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Wu, J.

Publications and source records attributed to Wu, J..

At least 127 records · Page 7Linked to original sources

Stimulus-Driven Thermodynamic Shifts and Geometric Reorganization in Mouse Primary Visual Cortex

Efficient coding is essential for sensory systems to extract meaningful information from the environment. Here, we investigate how stimulus-driven thermodynamic shifts and geometric reorganization enable efficient population coding. Using wide-field calcium imaging, we simultaneously recorded neuronal activity across the entire mouse V1 under the presentation of structured stimuli and characterized neural dynamics spanning microscale neuronal connectivity, mesoscale thermodynamic states, and macroscale manifold geometry. We found that stimulus presentation reorganized neuronal couplings into increasingly modular subnetworks, driving a shift from near-critical dynamics toward a more ordered regime. This shift coincided with a compression of neural population activity onto low-dimensional manifolds aligned with stimulus features, thereby enabling efficient coding. Furthermore, mathematical derivations and in silico perturbation experiments confirmed that selective modulations of neuronal connectivity altered both critical temperature and manifold geometry, establishing a causal link from neuronal couplings to efficient coding. Collectively, our findings suggest a mechanistic bridge between statistical physics and neural geometry, providing new theoretical insights into how neural networks dynamically transition into more stable and efficient coding states in response to the environment.

neuroscience↗

Intracellular TDP-43 amyloid nucleates from arrested nascent condensates

TDP-43 is a model protein for pathophysiological phase transitions, forming a multitude of intracellular assemblies with different physical properties. Physiological condensation is widely presumed to precede pathological aggregation, but the causal relationships between different modes of assembly in vivo are still unclear. Here we use Distributed Amphifluoric FRET (DAmFRET) and complementary approaches to map the phase space of TDP-43 self-assembly in yeast cells. We discovered that the low-complexity C-terminal domain (CTD) on its own partitions into soluble clusters that dynamically arrest en route to liquid-liquid phase separation. These clusters uniquely supported amyloid formation, and only when templated by pre-existing amyloids of other proteins. Self-interacting modules outside the CTD, whether in the full-length TDP-43, pathological C-terminal fragments, or fusion partners, all suppressed amyloid nucleation. They did this by promoting CTD condensation beyond the arrested state. Stress and cotranslational self-association had the same effect. We leveraged this property of condensation to stop CTD amyloid formation by co-expressing an oligomeric binder in cells. Our findings reveal that TDP-43 amyloid formation occurs only under very specific physical and biological circumstances that present new opportunities for therapeutic control.

biophysics↗

Decoupling Structure and Property in Discrete Protein Diffusion: Alignment Dynamics and Collapse Mechanisms

Direct Preference Optimization (DPO) has emerged as a powerful paradigm for aligning generative models, yet its temporal optimization dynamics in the discrete diffusion space of proteins remain poorly understood. Existing approaches often assume that maintaining structural integrity while optimizing physicochemical properties requires simultaneous, tightly coupled reinforcement learning constraints. In this work, we challenge this assumption by uncovering a fundamental temporal decoupling between structural and functional alignment. Using antibody design as a testbed, extensive trajectory analysis reveals two distinct regimes: (1) Instant Structural Alignment, where the strong generative prior of discrete diffusion rapidly eliminates structural hallucinations via denoising within the first few epochs; and (2) Slow Property Adaptation, where physicochemical attributes improve gradually over a prolonged optimization window. We further identify a critical transition point around Epoch 50, which empirically defines a Pareto-optimal boundary between property improvement and structural stability. Beyond this point, continued optimization induces a sharp phase transition into a Structural Collapse regime. To isolate the physical driver underlying this collapse, we introduce a counterfactual preference experiment targeting negative charge. We observe a striking symmetrical collapse: while hydrophilicity optimization induces a Poly-Arginine (+) degeneration, negative charge optimization drives a Poly-Aspartate (-) degeneration. Despite opposite physicochemical trajectories, including extreme shifts in isoelectric point (> 11 vs. < 4.5), both regimes converge to the same structural failure. This symmetry demonstrates that generic Coulombic repulsion, rather than residue-specific bias, constitutes the fundamental physical constraint being violated. Our findings reveal that discrete diffusion models possess strong intrinsic structural robustness, enabling minimalist alignment strategies provided optimization halts before this physical boundary. More broadly, this work offers a mechanistic warning against unchecked reward optimization in biological generation, illustrating a concrete manifestation of Goodharts Law in protein design. Code and data are available at https://github.com/Wu-Junqi/DPO-Protein-Diffusion.

