Live birth of chimeric monkey with high contribution from embryonic stem cells.

A formal demonstration that mammalian pluripotent stem cells possess preimplantation embryonic cell-like (naive) pluripotency is the generation of chimeric animals through early embryo complementation with homologous cells. Whereas such naive pluripotency has been well demonstrated in rodents, poor chimerism has been achieved in other species including non-human primates due to the inability of the donor cells to match the developmental state of the host embryos. Here, we have systematically tested various culture conditions for establishing monkey naive embryonic stem cells and optimized the procedures for chimeric embryo culture. This approach generated an aborted fetus and a live chimeric monkey with high donor cell contribution. A stringent characterization pipeline demonstrated that donor cells efficiently (up to 90%) incorporated into various tissues (including the gonads and placenta) of the chimeric monkeys. Our results have major implications for the study of primate naive pluripotency and genetic engineering of non-human primates.

SMURF2 predisposes cancer cell toward ferroptosis in GPX4-independent manners by promoting GSTP1 degradation.

Ferroptosis is a non-apoptotic form of regulated cell death. Glutathione (GSH) peroxidase 4 (GPX4) and GSH-independent ferroptosis suppressor protein 1 (FSP1) have been identified as major defenses. Here, we uncover a protective mechanism mediated by GSH S-transferase P1 (GSTP1) by monitoring proteinomic dynamics during ferroptosis. Dramatic downregulation of GSTP1 is caused by SMURF2-mediated GSTP1 ubiquitination and degradation at early stages of ferroptosis. Intriguingly, GSTP1 acts in GPX4- and FSP1-independent manners by catalyzing GSH conjugation of 4-hydroxynonenal and detoxifying lipid hydroperoxides via selenium-independent GSH peroxidase activity. Genetic modulation of the SMURF2/GSTP1 axis or the pharmacological inhibition of GSTP1's catalytic activity sensitized tumor responses to Food and Drug Administration (FDA)-approved ferroptosis-inducing drugs both in vitro and in vivo. GSTP1 expression also confers resistance to immune checkpoint inhibitors by blunting ferroptosis. Collectively, these findings demonstrate a GPX4/FSP1-independent cellular defense mechanism against ferroptosis and suggest that targeting SMURF2/GSTP1 to sensitize cancer cells to ferroptosis has potential as an anticancer therapy.

Asymmetric hydrogenation of ketimines with minimally different alkyl groups.

Asymmetric catalysis enables the synthesis of optically active compounds, often requiring the differentiation between two substituents on prochiral substrates1. Despite decades of development of mainly noble metal catalysts, achieving differentiation between substituents with similar steric and electronic properties remains a notable challenge2,3. Here we introduce a class of Earth-abundant manganese catalysts for the asymmetric hydrogenation of dialkyl ketimines to give a range of chiral amine products. These catalysts distinguish between pairs of minimally differentiated alkyl groups bound to the ketimine, such as methyl and ethyl, and even subtler distinctions, such as ethyl and n-propyl. The degree of enantioselectivity can be adjusted by modifying the components of the chiral manganese catalyst. This reaction demonstrates a wide substrate scope and achieves a turnover number of up to 107,800. Our mechanistic studies indicate that exceptional stereoselectivity arises from the modular assembly of confined chiral catalysts and cooperative non-covalent interactions between the catalyst and the substrate.

Architecture of chloroplast TOC-TIC translocon supercomplex.

