Nature Materials

Granular scaffolds have emerged as promising platforms for tissue regeneration, offering injectability and cell-scale porosity that support robust cell infiltration and tissue formation. However, the isotropic pore structure of spherical building blocks does not provide the directional cues needed to guide organized tissue formation. Addressing this requires asking not just whether granular scaffolds can be made anisotropic, but whether directional cues persist across the pore network at scales relevant to cell behavior. Using high aspect ratio GelMA hydrogel fibers as building blocks, we demonstrate that spherical granular materials lose orientational coherence at the cellular scale, confirming that isotropic building blocks are fundamentally incapable of providing structural guidance beyond individual pore neighborhoods. In contrast, fibrous building blocks extend persistence into the multicellular range, occupying an intermediate architectural regime exhibiting locally coherent but globally variable organization, rather than simple isotropic or uniaxial alignment, that has previously been inaccessible to granular scaffold design. We show this regime is functionally meaningful: myotubes undergo contact guidance through locally persistent but globally variable pore structure, and greater persistence is associated with increased myotube elongation and multinucleation in primary human muscle progenitor cells. Together these results expand the design space for granular scaffolds beyond pore size and porosity, and establish persistence as a variable linking granular scaffold architecture to organized tissue formation.

Gene order is a powerful design principle for protein nanomachines. In nature, gene organisation ensures the precise assembly of functional protein nanostructures. We demonstrate how genetic repositioning of the key structural gene pduN, within the operon encoding a self-assembling protein nanocompartment, sculpts the morphology and function of bacterial microcompartments (BMCs). Relocating pduN to new operonic positions dramatically altered the size, shape, and catalytic output of BMCs, despite identical protein sequences. These shifts reveal how gene order may control nanoscale assembly and compartmentalised function. Our findings establish operon architecture as a programmable genetic framework for nanostructure morphogenesis and provide a synthetic biology strategy to engineer self-assembling nanodevices with customised geometries and activities.

Understanding protein phase separation in cellular environments remains a major challenge, as ex vivo assays often fail to capture the influence of environmental context - such as crowding, multimodal interactions, and the dynamic properties of the cytosol or nucleus. Here, we introduce programmable DNA-based protonuclei (PN) as nucleus-inspired compartments to probe phase separation of the neurodegeneration-linked protein FUS. We show that FUS partitioning and condensate formation are highly sensitive to nucleic acid sequence, spatial confinement, and viscoelastic properties of the PN core. Notably, classical test-tube affinity assays fail to predict protein behavior within the crowded and multivalent PN environment. By tuning DNA crosslinking, we modulate condensate dynamics and suppress liquid-to-solid transitions of FUS - a hallmark of disease. These findings demonstrate that multivalent, confined environments fundamentally reshape protein-nucleic acid interactions and phase behavior. The PN platform complements test-tube assays and complex cellular settings and enables to dissect nuclear condensates under controllable conditions.

Protein immunotherapies can elicit potent tumor rejection, but reversible target engagement, incomplete tumor retention, and systemic leakage often erode spatial control. Here, we develop covalently anchored tumor immunotherapeutic proteins (CATIPs), a modular platform that uses proximity-enabled covalent chemistry to immobilize immune cues on tumor-cell surfaces after intratumoral administration. CATIPs combine tumor-targeting nanobodies with payloads for T cell engagement, co-stimulation, and cytokine support. In human PBMC-reconstituted NSG mice, CATIPs completely eradicated treated EGFR-positive tumors, outperforming matched non-covalent proteins while limiting redistribution, systemic T cell activation, cytokine release, xGVHD-associated morbidity, and on-target, off tumor toxicity. In immunocompetent melanoma models, CATIPs remodeled the tumor microenvironment, expanded antigen-specific CD8+ T cells, induced antigen-restricted abscopal control, and generated durable protection against local and metastatic rechallenge. CATIP-engineered tumor cells further functioned as whole-cell vaccines. Thus, covalent tumor anchoring converts local protein delivery into tumor-surface immune programming, enabling systemic, tumor-specific, durable antitumor immunity while limiting systemic immunopathology.

