Essential for eukaryotes, multiple copies of the exocyst complex tether each secretory vesicle to the plasma membrane (PM) in constitutive exocytosis. The exocyst higher-order structure (ExHOS) that coordinates the action of these multiple exocysts remains unexplored. We integrated particle tracking, super-resolution microscopy, and cryo-electron tomography to time-resolve the continuum conformational landscape of the ExHOS and to functionally annotate its different conformations. We found that 7 exocysts form a flexible ring-shaped ExHOS that tethers vesicles at <45 nm from the PM. The ExHOS rapidly expands while pulling the vesicle toward the PM in a stepwise mechanism comprising three metastable states at 27, 18, and 5 nm from the PM. After fusion, Sec18 mediates the disassembly of the stationary ExHOS, a function that controls the rate of exocytosis. By resolving biophysical principles in situ, we reconstructed the spatiotemporal dynamics of the multimeric architecture controlling vesicle tethering in exocytosis.

The rapid accessibility of calvarial immune cells to the brain, in principle, may offer transformative opportunities for overcoming drug delivery barriers in central nervous system (CNS) disorders. Here, we hijacked calvarial immune cells using drug-loaded nanoparticles (NPs) and leveraged their unique migration mechanism through skull-meninges microchannels to bypass the blood-brain barrier (BBB) for CNS drug delivery. We constructed NP-loaded immune cells in situ via intracalvariosseous (ICO) injection, validated their prompt migration in response to CNS perturbation, and targeted therapeutic delivery to CNS lesions. Compared with conventional delivery approaches, this strategy achieved promising therapeutic efficacy in improving both short- and long-term outcomes in preclinical stroke models. Our prospective clinical trial further supports the translational feasibility of ICO immune access in treating malignant stroke. These findings establish skull-based delivery as a promising, clinically translatable route for CNS drug delivery and highlight immune-assisted transport as a potentially transformative strategy for improving therapeutic outcomes in neurological disorders.

Targeted protein degradation (TPD) has transformed strategies for modulating protein function in both basic biology and therapeutic development. However, current strategies often lack the spatial and temporal precision required for in vivo applications. Herein, we report supramolecular targeting chimeras (SupTACs), a modular and programmable platform that enables tissue-specific and temporally controlled protein degradation in vivo. SupTACs self-assemble into supramolecular nanoparticles (SNPs) that co-localize target-binding ligands and E3 ligase recruiters, thereby facilitating proteasomal degradation through multivalent supramolecular proximity. This strategy achieves robust and tissue-specific degradation, including liver and lung specificity, in multiple species up to non-human primates. As a proof of concept, lung-specific degradation of acyl-coenzyme A (CoA) synthetase long-chain family member 4 (ACSL4) using SupTACs effectively mitigates ferroptosis and pulmonary inflammation in a murine model of acute lung injury. By integrating modularity, tissue specificity, and temporal regulation, SupTACs establish a versatile platform for precise control of protein degradation for interrogating dynamic signaling networks and developing targeted therapeutics.

The efficacy of B cell depletion therapies, and their association with Epstein-Barr virus (EBV), implicate B cells in the pathogenesis of multiple sclerosis (MS). In mice, we observed that viral infections induce infiltration of B cells into the brain, independent of phenotype and specificity, and that myelin-reactive B cells then capture antigens directly from parenchyma. Trafficking of these antigen-loaded B cells to draining lymph nodes was not observed, and without T cell help, antigen-capturing B cells die rapidly. CD40L signaling or EBV latent membrane protein 1 (LMP1) can override this checkpoint, leading to B cell-receptor- and/or antibody-dependent inflammatory demyelination. Myelin-reactive B cells were identified in the healthy human B cell repertoire, and expression of LMP1 was observed in the brains of a subset of MS patients. These observations can explain the dependency of disease incidence on prior EBV infection, and the increased risk associated with brain infections, and suggest possible treatment strategies.

Epstein-Barr virus (EBV) is involved in causing and probably also in perpetuating multiple sclerosis (MS). Among several mechanisms of how EBV may contribute are transcriptome alterations, including changes of antigen processing and preferential presentation of both viral and self-antigens. Here, we report that EBV reprograms the transcriptome and immunopeptidome presented on the MS-associated human leukocyte antigen (HLA)-DR15 molecules of infected B cells. Identical myelin basic protein (MBP) peptides were found to be presented on both EBV-infected B cells and MS brain tissue but not primary B cells and thymic tissue. Peripheral memory and cerebrospinal fluid (CSF)-derived CD4+ T cells of HLA-DR15+ MS patients responded to MBP peptides, MBP(78-90) and/or MBP(83-90), and T cell clones raised with these peptides recognized all MBP peptides ending at amino acid MBP90 in MS brain tissue. Our study provides a new mechanistic link for how the environmental and genetic risk factors, EBV infection and HLA-DR15 haplotype, may contribute jointly to MS.

