Localized protein translation occurs in numerous subcellular compartments and regulates diverse biological processes by rapidly changing protein compositions in response to subcellular needs. Existing assays for subcellular local translation either require physical isolation, which is prone to contamination and loss of material, or imaging-based readout, which is often hampered with low throughput. In this study, we report the development of the optoRibo-seq method that features photoactivatable enzyme-mediated proximity labeling of ribosomes in genetically specified subcellular locations. We demonstrate the spatial specificity of optoRibo-seq at the endoplasmic reticulum (ER) membrane, with a temporal resolution of 1 min. In cells undergoing chemically induced ER stress, optoRibo-seq allowed mapping of the dynamic changes in the ER-proximal translatome, identifying transcripts involved in protein folding and targeting. Our strategy provides a general platform for spatiotemporally resolved profiling of subcellular protein translation.

Cuproptosis, a copper-induced form of regulated cell death, holds therapeutic promise in cancer but remains mechanistically unclear. We developed Mito-TPCA, a mitochondrial thermal proximity coaggregation strategy combining enzyme-catalyzed proteome labeling with thermal profiling, to map mitochondrial protein-protein interaction dynamics during cuproptosis. This approach revealed that copper disrupts the association of pyruvate dehydrogenase kinases (PDKs) with the pyruvate dehydrogenase (PDH) complex by targeting lipoyl domains, triggering PDH dephosphorylation and aberrant activation. We demonstrate that this PDH activation is a key driver of cuproptosis and contributes to the heightened susceptibility of cancer cells. These findings establish PDH dephosphorylation/activation as a central mechanism of cuproptosis and a potential anti-cancer therapeutic target. Mito-TPCA offers a versatile platform to study mitochondrial protein complex dynamics in live cells.

Prohibitin 1 (PHB1) and Prohibitin 2 (PHB2), two conserved prohibitin members, are primarily localized to the mitochondrial inner membrane (MIM) to form a nanoscale macromolecular prohibitin complex. This prohibitin complex can facilitate the spatial organization of proteins and lipids, thus maintaining cellular metabolism and homeostasis, but its architecture remains largely unknown. Here, we report the cryo-EM structure of a prohibitin complex at 2.8 Å resolution, which contains 11 PHB1-PHB2 heterodimers. This complex displays a bell-like cage, consisting of a lid and a wall, which creates an intermembrane space-facing compartment for the MIM. The lid of the cage is stably assembled, and it is responsible for the prohibitin complex formation. In contrast, the wall of the cage is flexible and exhibits lateral openings, providing a channel for intramembrane exchange of proteins and lipids. These findings provide a structural basis for understanding the scaffold role of the prohibitin complex in organizing intramembrane proteins and lipids.

XPR1 is the sole protein known to transport inorganic phosphate (Pi) out of cells, a function conserved across species from yeast to mammals. Human XPR1 variants lead to cerebral calcium-phosphate deposition and primary familial brain calcification (PFBC), a hereditary neurodegenerative disorder. Here, we present the cryo-EM structure of human XPR1 in both its Pi-unbound and various Pi-bound states. XPR1 features 10 transmembrane α-helices forming an ion channel-like structure, with multiple Pi recognition sites along the channel. Pathogenic mutations in two arginine residues, which line the translocation channel, disrupt Pi transport. Molecular dynamics simulations reveal that Pi ion undergoes a stepwise transition through the sequential recognition sites during the transport process. Together with functional analyses, our results suggest that this sequential arrangement allows XPR1 to facilitate Pi ion passage via a “relay” process, and they establish a framework for the interpretation of disease-related mutations and for the development of future therapeutics. XPR1 is the only known protein that exports inorganic phosphate from cells. This study demonstrates that XPR1 uses a channel-like mechanism for phosphate transport, offering insights into potential therapeutic strategies for XPR1-linked diseases.

Mapping the spatial organization of proteins and cellular interactions is crucial for understanding their biological functions. Herein, we report a biocompatible, multi-functional luminescence-activated proximity beling (LAP) strategy for profiling subcellular proteomes and cell-cell interactions in live cells and animals. Our method capitalizes on fusing the photocatalyst miniSOG to NanoLuc luciferase, whose bioluminescence activates miniSOG via a resonance energy transfer mechanism, generating reactive oxygen species in situ mediate proximity labeling (PL). We demonstrated the high spatial specificity of LAP in a C57BL6/N mouse model transplanted with MC38 cells. Our data revealed tumor microenvironment-dependent remodeling secretome. LAP was further applied to identify ligand-receptor-mediated cell-cell interactions both in vitro and in vivo. We also achieved local transcriptome profiling by combining LAP with next-generation sequencing. Overall, LAP was proved to be a versatile PL technique with strong biocompatibility for spatial multi-omic applications.