Engineered living materials (ELMs) harness the adaptive and self-replicating capabilities of biological systems to create functional materials for sensing, catalysis, and biomineralization. While most ELM strategies rely on static microbial assemblies, the role of bacterial motility in structuring living materials remains unexplored. Here, for the first time, we demonstrate how swarming motility in Escherichia coli MG1655 can be induced to guide spatio-temporally organized calcium phosphate mineralization. The mineralized calcium phosphate is characterized by scanning electron microscopy and elemental analysis. By systematically varying phosphate sources and their concentrations in calcium-rich media, we observe the emergence of regularly spaced concentric mineralized patterns. The previously undocumented observation of the concentric patterns was rationalized through a continuum model that captures the spatiotemporal coupling between swarm expansion and mineral deposition. The model shows that this coupling can generate recurrent front arrest and restart, leading to concentric ring formation. Finally, we show that altering the phosphate species results in distinct mineral morphologies. Together, this work establishes a novel framework for integrating bacterial swarming with biomineralization, enabling dynamic and programmable pattern formation in ELMs.

Microstructures created with flow lithography exhibit distinct functionality depending on the shape and composition of the precursor fluids, enabling applications from tissue engineering to anti-counterfeiting. However, current techniques rely on static nozzle geometries or passive hydrodynamic focusing, which commit to a fixed structure and limit dynamic reconfiguration of material architecture during fabrication. Here, we introduce ActiSculpt, an acoustofluidic platform that replaces in-channel physical structures with programmable, electronically driven acoustic streaming. By exploiting the interplay between laminar stability and acoustic streaming, we decouple deterministic fluid deformation from chaotic mixing, achieving a continuous cross-sectional displacement sensitivity of ~15 m/V. We demonstrate the generation of a diverse library of hydrogel particles whose cross-sectional moments of inertia are tunable up to 5.5-fold, establishing a direct, geometry-mediated link between acoustic parameters and the moments that govern bending and torsional rigidity. We further demonstrate continuous fiber fabrication in which acoustic parameters are varied in real time, encoding structural variation along the fibers length. The result is a platform that overcomes the one-device, one-geometry constraint of existing techniques, enabling not only on-demand reconfiguration between fabrication runs but also real-time control of material architecture. This spatiotemporal control establishes a new design axis for soft-material manufacturing.

Cell migration is fundamental to various biological processes, including morphogenesis, wound healing, and cancer metastasis. Durotaxis--directed migration of cells in response to spatial variations in stiffness--has been extensively studied using engineered substrates with prescribed stiffness. However, recent work has increasingly shifted toward understanding cell migration in fibrous matrices that can be actively remodeled by the actomyosin contractility, as commonly observed in tumor and epithelial cells. Despite these advances, a theoretical framework explaining how cells structurally remodel their surrounding matrix to promote their own durotaxis, and which cellular forces govern this behavior, remains elusive. To address this gap, we developed a biomechanical model in which polarized cells contract and migrate over a fibrous matrix. Using this model, we first confirmed that cells on an externally strained matrix preferentially migrate along the direction of applied strain. Then, we investigated how cells autonomously remodel the matrix to create stiffness patterns favorable for durotaxis. In the presence of intercellular adhesion, cells acted collectively to stiffen the matrix, after which a small subset of cells escaped the main population and migrated outward. This behavior is reminiscent of intravasation during cancer metastasis, where cohesive cell clusters generate local matrix remodeling that facilitates the departure of more motile subpopulations. These results illustrate how matrix stiffening driven by cell cohesion and contractility regulates durotactic behavior and provide mechanistic insight into collective invasion processes relevant to cancer metastasis.

The vascular system exhibits complex, non-planar geometries that become further distorted during pathological remodeling, including arterial tortuosity and aneurysms. Although hemodynamic shear stress is a well-established regulator of vascular function, the direct effects of curvature as an intrinsic geometric cue remain poorly defined. This is largely because existing in vitro models are static and fail to capture the dynamic changes that accompany disease progression. To address this gap, we used a magnetoactive hydrogel platform that enables real-time, on-demand curvature of endothelial monolayers to reproduce clinically established tortuosity metrics. Using this system, we found that elevated curvature increased nuclear localization of yes-associated protein (YAP), with the strongest response in convex relative to concave regions of highly tortuous endothelial monolayers. This mechanosensitive response was accompanied by reduced VE-Cadherin junctional thickness and increased membrane localization of endothelial nitric oxide synthase. Together, these findings identify local curvature, independent of shear stress, as a regulator of endothelial cell mechanosensing and function, and establish a dynamic hydrogel platform for isolating geometric regulation from shear stress inputs in vascular mechanobiology.