bioinformatics↗

Ophiuchus-Ab: A Versatile Generative Foundation Model for Advanced Antibody-Based Immunotherapy

Antibodies exhibit extraordinary specificity and diversity in antigen recognition and have become a central class of therapeutics across a wide range of diseases. Despite this clinical success, antibody design remains fundamentally challenging. Antibody function emerges from intricate and highly coupled interactions between heavy and light chains, which complicate sequence-function relationships and limit the rational design of developable antibodies. Here, we reveal that modeling antibody sequence space at the level of paired heavy and light chains is essential to faithfully capture inter-chain dependencies, enabling a deeper understanding of antibody function and facilitating antibody discovery. We present Ophiuchus-Ab, a generative foundation model pre-trained on largescale paired antibody repertoires within a diffusion language modeling framework, unifying antibody generation and representation learning in a single probabilistic formulation. This framework excels diverse antibody design tasks, including CDR infilling, antibody humanization, and light-chain pairing. Beyond generation, diffusion-based pre-training yields transferable representations that enable accurate prediction of antibody properties, including developability, binding affinity, and specificity, even in low-data regimes. Together, these results establish Ophiuchus-Ab as a versatile foundation model for modeling antibodies, providing a foundation for next-generation antibody-based immunotherapy.

bioinformatics↗

cGAS-mediated IFN-I signaling contributes to disease progression in drug-refractory epilepsy

Epilepsy is a prevalent neurological disease with a third of patients becoming non-responsive to antiepileptic drugs and developing drug-refractory epilepsy (DRE). Here we report that DRE disease progression is contributed by overactive cyclic GMP-AMP synthase (cGAS), a double-stranded DNA sensor that induces type I interferon (IFN-I) signaling. In human DRE microglia, we observe a robust IFN-I signature and the activation of upstream cGAS-STING signaling. Further, in mouse models of Dravet syndrome, a genetic form of DRE, we observe the activation of the cGAS pathway. We show that microglial cGAS can be activated by DNA released from hyperexcitable neurons. Genetic reduction and pharmacological inhibition of cGAS reduces epileptic phenotypes, glial inflammatory signatures, and neuronal transcriptomic changes, underscoring the therapeutic potential of targeting cGAS for DRE treatment.

neuroscience↗

Spatial multi-omics unveils the monoclonal origin, neuroendocrine plasticity, and microenvironment niches in combined small cell lung cancer