Frizzled receptors (FZDs) are key contributors intrinsic to the Wnt signaling pathway, activation of FZDs triggering the Wnt signaling cascade is frequently observed in human tumors and intimately associated with an aggressive carcinoma phenotype. It has been shown that the abnormal expression of FZD receptors contributes to the manifestation of malignant characteristics in human tumors such as enhanced cell proliferation, metastasis, chemotherapy resistance as well as the acquisition of cancer stemness. Given the essential roles of FZD receptors in the Wnt signaling in human tumors, this review aims to consolidate the prevailing knowledge on the specific status of FZD receptors (FZD1-10) and elucidate their respective functions in tumor progression. Furthermore, we delineate the structural basis for binding of FZD and its co-receptors to Wnt, and provide a better theoretical foundation for subsequent studies on related mechanisms. Finally, we describe the existing biological classes of small molecule-based FZD inhibitors in detail in the hope that they can provide useful assistance for design and development of novel drug candidates targeted FZDs.

This article investigates the cultural politics of the Beijing subway. Drawing on diverse sources, we trace the evolution of the subway over the last half-century to reveal that it transcends its fluctuating, time-specific practicalities to serve as a potent conduit through which the Chinese state consistently shapes subjecthood. The article begins with the subway's Cold War inception as a military enterprise, spotlighting its deliberate concealment to safeguard the echelons of power and obscure both international and domestic tensions. The second section delves into the subway's rebirth in the wake of China's opening-up reform and rapid economic rise, as it transforms into a mobile gallery of political aesthetics that extols China's cultural heritage and triumphs, cultivating national pride under siege from unleashed market and social forces. The final section dissects the subway's orchestration of undesirable passengers, sculpting a socioeconomic hierarchy in the city's commuting system. As a multifaceted prism, the Beijing subway encapsulates a range of covert and overt, pragmatic and aesthetic, and inclusive and exclusive elements in the cultural politics of Chinese infrastructure at large; and it illustrates the sustained centrality of state power in shaping individual subjectivities and defining the cultural and representational significance of Chinese infrastructure, albeit amid growing contestation.

Cellular context is crucial for understanding the complex and dynamic kinase functions in health and disease. Systematic dissection of kinase-mediated cellular processes requires rapid and precise stimulation (‘pulse’) of a kinase of interest, as well as global and in-depth characterization (‘chase’) of the perturbed proteome under living conditions. Here we developed an optogenetic ‘pulse-chase’ strategy, termed decaging kinase coupled proteomics (DeKinomics), for proteome-wide profiling of kinase-driven phosphorylation at second-timescale in living cells. We took advantage of the ‘gain-of-function’ feature of DeKinomics to identify direct kinase substrates and further portrayed the global phosphorylation of understudied receptor tyrosine kinases under native cellular settings. DeKinomics offered a general activation-based strategy to study kinase functions with high specificity and temporal resolution under living conditions.

The full text of this preprint has been withdrawn by the authors while they make corrections to the work. Therefore, the authors do not wish this work to be cited as a reference. Questions should be directed to the corresponding author.

Civic organizations have long been heralded for promoting voluntarism and civic participation, yet observational studies, facing endogeneity concerns, have struggled to solidify the causal underpinnings of the neo-Tocquevillian theory. Furthermore, empirical examination of their recruitment efficacy relative to state entities, particularly in crises or authoritarian settings, remains scarce. Our survey experiment in COVID-period China provides a twofold crucial case study. By embodying both the "most likely case" of effective stateled mobilization and the "least likely case" of potent civic organization efforts, it offers unique analytical leverage and insights. Civic appeals significantly boosted volunteer inclinations, whereas governmental outreach had minimal or even negative effects. This civic boost was stronger among those with less social and political trust and channeled by heightened political efficacy and active citizenship norms. Our findings shed light on the pivotal roles of civic organizations, even in authoritarian settings featuring embryonic civil societies.