T-cells use molecular reactions with nonequilibrium error correction, i.e., proofreading, to discriminate between nearly identical antigens with high specificity and sensitivity. These receptor binding events are known to be force sensitive, yet traditional schemes of proofreading focus on reaction kinetics alone and do not consider the role of force dependent catch/slip bond behavior or interactions with mechanically engaged coreceptors such as adhesion molecules. To address this, we propose a minimal framework for proofreading of ligand discrimination by T-cell receptors (TCRs) that uses endogenous TCR mechanosensation and substrate-mediated mechanical interactions with adhesive proteins (load sharing) to improve recognition fidelity. We leverage the catch bond behavior of cognate antigens to delay decision making and amplify TCR signaling while discarding noncognate slip bond ligands in the presence of a force. By integrating our model with existing structural and molecular data, we show that substrate mechanics regulates the transmission of active cytoskeletal forces through a molecular clutch and controls the energization of bound TCRs needed for optimal proofreading. Our work demonstrates how mechanical forces and substrate properties can augment kinetic proofreading in T-cells, suggesting biomaterial design strategies for immunotherapies that tune the mechanical microenvironment of T-cells to achieve high fidelity TCR-ligand discrimination, antigen recognition, and activation.

Amyloid fibrils are intrinsically polymorphic protein assemblies that form distinct structural strains linked to diverse biological and pathological outcomes. Yet, the principles governing how sequence encodes diverse fibril architectures, and the extent to which a given fold constrains underlying amino-acid sequence compatibility, remain poorly understood. Here, we apply generative protein design to directly interrogate the sequence-structure relationship of defined fibril architectures, using -synuclein (S), a protein known to form highly polymorphic amyloid fibrils, as a model system. Sampling sequence space under structural constraints reveals a continuous compatibility manifold in which diverse sequences encode a common amyloid architecture. De novo designed sequences assemble into fibrils, often with enhanced aggregation efficiency relative to S. A subset exhibits strain-like behaviour, including similar morphologies, efficient cross-templating, and induction of S cellular propagation, thereby functionally validating structural compatibility with the native fibril fold. Energetic analysis shows that stability is achieved through distinct but compensatory interactions, supporting a non-unique mapping between sequence and structure. Together, our results define a continuous and constrained compatibility landscape underlying amyloid strains, providing a framework for understanding the determinants of polymorphism and establishing generative protein design as a strategy to access this space, interrogate amyloid sequence-structure relationships, and engineer fibrillar protein assemblies and functional biomaterials.

Transcription is usually framed as information transfer, yet it also injects a new polymer into a crowded, confined environment. Here we demonstrate how spatial confinement to surfaces in a minimal membrane-bound transcription (MBT) system displays the physical consequences of RNA synthesis. Within a dense membrane-tethered DNA network, transcription drives co-transcriptional RNA phase separation: nascent RNA oligomerizes, gels and demixes from a surrounding fluid DNA phase, generating stable spatial patterns while mechanically remodeling the DNA layer. RNA gelation sequesters T7 RNA polymerase, whereas RNA-binding and translation-associated factors reverse gelation and restore fluidity. Thus, in the absence of downstream regulatory machinery, transcription under confinement is sufficient to trigger RNA condensation and nucleic-acid phase separation. The membrane as confining interface catalyzes the onset of DNA-RNA demixing and modulates the morphology of the resulting patterns. Since such large-scale spatial unmixing may be detrimental to cellular physiology, we suggest that one fundamental role of translation is to actively prevent condensation effects created by continuous RNA production.

Chronic inflammation drives tissue dysfunction and aging, yet the dynamic interplay between persistent inflammatory signaling and structural deterioration remains difficult to study in human-relevant systems. Here, an advanced long-term human skin platform is presented that preserves native tissue architecture and epidermal, stromal, and immune-associated molecular programs for up to 4 weeks. Using this system, sustained cytokine-driven inflammation was modeled, demonstrating chronic inflammatory transcriptional programs, progressive histopathological changes, and persistent inflammatory mediator secretion that were broadly suppressed by the JAK inhibitor tofacitinib. Using aged donor tissue, prolonged senolytic-associated treatment attenuated inflammatory and remodeling pathways. Finally, UVB exposure triggered coordinated stress and inflammatory responses that were partially mitigated using topical sunscreen, demonstrating compatibility with environmental stress modeling and topical intervention within preserved tissue architecture. Together, these findings establish a versatile human skin platform for modeling chronic inflammation, aging-associated tissue remodeling, and environmental stress, providing a translational framework for investigating skin tissue dysfunction and evaluating therapeutic interventions.