Chloroplasts rely on the translocon complexes in the outer and inner envelope membranes (the TOC and TIC complexes, respectively) to import thousands of different nuclear-encoded proteins from the cytosol1-4. Although previous studies indicated that the TOC and TIC complexes may assemble into larger supercomplexes5-7, the overall architectures of the TOC-TIC supercomplexes and the mechanism of preprotein translocation are unclear. Here we report the cryo-electron microscopy structure of the TOC-TIC supercomplex from Chlamydomonas reinhardtii. The major subunits of the TOC complex (Toc75, Toc90 and Toc34) and TIC complex (Tic214, Tic20, Tic100 and Tic56), three chloroplast translocon-associated proteins (Ctap3, Ctap4 and Ctap5) and three newly identified small inner-membrane proteins (Simp1-3) have been located in the supercomplex. As the largest protein, Tic214 traverses the inner membrane, the intermembrane space and the outer membrane, connecting the TOC complex with the TIC proteins. An inositol hexaphosphate molecule is located at the Tic214-Toc90 interface and stabilizes their assembly. Four lipid molecules are located within or above an inner-membrane funnel formed by Tic214, Tic20, Simp1 and Ctap5. Multiple potential pathways found in the TOC-TIC supercomplex may support translocation of different substrate preproteins into chloroplasts.

Novel flight style and light wings boost flight performance of tiny beetles.

Flight speed is positively correlated with body size in animals1. However, miniature featherwing beetles can fly at speeds and accelerations of insects three times their size2. Here we show that this performance results from a reduced wing mass and a previously unknown type of wing-motion cycle. Our experiment combines three-dimensional reconstructions of morphology and kinematics in one of the smallest insects, the beetle Paratuposa placentis (body length 395 µm). The flapping bristled wings follow a pronounced figure-of-eight loop that consists of subperpendicular up and down strokes followed by claps at stroke reversals above and below the body. The elytra act as inertial brakes that prevent excessive body oscillation. Computational analyses suggest functional decomposition of the wingbeat cycle into two power half strokes, which produce a large upward force, and two down-dragging recovery half strokes. In contrast to heavier membranous wings, the motion of bristled wings of the same size requires little inertial power. Muscle mechanical power requirements thus remain positive throughout the wingbeat cycle, making elastic energy storage obsolete. These adaptations help to explain how extremely small insects have preserved good aerial performance during miniaturization, one of the factors of their evolutionary success.

Rolling back human pluripotent stem cells to an eight-cell embryo-like stage.

After fertilization, the quiescent zygote experiences a burst of genome activation that initiates a short-lived totipotent state. Understanding the process of totipotency in human cells would have broad applications. However, in contrast to in mice1,2, demonstration of the time of zygotic genome activation or the eight-cell (8C) stage in in vitro cultured human cells has not yet been reported, and the study of embryos is limited by ethical and practical considerations. Here we describe a transgene-free, rapid and controllable method for producing 8C-like cells (8CLCs) from human pluripotent stem cells. Single-cell analysis identified key molecular events and gene networks associated with this conversion. Loss-of-function experiments identified fundamental roles for DPPA3, a master regulator of DNA methylation in oocytes3, and TPRX1, a eutherian totipotent cell homeobox (ETCHbox) family transcription factor that is absent in mice4. DPPA3 induces DNA demethylation throughout the 8CLC conversion process, whereas TPRX1 is a key executor of 8CLC gene networks. We further demonstrate that 8CLCs can produce embryonic and extraembryonic lineages in vitro or in vivo in the form of blastoids5 and complex teratomas. Our approach provides a resource to uncover the molecular process of early human embryogenesis.

Genome-wide analyses reveal the contribution of somatic variants to the immune landscape of multiple cancer types.

It has been well established that cancer cells can evade immune surveillance by mutating themselves. Understanding genetic alterations in cancer cells that contribute to immune regulation could lead to better immunotherapy patient stratification and identification of novel immune-oncology (IO) targets. In this report, we describe our effort of genome-wide association analyses across 22 TCGA cancer types to explore the associations between genetic alterations in cancer cells and 74 immune traits. Results showed that the tumor microenvironment (TME) is shaped by different gene mutations in different cancer types. Out of the key genes that drive multiple immune traits, top hit KEAP1 in lung adenocarcinoma (LUAD) was selected for validation. It was found that KEAP1 mutations can explain more than 10% of the variance for multiple immune traits in LUAD. Using public scRNA-seq data, further analysis confirmed that KEAP1 mutations activate the NRF2 pathway and promote a suppressive TME. The activation of the NRF2 pathway is negatively correlated with lower T cell infiltration and higher T cell exhaustion. Meanwhile, several immune check point genes, such as CD274 (PD-L1), are highly expressed in NRF2-activated cancer cells. By integrating multiple RNA-seq data, a NRF2 gene signature was curated, which predicts anti-PD1 therapy response better than CD274 gene alone in a mixed cohort of different subtypes of non-small cell lung cancer (NSCLC) including LUAD, highlighting the important role of KEAP1-NRF2 axis in shaping the TME in NSCLC. Finally, a list of overexpressed ligands in NRF2 pathway activated cancer cells were identified and could potentially be targeted for TME remodeling in LUAD.