BAK permeabilizes the mitochondrial outer membrane, causing apoptosis. This apoptotic activity of BAK is stimulated by binding prodeath activators within its canonical hydrophobic groove. Parkin, an E3 ubiquitin (Ub) ligase, can ubiquitinate BAK, which inhibits BAK apoptotic activity. However, the molecular mechanism underlying the inhibition of ubiquitination remains structurally uncharacterized. Here, we utilize truncated and soluble BAK to construct a mimetic of K113-ubiquitinated BAK (disulfide-linked Ub G76C ~ BAK K113C ) and further present its NMR-derived structure model. The classical L8-I44-H68-V70 hydrophobic patch of the conjugated Ub subunit binds within the canonical hydrophobic groove of BAK. This Ub occludes the binding of prodeath BID activators in the groove and impairs BID-triggered BAK activation and membrane permeabilization. Reduced interaction between Ub and BAK subunits allows BID to activate K113-ubiquitinated BAK. These mechanistic insights suggest a nonsignaling function of Ub in that it directly antagonizes stimuli targeting Ub-modified proteins rather than by recruiting downstream partners for cellular messaging.

Enzyme- and catalyst-generated reactive species have been leveraged in the past decade to covalently label biomolecules within a short range of a defined site or space inside cells or at the cell-cell interface. Due to their high spatial resolution, such proximity labeling strategies have been coupled with various bioanalytical techniques for dissecting dynamic and complex biological processes. Here, we review the development of enzyme- and catalyst-triggered proximity chemistry and their applications to identifying protein interaction networks as well as cell-cell communications in living systems.

Abstract BAK permeabilizes mitochondrial outer membrane, resulting in apoptosis. This apoptotic activity of BAK is stimulated by binding prodeath activators and must be tightly controlled, otherwise it leads to cancers or neurodegenerative diseases. The Parkinson's disease-related E3 ubiquitin (Ub) ligase Parkin ubiquitinates BAK and inhibits the apoptotic activity. However, the molecular mechanism of how ubiquitination inhibits BAK remains uncharacterized. Here, we verify the Ub modification at BAK K113 by Parkin, and further resolve the solution structure of K113-ubiquitinated BAK complex. The conjugated Ub subunit employs its classical L8-I44-H68-V70 hydrophobic patch to bind within the canonical hydrophobic groove of BAK. This groove-harbored Ub occludes the binding of prodeath BID activators, impairs BID-triggered BAK activation and membrane permeabilization. Loosing Ub association with BAK allows BID to activate the K113-ubiquitinated BAK. Together with structure and function evidence, our study now provide mechanistic insights into the ubiquitination regulatory modality wherein Parkin targets BAK to fine-tune apoptosis.

本文聚焦美国政治中的反智主义,对反智主义的内涵、表现、成因及其影响进行了梳理和分析.反智主义的内涵包括反理性主义、反精英主义和对待知识的实用工具主义,也被扩展到反专家及专业知识.反智主义既表现为外显的反智观点,也表现为内在的反智心理.政治生活中的反智主义,集中体现为各政治行为者反理性智识、反知识分子和专家的言行.美国政治中反智主义的形成,受到个体心理与认知因素的影响,也植根于美国宗教、政治和社会发展的历史;媒体信息环境变化加剧其传播和培育,两党制、总统制、民粹主义以及对技治主义的恐惧使其成为一种政治策略.反智主义影响了民众和精英的政治行为,挑战了美国的民主制度,体现了知识与政治、技治与民主间的持续张力,为思考日益不平等的现代社会中民主危机和治理变迁提供了独特视角.

The UN. has intensified efforts to recruit female peacekeepers for peacekeeping missions. From 2006 to 2014, the number of female military personnel in UN peacekeeping missions nearly tripled. The theory driving female recruitment is that female peacekeepers employ distinctive skills that make units more effective along a variety of dimensions. Yet skeptics argue that deeper studies are needed. This paper explores the theoretical mechanisms through which female military personnel are thought to increase the effectiveness of peacekeeping units. Using new data, we document variation in female participation across missions over time, and we explore the impact of female ratio balancing on various conflict outcomes, including the level of female representation in post-conflict political institutions, the prevalence of sexual violence in armed conflict, and the durability of peace. We find evidence that a greater proportion of female personnel is systematically associated with greater implementation of women's rights provisions and a greater willingness to report rape, and we find no evidence of negative consequences for the risk of conflict recurrence. We conclude that the inclusion of more female peacekeepers in UN peacekeeping does not reduce the ability to realize mission goals.

Pyroptosis is an inflammatory cell death form triggered by protease-mediated truncation and release of the N-terminal pore-forming domain of the gasdermin (GSDM) family proteins in various cell types. We report a Bioorthogonally ACtivatable Base editor (BaseBAC) for in situ and on-demand initiation of cell-type-specific pyroptosis. We first made the enzymatic activity of a cytosine base editor (CBE) switchable by establishing a bioorthogonal blockage on the PAM-interacting residue to control its DNA-binding ability. The resulting BaseBAC allowed in situ control of base editing on the GSDME gene that switched to the truncated expression of its N-terminal domain to activate pyroptosis. BaseBAC offers a general method for on-demand awakening of functional domains of self-inhibiting proteins and the corresponding cellular processes with high specificity in living systems.