Biomolecular condensates organize cellular biochemistry, yet the principles governing their internal solvent architectures remain poorly understood. Most current models focus on macromolecular scaffolds while treating the solvent as a passive, spatially uniform background. Here, we introduce Condensate Spatial Topography via Emission Lifetimes (ConSTEL) to map the continuous solvent polarity landscape inside biomolecular condensates. Using PopZ as a model system, we show that the condensate interior contains a persistent, tunable mosaic of aqueous environments whose apparent polarity, reported by Nile Red fluorescence lifetimes, is organized by thermodynamic state and chemical cues. This microphase-separated solvent architecture defines distinct mesoscale rheological regimes, with intermediate aqueous niches supporting fast, confined tracer motion and highly polar or non-polar extremes forming a slower, viscoelastic mesh. We further demonstrate that drug-like small molecules partition non-uniformly across this landscape according to their physicochemical properties, and that exceeding local solubility limits drives "reciprocal sculpting", in which mismatched guests remodel the host solvent architecture. Together, these results highlight internal solvent organization as an active, tunable determinant of condensate material properties, molecular transport, and partitioning, and suggest that predictive models of condensate function and pharmacology would benefit from incorporating the spatial arrangement of solvent environments alongside bulk composition.

Biohybrid robots combining compliant synthetic support structures with biological actuators could enable future applications ranging from precision microsurgery to unmanned exploration. Machines actuated by living skeletal muscles are capable of adaptive behaviors, such as sensing and responding to environmental stimuli in real-time, offering functional advantages over non-biological actuators. However, typical skeletal muscle-powered biohybrid robots depend on 3D tissues which require large cell volumes and offer limited control of muscle fiber alignment, thus reducing efficiency of force generation and transduction. Here, we present a locomotive biohybrid robot powered by 2D monolayers, or thin films, of precisely aligned skeletal muscle fibers on a micropatterned hydrogel skeleton. We demonstrate how varying skeleton design parameters, ranging from material stiffness to microscale topology, impacts muscle fiber alignment and resultant actuation strains, generating forces 10X higher than previous 2D skeletal muscle actuators, improving untethered actuation longevity by [~]4500X from < 10 minutes to > 30 days, and increasing efficiency of muscle force output (force per unit volume of muscle) by 20X as compared to 3D muscles. Utilizing our optimized design for skeletal muscle thin films, we create a multi-limbed robot composed of independent muscle-powered fins capable of on/off control and frequency-dependent speed control. With these control inputs, we achieve steered multi-directional locomotion at speeds up to 4 body lengths per minute in straight movement and 1200 degrees per minute in rotational movement, highlighting potential for such actuators to be transformed into long-lasting functional soft robots.

Covalent chemistry has transformed small-molecule drug discovery, yet analogous strategies for proteins remain largely inaccessible because covalent warheads cannot be readily integrated into biologics. Conventional genetic code expansion requires engineering a dedicated aminoacyl-tRNA synthetase for each new amino acid, rendering broad warhead screening impractical. Here we introduce AminoX, a platform that bypasses this limitation through direct tRNA acylation, enabling site-specific incorporation of chemically diverse non-standard amino acids (nsAAs), including covalent warhead nsAAs compatible with scalable biologic manufacturing and multifunctional nsAAs. Using a pooled mRNA display workflow, we screened more than 2,000 warhead-position combinations in machine learning-designed de novo miniproteins targeting CTLA-4, enabling parallel interrogation of covalent chemistry, linker geometry, and incorporation site. We confirmed covalent engagement on cells together with enhanced functional blockade. Finally, we demonstrate multifunctional nsAAs that combine covalent warheads with fluorogenic reporters for real-time detection of target engagement, as well as dual nsAA incorporation for macrocyclization and fluorescent imaging of covalent binding on cell surfaces. By uniting synthetic biology, chemical biology, generative protein design, and high-throughput functional selection, AminoX compresses covalent protein engineering timelines by orders of magnitude, accelerating the development of next-generation therapeutics, biosensors, and chemical probes.

Planar cell polarity (PCP) aligns cells within the plane of a tissue through asymmetric localization of core PCP proteins. In vivo, PCP arises from integration of biochemical signaling and mechanical inputs, making it challendging to reconstruct in vitro. Here we reconstitute PCP in cultured MDCK epithelial cells using Wnt signaling, collective migration, and Prickle3 overexpression. Loss of Vangl1 disrupts reconstituted PCP, directional collective migration, and propagation of ERK traveling waves. Orthogonal manipulation of signaling and mechanics show that tissue tension contributes to establishing a polarity axis, but not its direction, whereas local Wnt input specifies polarity direction. Together, these inputs generate tissue-wide vectorial polarity. This system provides a tractable framework for dissecting how signaling and mechanics are integrated to organize tissues. TeaserWnt signaling and tissue tension reconstitute planar cell polarity in cultured epithelial cells through core-component interdependence and ERK dynamics.