Prohibitin 1 regulates mtDNA release and downstream inflammatory responses.

Exposure of mitochondrial DNA (mtDNA) to the cytosol activates innate immune responses. But the mechanisms by which mtDNA crosses the inner mitochondrial membrane are unknown. Here, we found that the inner mitochondrial membrane protein prohibitin 1 (PHB1) plays a critical role in mtDNA release by regulating permeability across the mitochondrial inner membrane. Loss of PHB1 results in alterations in mitochondrial integrity and function. PHB1-deficient macrophages, serum from myeloid-specific PHB1 KO (Phb1MyeKO) mice, and peripheral blood mononuclear cells from neonatal sepsis patients show increased interleukin-1ß (IL-1ß) levels. PHB1 KO mice are also intolerant of lipopolysaccharide shock. Phb1-depleted macrophages show increased cytoplasmic release of mtDNA and inflammatory responses. This process is suppressed by cyclosporine A and VBIT-4, which inhibit the mitochondrial permeability transition pore (mPTP) and VDAC oligomerization. Inflammatory stresses downregulate PHB1 expression levels in macrophages. Under normal physiological conditions, the inner mitochondrial membrane proteins, AFG3L2 and SPG7, are tethered to PHB1 to inhibit mPTP opening. Downregulation of PHB1 results in enhanced interaction between AFG3L2 and SPG7, mPTP opening, mtDNA release, and downstream inflammatory responses.

DSCAM gene triplication causes excessive GABAergic synapses in the neocortex in Down syndrome mouse models.

Down syndrome (DS) is caused by the trisomy of human chromosome 21 (HSA21). A major challenge in DS research is to identify the HSA21 genes that cause specific symptoms. Down syndrome cell adhesion molecule (DSCAM) is encoded by a HSA21 gene. Previous studies have shown that the protein level of the Drosophila homolog of DSCAM determines the size of presynaptic terminals. However, whether the triplication of DSCAM contributes to presynaptic development in DS remains unknown. Here, we show that DSCAM levels regulate GABAergic synapses formed on neocortical pyramidal neurons (PyNs). In the Ts65Dn mouse model for DS, where DSCAM is overexpressed due to DSCAM triplication, GABAergic innervation of PyNs by basket and chandelier interneurons is increased. Genetic normalization of DSCAM expression rescues the excessive GABAergic innervations and the increased inhibition of PyNs. Conversely, loss of DSCAM impairs GABAergic synapse development and function. These findings demonstrate excessive GABAergic innervation and synaptic transmission in the neocortex of DS mouse models and identify DSCAM overexpression as the cause. They also implicate dysregulated DSCAM levels as a potential pathogenic driver in related neurological disorders.

A new polymodal gating model of the proton-activated chloride channel.