Combined small-cell lung cancer (cSCLC) is a rare and aggressive subtype of small-cell lung cancer (SCLC) characterized by mixed histology comprising SCLC and non-small cell lung cancer (NSCLC) or large cell neuroendocrine carcinoma (LCNEC) components. Despite its histological heterogeneity and even poorer prognosis than de novo SCLC, cSCLC is clinically managed as pure SCLC, largely due to the lack of molecular insights into its biology, lineage plasticity, and tumor microenvironment (TME). Here, we perform multi-omics profiling, including spatially-resolved whole-exome sequencing (WES), spatial transcriptomics (ST) and single-nucleus RNA sequencing (snRNA-seq), across 19 treatment-naive cSCLC tumors spanning all major histological subtypes. Our analysis reveals that SCLC and NSCLC/LCNEC components share a monoclonal origin, with histological divergence characterized by distinct mutation and copy number alteration patterns. ST and snRNA-seq uncover spatially exclusive or interspersed tumor domains, with distinct TME compositions and immune landscapes. Notably, fibroblast-rich regions enriched for an aggressive fibroblast subtype form boundaries between tumor domains, potentially influencing immune TME and treatment responses. We identify extensive lineage plasticity within cSCLC, including active LUAD-to-SCLC transdifferentiation and SCLC subtype coexistence, suggesting transitional cellular states not captured by traditional diagnostics. Leveraging these insights, we developed the cSCLC Detector, a sensitive mutation-based diagnostic assay that improves the detection of cSCLC in tissue and liquid biopsy samples. Our findings offer critical insights into cSCLC lineage plasticity, cellular evolution, and microenvironmental interactions, underscoring the need for tailored treatment strategies and diagnostic frameworks for this aggressive cancer subtype.

cancer biology↗

Integrating UHPLC-MS and MALDI-MSI for Spatial Nucleoside Profiling in FFPE Breast Cancer: A Multimodal Molecular Pathology Framework

Formalin-fixed, paraffin-embedded (FFPE) tissues constitute the primary material for diagnostic pathology and retrospective clinical research, yet their use in metabolomics remains limited due to molecular cross-linking and analyte degradation. Here, we establish a cost-efficient molecular pathology workflow that integrates ultra-high-performance liquid chromatography mass spectrometry (UHPLC-MS) with matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) to quantify and spatially map nucleosides in FFPE breast cancer tissues. Optimized extraction using methanol yielded nucleoside profiles comparable to fresh-frozen tissues, while MALDI-MSI enabled the spatial visualization of nine nucleosides across distinct histological regions. Several nucleosides including deoxyadenosine and 5-formylcytosine showed strong discriminatory power between tumor stages, revealing progressive metabolic rewiring during breast cancer progression. Finally, spatial nucleoside patterns observed in a murine model were recapitulated in patient-derived FFPE tissues, underscoring the translational potential of nucleoside-based spatial metabolomics for clinical research and biomarker discovery. Together, this workflow establishes MALDI-MSI as a powerful and scalable spatial molecular pathology tool for interrogating nucleoside biology in archival breast cancer samples. Following MALDI-MSI, the same FFPE tissue sections can undergo laser capture microdissection, enabling genomic, proteomic, or targeted metabolomic profiling of MSI-defined tumor niches and microenvironmental regions. This integration directly links spatial nucleoside signatures to molecular alterations relevant to precision oncology in future.

cancer biology↗

p300/CBP-Driven Acetylation Stabilizes PARP7 to Reinforce Negative Feedback on IFN-I Signaling

Immune suppression within the tumor microenvironment (TME) remains a major barrier to effective immunotherapy, yet the molecular mechanisms that constrain type I interferon (IFN-I) signaling in tumor cells are incompletely understood. PARP7 (also known as TIPARP), a mono-ADP-ribosyltransferase, has been reported to suppress IFN-I signaling by modifying TBK1. Here, we identify PARP7 as a previously unrecognized acetylation substrate of the histone acetyltransferases p300 and CBP. We show that p300/CBP-mediated acetylation of PARP7 at lysine 32 markedly enhances its stability, thereby reinforcing repression of the TBK1-IRF3-STAT1 axis and suppressing IFN-{beta}-induced expression of interferon-stimulated genes, including CXCL10. Beyond their established function as transcriptional co-activators that support tumor cell proliferation, our findings uncover a direct role for p300/CBP in shaping innate antitumor immunity. Pharmacological inhibition of p300/CBP catalytic activity by A-485 abolishes PARP7 acetylation, accelerates its proteasomal turnover, and restores IFN-I signaling. In immunocompetent tumor models, this is accompanied by enhanced CD8+ T-cell infiltration and effector function. Together, these findings define a p300/CBP-PARP7 regulatory axis, uncover a non-canonical role for p300/CBP in innate immune suppression, and nominate this pathway as a tractable target for restoring antitumor immunity.

