorbate depletion increases quiescence and self-renewal potential in hematopoietic stem cells and multipotent progenitors.

Ascorbate (vitamin C) limits hematopoietic stem cell (HSC) function and suppresses leukemia development by promoting the function of the Tet2 tumor suppressor. In humans, ascorbate is obtained from the diet while in mice it is synthesized in the liver. In this study, we show that deletion of the Slc23a2 ascorbate transporter severely depleted ascorbate from hematopoietic cells. Slc23a2 deficiency increased HSC reconstituting potential and self-renewal potential upon transplantation into irradiated mice. Slc23a2 deficiency also increased the reconstituting and self-renewal potential of multipotent hematopoietic progenitors (MPPs), conferring the ability to long-term reconstitute irradiated mice. Slc23a2-deficient HSCs and MPPs divided much less frequently than control HSCs and MPPs. Increased self-renewal and reconstituting potential were observed particularly in quiescent Slc23a2-deficient HSCs and MPPs. The effect of Slc23a2 deficiency on MPP self-renewal was not mediated by reduced Tet2 function. Ascorbate thus regulates quiescence and restricts self-renewal potential in HSCs and MPPs such that ascorbate depletion confers MPPs with long-term self-renewal potential.

Fault-tolerant Hamiltonian cycle strategy for fast node fault diagnosis based on PMC in data center networks.

System-level fault diagnosis model, namely, the PMC model, detects fault nodes only through the mutual testing of nodes in the system without physical equipment. In order to achieve server nodes fault diagnosis in large-scale data center networks (DCNs), the traditional algorithm based on the PMC model cannot meet the characteristics of high diagnosability, high accuracy and high efficiency due to its inability to ensure that the test nodes are fault-free. This paper first proposed a fault-tolerant Hamiltonian cycle fault diagnosis (FHFD) algorithm, which tests nodes in the order of the Hamiltonian cycle to ensure that the test nodes are faultless. In order to improve testing efficiency, a hierarchical diagnosis mechanism was further proposed, which recursively divides high scale structures into a large number of low scale structures based on the recursive structure characteristics of DCNs. Additionally, we proved that $ 2(n-2){n^{k-1}} $ and $ (n-2){t_{n, k}}/{t_{n, 1}} $ faulty nodes could be detected for $ BCub{e_{n, k}} $ and $ DCel{l_{n, k}} $ within a limited time for the proposed diagnosis strategy. Simulation experiments have also shown that our proposed strategy has improved the diagnosability and test efficiency dramatically.

Affinity Purification of Membrane ß-Barrel Proteins via Biotin-Tagged Peptidiscs.

ß-barrel membrane proteins play a crucial role in bacterial pathogenesis and antibiotic resistance, making them a prime focus for the development of new antibiotics and therapeutics. However, their inherent hydrophobic nature and limited presence pose challenges for their high-throughput characterization using conventional methods. In this context, we present a simple but efficacious approach using peptidisc, a membrane mimetic, to overcome the low abundance and hydrophobicity of these proteins. Our methodology, illustrated here using Escherichia coli (E. coli) as a model organism, covers the entire process from outer membrane fraction preparation to data analysis. This detailed protocol outlines the purification of a diverse collection of ß-barrel membrane proteins, rendering them water-soluble and readily amenable to mass spectrometry and downstream drug screening strategies.

Capture of the Mouse Organ Membrane Proteome Specificity in Peptidisc Libraries.

Membrane proteins, particularly those on the cell surface, play pivotal roles in diverse physiological processes, and their dysfunction is linked to a broad spectrum of diseases. Despite being crucial biomarkers and therapeutic drug targets, their low abundance and hydrophobic nature pose challenges in isolation and quantification, especially when extracted from tissues and organs. To overcome these hurdles, we developed the membrane-mimicking peptidisc, enabling the isolation of the membrane proteome in a water-soluble library conducive to swift identification through liquid chromatography with tandem mass spectrometry. This study applies the method across five mice organs, capturing between 200 and 450 plasma membrane proteins in each case. More than just membrane protein identification, the peptidisc is used to estimate the relative abundance across organs, linking cell-surface protein molecular functions to organ biological roles, thereby contributing to the ongoing discourse on organ specificity. This contribution holds substantial potential for unveiling new avenues in the exploration of biomarkers and downstream applications involving knowledge of the organ cell-surface proteome.