The proton-activated chloride (PAC) channel plays critical roles in ischemic neuron death, but its activation mechanisms remain elusive. Here, we investigated the gating of PAC channels using its novel bifunctional modulator C77304. C77304 acted as a weak activator of the PAC channel, causing moderate activation by acting on its proton gating. However, at higher concentrations, C77304 acted as a weak inhibitor, suppressing channel activity. This dual function was achieved by interacting with 2 modulatory sites of the channel, each with different affinities and dependencies on the channel's state. Moreover, we discovered a protonation-independent voltage activation of the PAC channel that appears to operate through an ion-flux gating mechanism. Through scanning-mutagenesis and molecular dynamics simulation, we confirmed that E181, E257, and E261 in the human PAC channel serve as primary proton sensors, as their alanine mutations eliminated the channel's proton gating while sparing the voltage-dependent gating. This proton-sensing mechanism was conserved among orthologous PAC channels from different species. Collectively, our data unveils the polymodal gating and proton-sensing mechanisms in the PAC channel that may inspire potential drug development.

Light from Afield: Fast, High-Resolution, and Layer-Free Deep Vat 3D Printing.

Harnessing light for cross-linking of photoresponsive materials has revolutionized the field of 3D printing. A wide variety of techniques leveraging broad-spectrum light shaping have been introduced as a way to achieve fast and high-resolution printing, with applications ranging from simple prototypes to biomimetic engineered tissues for regenerative medicine. Conventional light-based printing techniques use cross-linking of material in a layer-by-layer fashion to produce complex parts. Only recently, new techniques have emerged which deploy multidirection, tomographic, light-sheet or filamented light-based image projections deep into the volume of resin-filled vat for photoinitiation and cross-linking. These Deep Vat printing (DVP) approaches alleviate the need for layer-wise printing and enable unprecedented fabrication speeds (within a few seconds) with high resolution (>10 µm). Here, we elucidate the physics and chemistry of these processes, their commonalities and differences, as well as their emerging applications in biomedical and non-biomedical fields. Importantly, we highlight their limitations, and future scope of research that will improve the scalability and applicability of these DVP techniques in a wide variety of engineering and regenerative medicine applications.

The moonlighting function of glycolytic enzyme enolase-1 promotes choline phospholipid metabolism and tumor cell proliferation.

Aberrantly upregulated choline phospholipid metabolism is a novel emerging hallmark of cancer, and choline kinase α (CHKα), a key enzyme for phosphatidylcholine production, is overexpressed in many types of human cancer through undefined mechanisms. Here, we demonstrate that the expression levels of the glycolytic enzyme enolase-1 (ENO1) are positively correlated with CHKα expression levels in human glioblastoma specimens and that ENO1 tightly governs CHKα expression via posttranslational regulation. Mechanistically, we reveal that both ENO1 and the ubiquitin E3 ligase TRIM25 are associated with CHKα. Highly expressed ENO1 in tumor cells binds to I199/F200 of CHKα, thereby abrogating the interaction between CHKα and TRIM25. This abrogation leads to the inhibition of TRIM25-mediated polyubiquitylation of CHKα at K195, increased stability of CHKα, enhanced choline metabolism in glioblastoma cells, and accelerated brain tumor growth. In addition, the expression levels of both ENO1 and CHKα are associated with poor prognosis in glioblastoma patients. These findings highlight a critical moonlighting function of ENO1 in choline phospholipid metabolism and provide unprecedented insight into the integrated regulation of cancer metabolism by crosstalk between glycolytic and lipidic enzymes.

Direct identification of A-to-I editing sites with nanopore native RNA sequencing.

Inosine is a prevalent RNA modification in animals and is formed when an adenosine is deaminated by the ADAR family of enzymes. Traditionally, inosines are identified indirectly as variants from Illumina RNA-sequencing data because they are interpreted as guanosines by cellular machineries. However, this indirect method performs poorly in protein-coding regions where exons are typically short, in non-model organisms with sparsely annotated single-nucleotide polymorphisms, or in disease contexts where unknown DNA mutations are pervasive. Here, we show that Oxford Nanopore direct RNA sequencing can be used to identify inosine-containing sites in native transcriptomes with high accuracy. We trained convolutional neural network models to distinguish inosine from adenosine and guanosine, and to estimate the modification rate at each editing site. Furthermore, we demonstrated their utility on the transcriptomes of human, mouse and Xenopus. Our approach expands the toolkit for studying adenosine-to-inosine editing and can be further extended to investigate other RNA modifications.