cancer biology↗

Machine Learning Ensemble Reveals Distinct Molecular Pathways of Retinal Damage in Spaceflown Mice

BackgroundSpaceflight-associated neuro-ocular syndrome (SANS) poses significant risks to astronaut visual health during long-duration missions, yet its underlying molecular mechanisms remain incompletely understood. Oxidative stress and apoptosis are candidate molecular drivers, but their transcriptomic signatures in spaceflight-exposed retinal tissue have not been systematically characterized. MethodsWe applied a machine learning ensemble of linear regression models to predict two ocular phenotypes: 4-hydroxynonenal (4-HNE) immunostaining as a marker of lipid peroxidation-mediated oxidative damage; and TUNEL positivity as a marker of apoptotic cell death. In this observational study, we use bulk retinal gene expression data obtained from a controlled experiment with ground control and spaceflown mice to predict these phenotypes. Gene Ontology pathway enrichment was performed on the most predictive gene sets for each phenotype. ResultsThe 4-HNE phenotype was predicted by genes that converge on membrane-associated pathways, photoreceptor protein modification, synaptic dysfunction, and extracellular matrix dysregulation, including B2m, Tf, Cnga1, mt-Nd1, Snap25, and Efemp1. The genes predicting the TUNEL phenotype revealed a distinct signature emphasizing stress-induced apoptosis, rod photoreceptor degeneration, and endoplasmic reticulum dysfunction, with Ddit4, Nrl, Rom1, Reep6, and Gabarapl1 emerging as central regulators. ConclusionsOxidative lipid peroxidation and apoptotic cell death represent complementary and molecularly distinct pathological mechanisms in spaceflight-exposed murine retinal tissue. The gene signatures provide a putative molecular framework for developing noninvasive biomarkers and therapeutic targets to monitor and protect astronaut visual health during long-duration and deep-space missions.

bioinformatics↗

Myeloid CD209 impairs T-cell activation by inducing ICAM-2-ERM-dependent cortical stiffening

Resistance to PD-1-based immunotherapy remains a major challenge in cancer treatment, especially in immunologically "cold" tumors characterized by T-cell exclusion and impaired effector function. Tumor-associated myeloid cells are important mediators of this resistance, but whether they directly suppress T-cell immunity through mechanical regulation remains unclear. Here, we identify DC-SIGN (CD209), a myeloid-expressed C-type lectin receptor, as a mechanosuppressive regulator of antitumor T-cell immunity. DC-SIGN is selectively enriched in tumor-infiltrating M2-like macrophages and monocytes, and its expression is associated with poor clinical response to PD-1 blockade and elevated TIDE scores across multiple cancer types. Mechanistically, DC-SIGN directly engages ICAM-2 on T cells and activates ROCK1-ERM signaling. This interaction increases T-cell cortical stiffness, destabilizes T cell-antigen-presenting cell interactions, and attenuates T-cell receptor signaling. Blocking the DC-SIGN-ICAM-2-ERM pathway restores T-cell activation and promotes CD8+ T-cell infiltration and effector function in mouse, humanized, and ex vivo human tumor systems. Importantly, DC-SIGN blockade overcomes resistance to PD-1-based immunotherapy in refractory and immunologically "cold" tumor models, and further enhances the response to PD-1 inhibition. Together, our findings define a myeloid-derived mechanical immune checkpoint and reveal T-cell mechanosuppression as a targetable mechanism of tumor immune evasion and immunotherapy resistance.

immunology↗

Dye-cycling DNA origami rotors for long-term tracking of transcription at base-pair resolution