Biological tissues require branched cellular architectures to maximize spatial coverage while minimizing redundancy. Yet, how cells decode local spatial information to collectively tile territories without a global blueprint remains a key open question. Here, we develop a biophysical theory of interacting branched cells, and show that coupling their growth to short-range repulsion drives efficient tiling with minimal territorial overlap. Our model predicts that the same local mechanism simultaneously suppresses long-range density fluctuations, driving the cellular collective toward hyperuniformity. We confirm these theoretical predictions with experiments on microglial patterning in the developing retina, and show that perturbations resulting in limited cell growth disrupt both tiling and fluctuation suppression. Our results reveal that two seemingly distinct optimization principles of biological patterning, large-scale regularity and efficient tiling, are intimately linked and can arise from a single growth-repulsion feedback, suggesting a general principle for self-organized tissue coverage.

Cells can respond to alterations in the abundances of specific proteins through transcriptional outputs. Synthetic approaches inspired by native post-transcriptional circuits that convert protein abundance changes into programmable gene expression would be transformative. Here, we discover and describe design principles that effectively convert protein degradation into transcriptional outputs in live cells. We define ratiometric transcriptional activation, where control over the ratio between a transcriptional inhibitor-protein of interest fusion and transcription factor enables detection of abundance changes with high sensitivity at scale. We show that ratiometric transcriptional activation can be implemented in single cells using triply orthogonal circuits or in multicellular pools, operating independently of mechanism of protein downregulation and enabling simultaneous detection of multiple protein downregulation events through outputs such as cell survival, fluorescent protein expression, or barcode sequencing. These circuits can be applied to oncogenic targets and enable discovery of new molecular glue degraders.

Ischemic stroke triggered by carotid atherosclerosis remains unpredictable because the thrombosis embolization is governed by patient-specific vascular geometry and hemodynamics that cannot be recapitulated in conventional models. Here, we engineered "Physical Twin" artery-on-chip platforms that reproduce individualized carotid anatomy with humanized subendothelial matrix composition and arterial endothelial phenotype under physiological flow patterns. We then established thrombotic microenvironments following laser-induced injury. Computational fluid dynamics-guided experiments across distinct patient geometries reveal that local shear dictates a mechanobiological hierarchy: high-shear bifurcations ([~]3,000 s-{superscript 1}) produce von Willebrand factor (VWF) A1-dependent thrombi suppressible by conformationally sensitive inhibitors, whereas low-shear stenoses generate fragile aggregates where VWF-integrin IIb{beta} coupling governs embolization overgrowth. Pulsatile flow suppresses thrombotic growth independent of mean shear. Molecular dynamics simulations reveal why conformationally sensitive nanobodies (caplacizumab) outperform shear-independent aptamers (ARC1172) in high-shear bifurcation flow zones. This Physical Twin platform provides a mechanistic blueprint for geometry-informed, personalized antithrombotic therapy selection.

Nucleic acid nanostructures provide programmable architectures for molecular delivery but remain limited by rapid nuclease degradation, poor in vivo persistence and inefficient intracellular cargo release. Here we report a mirror-image L-DNA nanocube as a biologically persistent and modular therapeutic delivery platform. The nanocube self-assembles from synthetic L-DNA oligonucleotides into a structurally defined architecture that exhibits substantially enhanced resistance to enzymatic degradation and prolonged stability under physiological conditions compared with the corresponding D-DNA nanostructure. Surface functionalization with folic acid enables selective tumour targeting in vitro and in vivo. The L-DNA nanocube supports the delivery of chemically distinct therapeutic cargos, including doxorubicin, a bortezomib prodrug and MCL1-targeting small interfering RNA (siRNA). In tumour-bearing mice, L-DNA nanocube-mediated delivery improves therapeutic efficacy while reducing systemic toxicity relative to free drug and D-DNA nanocube controls. For siRNA delivery, we engineer a pH-responsive release mechanism that promotes endosomal escape and cytosolic cargo localization, as visualized by cryo-electron tomography, resulting in efficient gene silencing. Together, these results establish mirror-image nucleic acid nanostructures as a class of biologically functional nanomaterials for programmable intracellular therapeutic delivery.