SAM-DTA: a sequence-agnostic model for drug-target binding affinity prediction.

Drug-target binding affinity prediction is a fundamental task for drug discovery and has been studied for decades. Most methods follow the canonical paradigm that processes the inputs of the protein (target) and the ligand (drug) separately and then combines them together. In this study we demonstrate, surprisingly, that a model is able to achieve even superior performance without access to any protein-sequence-related information. Instead, a protein is characterized completely by the ligands that it interacts. Specifically, we treat different proteins separately, which are jointly trained in a multi-head manner, so as to learn a robust and universal representation of ligands that is generalizable across proteins. Empirical evidences show that the novel paradigm outperforms its competitive sequence-based counterpart, with the Mean Squared Error (MSE) of 0.4261 versus 0.7612 and the R-Square of 0.7984 versus 0.6570 compared with DeepAffinity. We also investigate the transfer learning scenario where unseen proteins are encountered after the initial training, and the cross-dataset evaluation for prospective studies. The results reveals the robustness of the proposed model in generalizing to unseen proteins as well as in predicting future data. Source codes and data are available at https://github.com/huzqatpku/SAM-DTA.

EEG biomarkers of behavioral inhibition in patients with depression who committed violent offenses: a Go/NoGo ERP study.

Understanding the neurobiological correlates of behavioral inhibition in patients with depression who committed violent offenses could contribute to the prediction and prevention of violence. The present study recruited 29 depressed patients with violent offenses (VD group), 27 depressed patients without violent behavior (NVD group), and 28 healthy controls (HC group) to complete a visual Go/NoGo task, during which their responses and electroencephalography were simultaneously recorded using an event-related potentiometer. The results showed that the VD group made more commission errors and responded more slowly relative to the NVD and HC groups. The P3 amplitude of the VD group was reduced in the frontal and central brain regions compared to the HC group and increased in the parietal regions compared to the NVD group. In comparison to Go stimuli, NoGo stimuli induced longer P3 latencies in frontal regions in both the VD and NVD groups; however, this difference was not statistically significant in the HC group. These results provide electrophysical evidence of behavioral inhibition deficits in patients with depression, especially in those with violent behaviors. The reduced P3 amplitude in the frontal-central regions, increased P3 amplitude in the parietal regions, and increased NoGo P3 latency may be potential electrophysiological features that can predict violent behavior in patients with depression.

Obtaining Functional Proteomics Insights From Thermal Proteome Profiling Through Optimized Melt Shift Calculation and Statistical Analysis With InflectSSP.

Thermal proteome profiling (TPP) is an invaluable tool for functional proteomics studies that has been shown to discover changes associated with protein-ligand, protein-protein, and protein-RNA interaction dynamics along with changes in protein stability resulting from cellular signaling. The increasing number of reports employing this assay has not been met concomitantly with new approaches leading to advancements in the quality and sensitivity of the corresponding data analysis. The gap between data acquisition and data analysis tools is important to fill as TPP findings have reported subtle melt shift changes related to signaling events such as protein posttranslational modifications. In this study, we have improved the Inflect data analysis pipeline (now referred to as InflectSSP, available at https://CRAN.R-project.org/package=InflectSSP) to increase the sensitivity of detection for both large and subtle changes in the proteome as measured by TPP. Specifically, InflectSSP now has integrated statistical and bioinformatic functions to improve objective functional proteomics findings from the quantitative results obtained from TPP studies through increasing both the sensitivity and specificity of the data analysis pipeline. InflectSSP incorporates calculation of a "melt coefficient" into the pipeline with production of average melt curves for biological replicate studies to aid in identification of proteins with significant melts. To benchmark InflectSSP, we have reanalyzed two previously reported datasets to demonstrate the performance of our publicly available R-based program for TPP data analysis. We report new findings following temporal treatment of human cells with the small molecule thapsigargin that induces the unfolded protein response as a consequence of inhibition of sarcoplasmic/endoplasmic reticulum calcium ATPase 2A. InflectSSP analysis of our unfolded protein response study revealed highly reproducible and statistically significant target engagement over a time course of treatment while simultaneously providing new insights into the possible mechanisms of action of the small molecule thapsigargin.