Architecture of Vanadium-Based MXene Dysregulating Tumor Redox Homeostasis for Amplified Nanozyme Catalytic/Photothermal Therapy.

Taking the significance of the special microenvironment for tumor cell survival into account, disrupting tumor redox homeostasis is highly prospective for improving therapeutic efficacy. Herein, a multifunctional 2D vanadium-based MXene nanoplatform, V4 C3 /atovaquone@bovine albumin (V4 C3 /ATO@BSA, abbreviated as VAB) has been elaborately constructed for ATO-enhanced nanozyme catalytic/photothermal therapy. The redox homeostasis within the tumor cells is eventually disrupted, showing a remarkable anti-tumor effect. The VAB nanoplatform with mixed vanadium valence states can induce a cascade of catalyzed reactions in the tumor microenvironment, generating plenty of reactive oxygen species (ROS) with effective glutathione consumption to amplify oxidative stress. Meanwhile, the stable and strong photothermal effect of VAB under near-infrared irradiation not only causes the necrosis of tumor cells, but also improves its peroxidase-like activity. In addition, the release of ATO can effectively alleviate endogenous oxygen consumption to limit triphosadenine formation and inhibit mitochondrial respiration. As a result, the expression of heat shock proteins is effectively suppressed to overcome thermoresistance and the production of ROS can be further promoted due to mitochondrial injury. Moreover, VAB also presents high photoacoustic and photothermal imaging performances. In brief, the multifunctional nanoplatform can provide ATO-enhanced nanozyme catalytic/photothermal therapy with broadening the biomedical applications of vanadium-based MXene.

Electrochemistry detection of estrogenic effect: Regulation of de novo purine synthesis and catabolism by gibberellin and fulvestrant.

The estrogenic effect of plant growth regulators has been received little attention, which leads to the lack of relevant toxicity data. In this study, the estrogenic effect induced by gibberellin with ERα-dependent manner was found by E-screen and western blot methods, and the electrochemical signals of MCF-7 cells regulated by gibberellin and fulvestrant were investigated. The results showed that the electrochemical signals of MCF-7 cells were increased by gibberellin, while reduced by fulvestrant significantly, and displayed an extremely sensitive response to the effects of estrogenic effect induced by ERα agonist and antagonist. Western blot results showed that the expressions of phosphoribosyl pyrophosphate amidotransferase and hypoxanthine nucleotide dehydrogenase in de novo purine synthesis and adenine deaminase in catabolism were more effective regulated by gibberellin and fulvestrant, resulting in significant changes of the levels of guanine, hypoxanthine and xanthine in cells, and then electrochemical signals. The results provide a theoretical basis for the establishment of new electrochemical detection method of the estrogenic effect of plant regulators.

Mitochondria-targeting Cu3VS4 nanostructure with high copper ionic mobility for photothermoelectric therapy.

Thermoelectric therapy has emerged as a promising treatment strategy for oncology, but it is still limited by the low thermoelectric catalytic efficiency at human body temperature and the inevitable tumor thermotolerance. We present a photothermoelectric therapy (PTET) strategy based on triphenylphosphine-functionalized Cu3VS4 nanoparticles (CVS NPs) with high copper ionic mobility at room temperature. Under near-infrared laser irradiation, CVS NPs not only generate hyperthermia to ablate tumor cells but also catalytically yield superoxide radicals and induce endogenous NADH oxidation through the Seebeck effect. Notably, CVS NPs can accumulate inside mitochondria and deplete NADH, reducing ATP synthesis by competitively inhibiting the function of complex I, thereby down-regulating the expression of heat shock proteins to relieve tumor thermotolerance. Both in vitro and in vivo results show notable tumor suppression efficacy, indicating that the concept of integrating PTET and mitochondrial metabolism modulation is highly feasible and offers a translational promise for realizing precise and efficient cancer treatment.

Leptin receptor+ cells promote bone marrow innervation and regeneration by synthesizing nerve growth factor.