Protein interactions with DNA often result in rotational movements. These movements underlie many genetic processes such as DNA transcription by RNA polymerase (RNAP). To illuminate these movements, we previously developed Origami Rotor-Based Imaging and Tracking (ORBIT), a fluorescence-based method that enables high-speed tracking of DNA rotation. When used to track DNA rotation during transcription by E. coli RNAP, ORBIT enabled detection of single base-pair steps. However, the duration of ORBIT experiments was limited due to fluorescence photobleaching. To overcome this limitation and extend the total possible observation time, we here introduce dye-cycling ORBIT (dcORBIT), a method that enables observation of protein-DNA interactions for over 10 minutes at 20 Hz while maintaining single base-pair precision. dcORBIT thereby opens new possibilities to study the fundamental rotational movements underlying gene expression.

biophysics↗

ISG15 orchestrates dynamic crosstalk between mitochondrial fat oxidation and type 1 interferon in myeloid cells

In contrast to Krebs cycle intermediates, the role of mitochondrial fatty acid oxidation (FAO) in immunometabolism remains incompletely characterized. Studying primary bone marrow-derived macrophages (BMDMs), we show that IFN-{beta} and STING activation augments FAO and associated enzymes carnitine palmitoyltransferase 1a (CPT1a) and acetyl-CoA acetyltransferase 1 (ACAT1). Depleting BMDM Cpt1a reduces FAO and dampens type 1 interferon (IFN) signaling due to decreased histone H3-K9/K14 acetylation, supporting the prior finding of an epigenetic role of FAO in sustaining type 1 IFN. Interestingly, FAO induction by IFN was dynamic as a heightened IFN response suppressed FAO, suggesting a concurrent negative-feedback mechanism. This FAO blunting correlated with increased expression of interferon-stimulated gene 15 (ISG15), a ubiquitin-like modifier known to modulate metabolic proteins through ISGylation. This immune-metabolic signature was similarly operational in mice infected with lymphocytic choriomenigitis virus (LCMV) with temporal discordance between ISG15 levels and FAO. The role of ISG15 in this negative feedback was shown with increased FAO and type 1 IFN response in Isg15 knockout BMDMs. Parallely, endogenous co-immunoprecipitation showed interactions between ISG15 and CPT1a/ACAT1. This ISG15-FAO regulatory interaction was also evident in systemic lupus erythematosus (SLE)-associated interferonopathy and in primary monocytes from SLE individuals that exhibited increased ISG15 levels and reduced FAO rates. Collectively, these findings support a model in which type 1 IFN initially enhances FAO to amplify interferon production, but excessive IFN-signaling induces ISG15-mediated inhibition of FAO, a putative feedback loop that restrains inflammation and preserves immune homeostasis. Together these data identify a novel biphasic FAO-dependent immunometabolic regulatory program. Graphical Summary O_FIG O_LINKSMALLFIG WIDTH=190 HEIGHT=200 SRC="FIGDIR/small/700051v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@13f572eorg.highwire.dtl.DTLVardef@174b6aeorg.highwire.dtl.DTLVardef@1856483org.highwire.dtl.DTLVardef@89e5a6_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIType 1 IFN rewires myeloid cell metabolism towards FAO. C_LIO_LIFAO epigenetically fuels Type 1 1IFN response. C_LIO_LIEffect of IFN on FAO is dynamic: inducing FAO at low concentrations while suppressing it during a heightened IFN response. C_LIO_LIEnhanced Type 1 IFN response produces ISG15 which ISGylates key enzymes of FAO altering their stability C_LIO_LIISG15 negatively regulates FAO to control Type 1 IFN homeostasis in a negative feedback loop. C_LI

immunology↗

Co-option of ILF2/3 in primates restrains Alu hyper-editing to enable cell fate transitions