Epstein-Barr virus (EBV) infection constitutes a prerequisite for multiple sclerosis (MS) development, and cross-reactivity between EBV nuclear antigen 1 (EBNA1) and anoctamin-2 (ANO2) antibodies was previously demonstrated in persons with MS (pwMS). Here, we show that ANO2-specific CD4+ T cells are more frequent in pwMS. Immunization of SJL/J mice with ANO2 or EBNA1 led to cross-reactive CD4+ T cell and antibody responses. ANO2 pre-immunization led to exacerbated experimental autoimmune encephalomyelitis (EAE), an effect mediated by CD4+ T cells, as confirmed by adoptive transfer experiments. T cell clones with cross-reactivity to EBNA1 and ANO2 could be isolated from natalizumab-treated pwMS, and sequencing of EBNA1- and ANO2-specific T cell receptors (TCRs) revealed a significant repertoire overlap. We thus report the first mechanistic evidence that EBNA1 CD4+ T cells can target the MS autoantigen ANO2, thereby establishing a link between EBV infection and neuroinflammation.

The guanosine triphosphate (GTP)-bound state of the heterodimeric Rag GTPases functions as a molecular switch regulating mechanistic target of rapamycin complex 1 (mTORC1) activation at the lysosome downstream of amino acid fluctuations. Under low amino acid conditions, GTPase-activating protein (GAP) activity toward Rags 1 (GATOR1) promotes RagA GTP hydrolysis, preventing mTORC1 activation. KICSTOR recruits and regulates GATOR1 at the lysosome by undefined mechanisms. Here, we resolve the KICSTOR-GATOR1 structure, revealing a striking ∼60-nm crescent-shaped assembly. GATOR1 anchors to KICSTOR via an extensive interface, and mutations that disrupt this interaction impair mTORC1 regulation. The S-adenosylmethionine sensor SAMTOR binds KICSTOR in a manner incompatible with metabolite binding, providing structural insight into methionine sensing via SAMTOR-KICSTOR association. We discover that KICSTOR and GATOR1 form a dimeric supercomplex. This assembly restricts GATOR1 to an orientation that favors the low-affinity active GAP mode of Rag GTPase engagement while sterically restricting access to the high-affinity inhibitory mode, consistent with a model of an active lysosomal GATOR1 docking complex.

Recent advances in regulatory T cell (Treg) biology and clinical application of Treg-based treatments show promise as a new generation of transforming therapeutics for immune-related disorders, positioning Tregs as a "living drug" to rebuild immune tolerance and repair damaged tissues simultaneously. This perspective summarizes the key knowledge on Treg biology and highlights the recent important discoveries in the development of clinical applications based on Treg biology, from low-dose interleukin-2 therapy showing promising results in trials for ALS and adoptive Treg transfer demonstrating efficacy in preventing GVHD to early pilot studies of CAR Tregs. Drawing on these advances, we provide perspectives on key research priorities and translational challenges and set forth a roadmap that integrates basic and clinical insights into developing next-generation therapies focusing on precision tolerance strategies.

Heme carries oxygen and is critical for the control of redox reactions. In this issue of Cell, Lewis and Gruber et al. demonstrate how low concentrations of heme destabilize complex IV of the respiratory chain to release copper and kill acute myeloid leukemia cells by cuproptosis.

Ancient DNA has become a workhorse for evolutionary biologists. In contrast, ancient RNA studies have been rare and often methodologically controversial. Mármol-Sánchez et al. provide compelling evidence that aRNA survived up to ca. 50,000 years in permafrost-preserved mammoth (Mammuthus primigenius) soft tissues. Will this finally pave the way for paleotranscriptomics?

Extrachromosomal DNA (ecDNA) amplifications are key drivers of human cancers. Here, we show that ecDNAs are major platforms for generating and amplifying oncogene fusion transcripts across diverse cancer types. By integrating analysis of whole-genome and transcriptome sequences from tumor samples and cancer cell lines of a wide variety of tissue types, we reveal that ecDNAs have the highest rate of oncogene fusion events of any copy-number alteration. Focusing on the most common ecDNA fusion hotspot, we find that fusion of the 5' end of the long noncoding RNA gene, PVT1-with exon 1 joined to diverse 3' partners-confers increased RNA stability, potentially via an SRSF1-dependent mechanism, and enhances MYC-dependent transcription and cancer cell survival. These results demonstrate that ecDNA fosters genome instability and frequent oncogene fusion formation in cancer.