The bone marrow contains peripheral nerves that promote haematopoietic regeneration after irradiation or chemotherapy (myeloablation), but little is known about how this is regulated. Here we found that nerve growth factor (NGF) produced by leptin receptor-expressing (LepR+) stromal cells is required to maintain nerve fibres in adult bone marrow. In nerveless bone marrow, steady-state haematopoiesis was normal but haematopoietic and vascular regeneration were impaired after myeloablation. LepR+ cells, and the adipocytes they gave rise to, increased NGF production after myeloablation, promoting nerve sprouting in the bone marrow and haematopoietic and vascular regeneration. Nerves promoted regeneration by activating ß2 and ß3 adrenergic receptor signalling in LepR+ cells, and potentially in adipocytes, increasing their production of multiple haematopoietic and vascular regeneration growth factors. Peripheral nerves and LepR+ cells thus promote bone marrow regeneration through a reciprocal relationship in which LepR+ cells sustain nerves by synthesizing NGF and nerves increase regeneration by promoting the production of growth factors by LepR+ cells.

Design and Mechanism Insight of Monodispersed AuCuPt Alloy Nanozyme with Antitumor Activity.

The abrogation of the self-adaptive redox evolution of tumors is promising for improving therapeutic outcomes. In this study, we designed a trimetallic alloy nanozyme AuCuPt-PpIX (ACPP), which mimics up to five naturally occurring enzymes: glucose oxidase (GOD), superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and glutathione peroxidase (GPx). Facilitated by these enzyme-mimicking traits, the constructed ACPP nanozymes can not only disrupt the established redox homeostasis in tumors through a series of enzymatic cascade reactions but also achieve cyclic regeneration of the relevant enzyme substrates. Density functional theory (DFT) calculations have theoretically explained the synergistic effect of multimetallic doping and the possible mechanism of enzymatic catalysis. The doped Cu and Pt sites are conducive to the adsorption, activation, and dissociation of reactant molecules, whereas the Au sites are conducive to desorption, which significantly improves catalytic efficiency via a synergistic effect. Additionally, ACPP nanozymes can improve the effect of protoporphyrin (PpIX)-enabled sonodynamic therapy (SDT) by alleviating hypoxia and initiating ferroptosis by inducing lipid peroxidation (LPO) and inhibiting GPX4 activity, thus achieving multimodal synergistic therapy. This study presents a typical paradigm to enable the use of multimetallic alloy nanozymes for the treatment of tumor cells with self-adaptive properties.

Piezoelectric enhanced sulfur doped graphdiyne nanozymes for synergistic ferroptosis-apoptosis anticancer therapy.

Graphdiyne has excellent potential due to its enzymatic properties. Metal-free sulfur-doped Graphdiyne (S-GDY) has piezoelectric characteristics, and ultrasonic excitation of S-GDY enhances peroxidase activity. It can turn hydrogen peroxide into toxic hydroxyl radicals and induce apoptosis in 4T1 cells. More importantly, the ultrasound (US) enhanced nanozyme induced 4T1 cell ferroptosis by promoting an imbalanced redox reaction due to glutathione depletion and glutathione peroxidase 4 inactivation. S-GDY exhibited enhanced nanozyme activity in vitro and in vivo that may directly trigger apoptosis-ferroptosis for effective tumor therapy. Altogether, this study was expected to provide new insights into the design of piezoelectric catalytic nanozyme and expand their application in the catalytic therapy of tumors.

Metabolite profiling identifies chemical markers associated with the cytotoxic properties of roasted fermented avocado seeds.

Studies have demonstrated avocado seeds are a good source of bioactive compounds. This study investigated the effects of roasting on the metabolites and anticancer activities of fermented avocado seeds. All three anti-cancer activities of fermented avocado seeds were higher at lower roasting temperature and time. The best inhibition effect was found against Hep G2 followed by the MDA-MB-231 and MCF-7 cancer cell lines. Untargeted metabolite profiling using gas chromatography-mass spectrometry resulted in identification of 208 metabolites. In total, 41 metabolites identified had VIP values more than 1 using PLS-R that were related to anticancer activities. All amino acids and most sugars were higher at lower roasting temperature and positively correlated to anticancer activity. The roasting conditions for optimal antioxidant and anticancer activities were determined to be 121 °C for 9 min. Findings showed that fermented avocado seed powder has the potential to become a functional food ingredient with beneficial bioctive properties.

Pathogenic mitochondrial DNA mutations inhibit melanoma metastasis.