Understanding human specific mechanisms of cell fate control is essential for advancing developmental biology and regenerative medicine. Here, we identify the ILF2/3 complex as a critical regulator of primate cell fate transitions. Using genetically and epigenetically engineered gastruloids and adult stem cells, we show that ILF2/3 is required for gastrulation in primates but not mice, and for differentiation of adult progenitor cells. Mechanistically, ILF2/3 directly binds Alu elements in chromatin-associated RNAs and shields them from ADAR1-mediated adenosine-to-inosine (A-to-I) editing. Acute ILF2/3 degradation increases A-to-I editing at Alu elements, but not murine retrotransposons, leading to aberrant splicing and nonsense-mediated decay of transcripts encoding key chromatin regulators in primate cells. This in turn destabilizes the epigenetic landscape and blocks lineage commitment across all three germ layers. Re-expression of correctly spliced chromatin regulators rescues differentiation defects in ILF2/3-deficient cells, functionally linking Alu editing control to chromatin regulation and cell fate. These findings define an evolutionary mechanism that restrains retrotransposon-associated RNA editing to preserve proteome integrity and enable primate-specific developmental programs.

genetics↗

Dual-modality, deep-learning-enabled endomicroscope with large field-of-view and depth-of-field for real-time in vivo imaging of epithelial hallmarks of cancer

In vivo microscopy (IVM) has shown great promise to improve early detection of epithelial precancer, but it suffers from fundamental trade-offs that limit the resolution, field-of-view (FOV) and depth-of-field (DOF). Here, we present PrecisionView, a compact, deep-learning-enabled endomicroscope that breaks these constrains and achieves 20 mm2 FOV and 500 {micro}m DOF with 4 {micro}m resolution, representing approximately 5x increase in FOV and 8x larger DOF compared to conventional IVM with similar resolution. PrecisionView integrates a deep-learning optimized phase mask and real-time reconstruction, enabling rapid in vivo assessment of two key hallmarks of cancer: epithelial cell nuclear morphology and subsurface microvasculature through fluorescence and reflectance imaging. By imaging oral cavity of healthy volunteers and cervical specimens with precancerous lesions, PrecisionView generates large-scale (1-3 cm2) co-registered maps of cellular and vascular structures, revealing distinct microscopic patterns associated with anatomic structures and precancerous lesions. Our results suggest the potential of this computational endomicroscope to address the unmet need for early cancer detection at the point-of-care.

bioengineering↗

Development of Integrin α5β1-targeted PET/NIR imaging probes for glioblastoma intraoperative navigation and intracavity targeted radionuclide therapy

Incomplete extent of tumor resection is the major factor of glioblastoma recurrence, leading to its poor prognosis. Developing therapies that enable the minimal tumor cell remnant during surgical resection is a major challenge to combat this aggressive cancer. This study develops a glioblastoma integrin 5{beta}1-selective peptide binder through series strategy screening and labeling it with 68Ga, indocyanine green (ICG), or 177Lu for efficacious multimodal treatment. This strategy combines positron emission tomography (PET) imaging for preoperative diagnosis, intraoperative NIR-II fluorescence-guided surgery for maximal tumor resection, and followed by intracavity targeted radionuclide therapy for elimination of residual tumor cells. We identified GR, with five arginine mutation, exhibiting superior integrin 5{beta}1 binding affinity and brain penetration. [68Ga]GR demonstrated substantial tumor uptake and prolonged retention time in both mouse models and tissues from glioblastoma patients. The guidance of ICG-GR achieved accurately intraoperative tumor imaging and maximal tumor resection in orthotopic mouse glioblastoma models. Notably, combining intracavity administration of [177Lu]GR following ICG-GR guiding resection markedly inhibited tumor growth and reduced possibility of tumor recurrence compared with surgery alone (unguided or ICG-GR-guided) or unguided surgery followed by [177Lu]GR. Therefore, we have reported the integrin 5{beta}1-targeted radionuclide and optical agents for maximal elimination of tumor cells during surgical resection in glioblastoma patients. HIGHLIGHTSO_LIGR is a glioblastoma-specific peptide with both favorable binding affinity superior brain penetration. C_LIO_LIICG-GR is a glioblastoma-targeted NIR probe enabling maximal surgical resection. C_LIO_LIICG-GR-guided surgery with [177Lu]GR-based adjuvant therapy allows robust elimination of tumor cells. C_LI