While much is known about the identity and regulation of cytokine-producing cells, the cell types that respond to cytokines remain largely uncharacterized. To address this knowledge gap, we developed "cytokine cellular locating platforms" (CyCLoPs), a reporter system that translates cytokine receptor engagement into a genetically traceable signal. In vitro, CyCLoPs demonstrated high specificity, robust signal-to-background ratios, and broad applicability for probing diverse cytokine receptor interactions. In vivo, interleukin (IL)-17A-CyCLoPs reporter mice enabled the identification of IL-17A-responsive intestinal epithelial cells predominantly localized in the ileal villi following commensal bacterial colonization. Interferon-gamma (IFN-γ)-CyCLoPs reporter mice allowed for the detection of IFN-γ-exposed CD8+ T cells within tumors, which expressed CD36, CD38, and leptin receptor and displayed gene signatures associated with reduced effector function. Collectively, CyCLoPs offers a platform for the direct visualization and characterization of cytokine-induced cellular responses and provides a tool for investigating how cytokines orchestrate distinct immunological outcomes in health and disease.

The evolutionary history of angiosperms (flowering plants) is characterized by a highly accelerated rate of diversification. Although a number of hypotheses have been proposed to explain the ecological success and rapid rise of angiosperms, the molecular mechanisms underlying their speciation are only partially understood. In this study, we analyzed the developmental transcriptomes of seven angiosperm species spanning 160 million years of evolution. We demonstrate that angiosperm protein-coding gene expression patterns diverged rapidly. Particularly high rates of expression changes were found for genes that are involved in the response to endogenous and environmental stimuli, promoting broad ecological tolerances, adaptive evolution, and speciation. This study highlights how ecological and evolutionary dynamics are highly interrelated, shows that the rates of expression divergence differ from those previously described for mammals, and provides a comprehensive resource to perform cross-kingdom comparative studies of transcriptome evolution.

Understanding the immunogenic properties of different forms of cell death is critical for rationalized antineoplastic therapeutic development. Here, we identify a regulatory axis that suppresses the immunogenicity of ferroptosis. During ferroptosis, but not apoptosis, cuproptosis, or necroptosis, cancer cells release glutathione peroxidase 4 (GPX4), which binds to zona pellucida glycoprotein 3 (ZP3) on the surface of dendritic cells (DCs), activates the 3',5'-cyclic adenosine monophosphate (cAMP)-protein kinase AMP-activated (PRKA) signaling cascade, inhibits glycolysis, and impairs maturation and activation of DCs, leading to a T cell priming defect. Disrupting the interaction between GPX4 and ZP3 restores DC metabolic activity and enhances antitumor immunity. In preclinical models, blockade of this pathway improves cancer immunosurveillance and potentiates cytotoxic T cell responses when combined with chemotherapy, immunochemotherapy, or radiotherapy. Clinically, high ZP3 expression predicts poor prognosis across multiple solid tumor types, while increased circulating GPX4 levels and ZP3 expression in DCs correlate with resistance to first-line therapies. These findings reveal an immunosuppressive danger signal that limits tumor immunity.

Homeostasis and repair in the nervous system are thought to rely on distinct molecular programs. Here, we uncover an unexpected role for the pentose phosphate pathway (PPP) in peripheral sensory axons, where it supports both homeostatic mechanosensation and axonal regeneration after injury. We show that the PPP is enriched and active in sciatic nerve axoplasms, where it maintains redox balance via NADPH production, enabling physiological mechanical sensation. However, following sciatic nerve injury, the PPP is required for regeneration by fueling ribonucleotide synthesis through ribose-5-phosphate. In contrast, this pathway remains inactive after spinal cord injury (SCI), contributing to regenerative failure. Reactivation of the PPP, through neuronal transketolase overexpression or oral ribose supplementation, promotes metabolic reprogramming, restores sensory and motor axonal growth, and improves neurological recovery after SCI. These findings propose the PPP as a metabolic checkpoint in sensory neuron physiology and regeneration, highlighting its therapeutic potential for central nervous system repair.