Mitochondrial DNA (mtDNA) mutations are frequently observed in cancer, but their contribution to tumor progression is controversial. To evaluate the impact of mtDNA variants on tumor growth and metastasis, we created human melanoma cytoplasmic hybrid (cybrid) cell lines transplanted with wildtype mtDNA or pathogenic mtDNA encoding variants that partially or completely inhibit oxidative phosphorylation. Homoplasmic pathogenic mtDNA cybrids reliably established tumors despite dysfunctional oxidative phosphorylation. However, pathogenic mtDNA variants disrupted spontaneous metastasis of subcutaneous tumors and decreased the abundance of circulating melanoma cells in the blood. Pathogenic mtDNA did not induce anoikis or inhibit organ colonization of melanoma cells following intravenous injections. Instead, migration and invasion were reduced, indicating that limited circulation entry functions as a metastatic bottleneck amidst mtDNA dysfunction. Furthermore, analysis of selective pressure exerted on the mitochondrial genomes of heteroplasmic cybrid lines revealed a suppression of pathogenic mtDNA allelic frequency during melanoma growth. Collectively, these findings demonstrate that functional mtDNA is favored during melanoma growth and enables metastatic entry into the blood.

Sonodynamic Therapy of NRP2 Monoclonal Antibody-Guided MOFs@COF Targeted Disruption of Mitochondrial and Endoplasmic Reticulum Homeostasis to Induce Autophagy-Dependent Ferroptosis.

The lethality and chemotherapy resistance of pancreatic cancer necessitates the urgent development of innovative strategies to improve patient outcomes. To address this issue, we designed a novel drug delivery system named GDMCN2,which uses iron-based metal organic framework (Fe-MOF) nanocages encased in a covalent organic framework (COF) and modified with the pancreatic cancer-specific antibody, NRP2. After being targeted into tumor cells, GDMCN2 gradually release the sonosensitizer sinoporphyrin sodium (DVDMS) and chemotherapeutic gemcitabine (GEM) and simultaneously generated reactive oxygen species (ROS) under ultrasound (US) irradiation. This system can overcome gemcitabine resistance in pancreatic cancer and reduce its toxicity to non-targeted cells and tissues. In a mechanistic cascade, the release of ROS activates the mitochondrial transition pore (MPTP), leading to the release of Ca2+ and induction of endoplasmic reticulum (ER) stress. Therefore, microtubule-associated protein 1A/1B-light chain 3 (LC3) is activated, promoting lysosomal autophagy. This process also induces autophagy-dependent ferroptosis, aided by the upregulation of Nuclear Receptor Coactivator 4 (NCOA4). This mechanism increases the sensitivity of pancreatic cancer cells to chemotherapeutic drugs and increases mitochondrial and DNA damage. The findings demonstrate the potential of GDMCN2 nanocages as a new avenue for the development of cancer therapeutics.

Autophagy counters inflammation-driven glycolytic impairment in aging hematopoietic stem cells.

Aging of the hematopoietic system promotes various blood, immune and systemic disorders and is largely driven by hematopoietic stem cell (HSC) dysfunction ( 1 ). Autophagy is central for the benefits associated with activation of longevity signaling programs ( 2 ), and for HSC function and response to nutrient stress ( 3,4 ). With age, a subset of HSCs increases autophagy flux and preserves some regenerative capacity, while the rest fail to engage autophagy and become metabolically overactivated and dysfunctional ( 4 ). However, the signals that promote autophagy in old HSCs and the mechanisms responsible for the increased regenerative potential of autophagy-activated old HSCs remain unknown. Here, we demonstrate that autophagy activation is an adaptive survival response to chronic inflammation in the aging bone marrow (BM) niche ( 5 ). We find that inflammation impairs glucose metabolism and suppresses glycolysis in aged HSCs through Socs3-mediated impairment of AKT/FoxO-dependent signaling. In this context, we show that inflammation-mediated autophagy engagement preserves functional quiescence by enabling metabolic adaptation to glycolytic impairment. Moreover, we demonstrate that transient autophagy induction via a short-term fasting/refeeding paradigm normalizes glucose uptake and glycolytic flux and significantly improves old HSC regenerative potential. Our results identify inflammation-driven glucose hypometabolism as a key driver of HSC dysfunction with age and establish autophagy as a targetable node to reset old HSC glycolytic and regenerative capacity. One-Sentence Summary: Autophagy compensates for chronic inflammation-induced metabolic deregulation in old HSCs, and its transient modulation can reset old HSC glycolytic and regenerative capacity.