cancer biology↗

Glucose is a ligand-like trigger for secretion of postprandial GLP-1

Glucagon-like peptide 1 (GLP-1) is secreted from the intestinal L cells in a glucose-dependent manner. However, the mechanism underlying how glucose stimulates the secretion remains unclear. Here, we found that glucose at postprandial concentrations near the Km of GK for glucose ([~]10 mM), as well as nonmetabolizable glucose analogs, triggers GLP-1 secretion by intestinal L cells through directly binding to glucokinase (GK), without increasing intracellular ATP levels. We also show that in postprandially high glucose, glucose-bound GK interacts and inhibits KATP, as assessed by immunoprecipitation and FRET-FLIM analyses, as well as patch clamping. In addition, we have utilized GK mutants identified in diabetics, demonstrating that those mutants defective in binding to glucose impair GLP-1 secretion in however high glucose, whereas mutants constitutively bound to glucose with defects in catalysis, promote GLP-1 secretion even in low glucose. Furthermore, the KATP mutant unable to bind to GK blocks GLP-1 secretion in high glucose. We have also provided evidence showing that sodium-glucose cotransporter 1 (SGLT1) plays an important role in the basal secretion of GLP-1, but not in high glucose. We have thus revealed that glucose the molecule itself and GK act as a ligand and the receptor, respectively, to induce GLP-1 secretion in response to high glucose, providing new therapeutic avenues targeting metabolic diseases.

biochemistry↗

Kilohertz-rate two-photon voltage imaging of population dynamics in vivo

Understanding how neural circuits compute requires capturing voltage dynamics across large neuronal populations with millisecond resolution in vivo. However, two-photon voltage imaging remains fundamentally limited by a trade-off among imaging speed, field of view, and excitation efficiency. We introduce HS2PM, a hybrid scanning two-photon microscope that overcomes this bottleneck, enabling kilohertz-rate imaging over a 650 x 524 m2 field of view at single-cell resolution while preserving the photon efficiency of single-point excitation. HS2PM stably records deep-layer membrane potentials from over 160 neurons simultaneously, with minimal photobleaching and phototoxicity. It resolves both spikes and subthreshold voltage dynamics in vivo, revealing how these jointly shape sensory adaptation and population coding. Beyond voltage imaging, HS2PM supports high-speed fluorescence lifetime and vascular flow imaging, establishing a multimodal platform for dissecting fast circuit dynamics with precision previously inaccessible to optical methods.

neuroscience↗

Macrophage-Dendritic Cell-T-Cell Tetrads Orchestrate Antitumor Immunity and Response to Checkpoint Blockade

Immune checkpoint inhibitors (ICIs) elicit durable responses in only a subset of patients with solid tumors, underscoring the need to define the cellular architectures that govern effective antitumor immunity. Here we identify a spatially organized multicellular immune unit comprising macrophages, cDC1s, CD4 T-cells, and CD8 T-cells that emerges in response to anti-CTLA-4 or dual checkpoint blockade. We term these structures tetrads. Using multiplexed imaging and spatial transcriptomics in mouse and human tumors, we show that tetrads assemble early during immune priming, depend on the ICOS-ICOSL pathway, and are enriched for ICOS Th1-like CD4 T cells and ICOSLhigh cDC1s. CD8 T-cells within tetrads exhibit an activated, non-terminally differentiated state, while tetrad-associated macrophages display an interferon-{gamma}-responsive program that sustains CD8 T-cell function and prevents dysfunction. Functionally, ICOSL cDC1s are required for tumor eradication in vivo. In patients with bladder cancer treated with neoadjuvant dual checkpoint blockade, tetrad, but not triad or dyad formation correlates with clinical response. These findings establish tetrads as a fundamental cellular unit coordinating antitumor immunity and responsiveness to ICIs.

immunology↗