Unifying the order and disorder dynamics in photoexcited VO2.

Photoinduced phase transition (PIPT) is always treated as a coherent process, but ultrafast disordering in PIPT is observed in recent experiments. Utilizing the real-time time-dependent density functional theory method, here we track the motion of individual vanadium (V) ions during PIPT in VO2 and uncover that their coherent or disordered dynamics can be manipulated by tuning the laser fluence. We find that the photoexcited holes generate a force on each V-V dimer to drive their collective coherent motion, in competing with the thermal-induced vibrations. If the laser fluence is so weak that the photoexcited hole density is too low to drive the phase transition alone, the PIPT is a disordered process due to the interference of thermal phonons. We also reveal that the photoexcited holes populated by the V-V dimerized bonding states will become saturated if the laser fluence is too strong, limiting the timescale of photoinduced phase transition.

Correction: Virus-mimicking nanosystems: from design to biomedical applications.

Correction for 'Virus-mimicking nanosystems: from design to biomedical applications' by Hao-Yang Liu et al., Chem. Soc. Rev., 2023, 52, 8481-8499, https://doi.org/10.1039/D3CS00138E.

One-Step Dual-Color Labeling of Viral Envelope and Intraviral Genome with Quantum Dots Harnessing Virus Infection.

Labeling the genome and envelope of a virus with multicolor quantum dots (QDs) simultaneously enables real-time monitoring of viral uncoating and genome release, contributing to our understanding of virus infection mechanisms. However, current labeling techniques require genetic modification, which alters the virus's composition and infectivity. To address this, we utilized the CRISPR/Cas13 system and a bioorthogonal metabolic method to label the Japanese encephalitis virus (JEV) genome and envelopes with different-colored QDs in situ. This technique allows one-step two-color labeling of the viral envelope and intraviral genome with QDs harnessing virus infection. In combination with single-virus tracking, we visualized JEV uncoating and genome release in real time near the endoplasmic reticulum of live cells. This labeling strategy allows for real-time visualization of uncoating and genome release at the single-virus level, and it is expected to advance the study of other viral infection mechanisms.

Single-cell sequencing reveals the impact of endothelial cell PIEZO1 expression on thoracic aortic aneurysm.

INTRODUCTION: Thoracic aortic aneurysm (TAA) is a severe vascular disease that threatens human life, characterized by focal dilatation of the entire aortic wall, with a diameter 1.5 times larger than normal. PIEZO1, a mechanosensitive cationic channel, monitors mechanical stimulations in the environment, transduces mechanical signals into electrical signals, and converts them into biological signals to activate intracellular signaling pathways. However, the role of PIEZO1 in TAA is still unclear. METHODS: We analyzed a single-cell database to investigate the expression level of PIEZO1 in TAA. We constructed a conditional knockout mouse model of Piezo1 and used the PIEZO1 agonist Yoda1 to intervene in the TAA model mice established by co-administration of BAPN and ANG-II. Finally, we explored the effect of Yoda1 on TAA in vitro. RESULTS AND DISCUSSION: We observed decreased PIEZO1 expression in TAA at both RNA and protein levels. Single-cell sequencing identified a specific reduction in Piezo1 expression in endothelial cells. Administration of PIEZO1 agonist Yoda1 prevented the formation of TAA. In PIEZO1 endothelial cell conditional knockout mice, Yoda1 inhibited TAA formation by interfering with PIEZO1. In vivo and in vitro experiments demonstrated that the effect of Yoda1 on endothelial cells involved macrophage infiltration, extracellular matrix degradation, and neovascularization. This study highlights the role of PIEZO1 in TAA and its potential as a therapeutic target, providing opportunities for clinical translation.