Type 2 diabetes (T2D) impacts the quality of life and lifespan of nearly 10% of the global population. Human islet amyloid polypeptide (hIAPP) constitutes a major component of islet amyloid deposition in patients with T2D, with hIAPP fibrils believed to play a key role in the pathogenesis of T2D. In this study, we determined the cryo-electron microscopy (cryo-EM) structure of hIAPP fibrils extracted from surgically resected pancreases of three donors with T2D. These fibrils exhibit a uniform morphology, comprising two symmetrical protofilaments encompassing residues 2-37 of hIAPP and adopting an Ω-shaped fold. The structure of pancreatic hIAPP fibrils differs from that of fibrils formed in vitro. Additional densities were observed in the pancreatic hIAPP fibrils, suggesting ligand binding that may play significant roles in the pathogenesis of T2D. Collectively, our study presents the atomic structure of pathological hIAPP fibrils, contributing to the therapeutic and mechanistic exploration of T2D.

Fungal infections cause millions of deaths annually and are challenging to treat due to limited therapeutic options and rising resistance. Cryptococci are intrinsically resistant to the latest generation of antifungals, echinocandins, while Candida auris, a notorious global threat, is also increasingly resistant. We performed a natural product screen to rescue caspofungin fungicidal activity against Cryptococcus neoformans H99 and identified butyrolactol A, which restores echinocandin efficacy against resistant fungal pathogens, including multidrug-resistant C. auris. Mode-of-action studies reveal that butyrolactol A inhibits the phospholipid flippase Apt1-Cdc50, blocking phospholipid transport. Cryo-electron microscopy analysis of the Apt1-butyrolactol A complex reveals that the flippase is trapped in a dead-end state. Apt1 inhibition disrupts membrane asymmetry, vesicular trafficking, and cytoskeletal organization, thereby enhancing echinocandin uptake and potency. This study identifies lipid flippases as promising antifungal targets and demonstrates the potential of revisiting natural products to expand the antifungal arsenal and combat resistance.

Although N6-methyladenosine (m6A) is a pervasive RNA modification essential for gene regulation, dissecting the functions of individual m6A sites remains technically challenging. To overcome this, we developed functional m6A sites detection by CRISPR-dCas13b-FTO screening (FOCAS), a CRISPR-dCas13b-based platform enabling high-throughput, site-specific functional screening of m6A. Applying FOCAS to four human cancer cell lines identified 4,475 m6A-regulated genes influencing cell fitness via both mRNAs and non-coding RNAs (ncRNAs), many of which are newly linked to cancer and exhibit dynamic developmental expression. FOCAS uncovered context-dependent and reader-specific effects of m6A within the same gene, revealing its intricate regulatory logic. We further uncovered universal and cell-type-specific m6A patterns, with unique sites enriched in ncRNAs and universal ones in transcription-related genes. In SMMC-7721 cells, we identified m6A-regulated transcriptional networks that demonstrated extensive epitranscriptome-transcriptome crosstalk. Overall, this study established a powerful, unbiased approach for the functional dissection of m6A, advancing the understanding of its complexity and therapeutic relevance in cancers.

Herpesviruses are widespread double-stranded DNA viruses that establish lifelong latency and cause various diseases. Although DNA-polymerase-targeting antivirals are effective, increasing drug resistance underscores the need for alternatives. Helicase-primase inhibitors (HPIs) are promising antivirals, but their mechanisms of action are poorly defined. Furthermore, how the helicase-primase (H/P) complex and DNA polymerase coordinate genome replication is not well understood for herpesviruses. Here, we report cryo-electron microscopy (cryo-EM) structures of the herpes simplex virus 1 H/P complex bound to HPIs, showing that these lock the H/P complex in an inactive state. Single-molecule assays reveal that HPIs cause H/P complexes to pause in unwinding activity on DNA. The structure of an HPI-bound replication fork complex, comprising the H/P complex (UL5, UL52, and UL8) and the polymerase holoenzyme (UL30 and UL42), reveals a previously uncharacterized interface bridging these complexes. These findings provide a structural framework for understanding herpesvirus replisome assembly and advancing inhibitor development.

A typical neuron signals to downstream cells when it is depolarized and fires sodium spikes. Some neurons, however, also fire calcium spikes when hyperpolarized. The function of such bidirectional signaling remains unclear in most circuits. Here, we show how a neuron class that participates in vector computation in the fly central complex employs hyperpolarization-elicited calcium spikes to invert two-dimensional mathematical vectors. By switching from firing sodium to calcium spikes, these neurons implement a ∼180° realignment between the vector encoded in the neuronal population and the fly's internal compass signal, thus inverting the vector. We show that calcium spikes rely on the T-type calcium channel Ca-α1T and argue via analytical and experimental approaches that these spikes enable vector computations in portions of angular space that would otherwise be inaccessible. These results reveal a seamless interaction between molecular, cellular, and circuit properties for implementing vector mathematics in the brain.