本文聚焦美国政治中的反智主义,对反智主义的内涵、表现、成因及其影响进行了梳理和分析.反智主义的内涵包括反理性主义、反精英主义和对待知识的实用工具主义,也被扩展到反专家及专业知识.反智主义既表现为外显的反智观点,也表现为内在的反智心理.政治生活中的反智主义,集中体现为各政治行为者反理性智识、反知识分子和专家的言行.美国政治中反智主义的形成,受到个体心理与认知因素的影响,也植根于美国宗教、政治和社会发展的历史;媒体信息环境变化加剧其传播和培育,两党制、总统制、民粹主义以及对技治主义的恐惧使其成为一种政治策略.反智主义影响了民众和精英的政治行为,挑战了美国的民主制度,体现了知识与政治、技治与民主间的持续张力,为思考日益不平等的现代社会中民主危机和治理变迁提供了独特视角.

Green power certificates consumption among individuals can offer environmental public good to minimize pollution caused by the overuse of traditional energy. Studies related to the consumption intention of green power certificates in a developing nation were rare due to the late start market. Regarding this, the present research has attempted to explore consumer purchase intention towards green power certificates in an economic and cultural context of a developing nation, China. The study has adopted the Theory of Planned Behavior (TPB) as the basic theoretical framework and further attempted to incorporate additional constructs (environmental concern, face concern and green perceived value) in measuring its applicability for predicting purchase intention towards green power certificate. A total of 796 effective data samples was collected from the one-on-one field investigation evidenced by latitude and longitude, geographic location of internet protocol, photos and recording backup of each respondent by a convenience sampling approach and analyzed by the Structural Equation Modeling (SEM) in evaluating relationships among constructs. The findings reported that the usefulness and applicability of TPB in predicting the Chinese adult’ purchase intention. The research result supported the applicability of additional construction in TPB as well, as this result has improved the stability of the proposed theoretical model (from 0.063 to 0.059) and made a better understanding on the purchase intention of the Chinese consumer towards green power certificates. In the end, the implications towards policymakers and further research direction have been discussed.

How do Chinese citizens strategize their political participation in the decentralized authoritarian regime? Going beyond the conventional focus on the volume of overall or a specific type of participation, this article focuses on citizens' varying degrees of participation across a range of political activities. It considers the perceived Chinese divided leviathan as a crucial cognitive shortcut for ordinary citizens to assess the uncertain activism environment, estimate the relative costs and benefits of different political activities, and strategize their participation portfolios. Using nationally representative survey data, the article exploits latent class analysis to uncover four distinct mass participatory modes-outsiders, conventionals, agitators, and activists-and examines the impact of perceptual government trustworthiness and integrity on modal transition. The empirical results reveal that citizens' perceptions of a division between the central and local government affect their choice of participatory activities but not their overall participation levels: people who perceive a greater integrity division tend to engage the state in an agitative and contentious mode, and are less likely to do so in an institutionalized, conventional mode. By detecting latent participatory modes and one important underlying psychological rationale, the article contributes to understanding political participation and state-society relations in China.

Temporal activation of proteins of interest (POIs) offers a gain‐of‐function approach to investigate protein functions in dynamic biological processes. Fusion of photo/chemical‐switchable proteins to a POI, or site‐specific blockage/decaging of catalytic residue(s) on a POI, are the most widely utilized strategies for selective protein activation. These methods, however, either lack generality (e.g., active site decaging) or would modify the POI with a bulky tag (e.g., genetic fusion). Recently, a computationally aided and genetically encoded proximal decaging strategy (CAGE‐prox) has been developed for time‐resolved photoactivation of a broad range of proteins in living systems. In contrast to the direct decaging of the active site of a POI, CAGE‐prox relies on a unified caged amino acid that can be anchored in proximity to a protein's functional site for temporal blockage of its activity until rescued by photo/chemical decaging. In order to identify the optimal site for photo‐caged unnatural amino acid insertion, which is key for the effective blockade and re‐activation of the POI, a computational algorithm was developed to screen all possible positions in close proximity to the functional site that would enable turning off/on protein activity via caging/decaging operations. Here, we describe the CAGE‐prox strategy, from in silico design to experimental validation, and provide various examples of its application. © 2021 Wiley Periodicals LLC