Biology subjects

Zhao, Y.

Publications and source records attributed to Zhao, Y..

At least 163 records · Page 9Linked to original sources

bioRxiv2026

Distinct and cooperative roles of host and tumor Osteopontin in colorectal cancer liver metastasis

Osteopontin (OPN) is a secreted phosphoprotein implicated in colorectal cancer liver metastasis (CRCLM), yet the distinct spatial contributions of host-and tumor-derived OPN in driving this disease remain unclear. Using a 2 x 2 genetic knockout mouse model targeting OPN in host and tumor compartments, combined with spatial transcriptomics, we investigated compartment-specific OPN functions in CRCLM. Tumor-derived OPN promotes tumor proliferation through MEK/ERK signaling. Host OPN licenses monocyte-to-macrophage differentiation, while tumor OPN polarizes macrophages towards an M2-like state. Both host and tumor OPN suppress T cells in the tumor microenvironment, whereas loss of host OPN reveals an interferon-driven, anti-tumor niche. Translational studies using OPN-blockade immunotherapy in syngeneic and patient-derived xenograft mouse models reduced tumor burden and enhanced T cell infiltration. Together, these findings redefine the OPN-myeloid paradigm in CRC and nominate OPN as a potential therapeutic target.

immunology↗

bioRxiv2026

Membrane lipid composition and endocytosis modulate Wingless release from secreting cells

Wnts are secreted signalling molecules that regulate development and adult homeostasis. Most Wnts carry a lipid moiety that must be shielded from the aqueous environment. In the secretory pathway, this is achieved by a hydrophobic tunnel in Wntless, a multipass transmembrane protein. However, the Wnt lipid moiety must be released from Wntless before Wnts can engage with Frizzled receptors on receiving cells. Here we address the cell biological basis of Wnt-Wntless dissociation, using as a model the secretion of Drosophila Wingless in wing primordia. Super-resolution microscopy shows that Wingless first reaches the apical surface before being re-internalized to reach, without Wntless, specialized Rab7/Rab4-positive endosomes. From there Wingless traffics to the basolateral membrane where it can engage with glypicans to form a basolateral gradient. Acute inhibition of endocytosis, either with a temperature-sensitive dynamin mutant or a novel optogenetic means of inhibiting clathrin, leads to apical Wingless release in abnormal punctae devoid of Wntless, suggesting that Wingless-Wntless dissociation commences at the apical surface, perhaps because of a distinct lipid composition there. Indeed, similar looking punctae are produced upon genetic abrogation of the ceramide synthase Schlank, specifically in Wingless-producing cells. These punctae resemble insoluble aggregates that form in vitro upon detergent removal. Accordingly, punctae formation can be prevented by shielding the Wingless lipid, in vivo with excess Dally-like protein (Dlp) or in vitro with liposomes. Our results show that membrane lipid composition modulates the orderly transfer of Wingless lipid from Wntless to the inner endosomal surface thus preventing aggregation and ensuring seamless secretion in the basolateral space.

cell biology↗

bioRxiv2026

Pharmacological recruitment of a VTA glutamatergic arousal circuit by the natural product BN3 drives broad-spectrum emergence from general anesthesia

Pharmacological control of emergence from general anesthesia remains strikingly limited. Clinically available antagonists are largely drug-class specific, leaving recovery dependent on anesthetic clearance rather than to an actively programmable switch in brain state. Through phenotypic screening of a structurally diverse natural-product library, we identified BN3 (L-(-)-camphor) as a broad-spectrum emergence agent. BN3 robustly shortened recovery from propofol anesthesia across species--from mice and rats to rabbits, dogs, and rhesus monkeys--and remained effective after continuous infusion. In rodents, BN3 also accelerated emergence from mechanistically distinct anesthetics, including etomidate, remimazolam, sodium {gamma} - hydroxybutyrate, midazolam, ketamine, sodium pentobarbital, and isoflurane. EEG recordings revealed a rapid cortical shift from slow-wave dominance to a wake-like high-frequency spectrum. Mechanistically, BN3 preferentially recruited ventral tegmental area (VTA) glutamatergic neurons: chemogenetic silencing abolished the BN3-evoked EEG spectral shift and strongly attenuated behavioral acceleration of emergence, whereas optogenetic activation of these neurons was sufficient to promote recovery. BN3 increased glutamate release in the nucleus accumbens and medial prefrontal cortex, and projection-specific inhibition of VTA[->]NAc or VTA[->]mPFC outputs blunted BN3-induced emergence. These findings define a VTA-centered glutamatergic arousal circuit that can be pharmacologically recruited as a common gateway for anesthetic-independent emergence.

neuroscience↗

bioRxiv2026

Cost-effective hybrid long- and short-read sequencing enables accurate somatic structural variant detection

Somatic structural variant (SSV) calling typically requires matched normal data. Incorporating relatively inexpensive short-read sequencing not only provides essential germline information but can also replace a substantial portion of long-read sequencing, thereby enabling more cost-effective somatic SV detection. Here, we present SomaSV, a hybrid sequencing framework that integrates 30x tumor long-read data with matched normal data comprising 10x long-read and 30x short-read sequencing. This design achieves high-accuracy somatic SV detection while remaining cost-competitive. Comprehensive benchmarking demonstrates that SomaSV outperforms current state-of-the-art methods by more than 13% in F1 score while reducing sequencing costs by approximately 19%. Moreover, SomaSV identifies clinically relevant cancer-associated genes, including CLDN4 and ROBO2, highlighting its potential for discovering valuable biomarkers to support early cancer screening and diagnosis. The source code for SomaSV can be accessed at https://github.com/eioyuou/SomaSV.

bioinformatics↗

bioRxiv2026

Genomic Isomerism and Environmental Adaptation in Cyanophages Infecting Freshwater Cyanobacteria

Cyanophages infecting multicellular, heterocyst-forming cyanobacteria play diverse pivotal roles in freshwater ecosystems, yet their infection strategies and genomic adaptability remain poorly understood. Here, we isolated and characterized three Caudoviricetes cyanophages (A-Lf14, A-Alj1 and A-Hlh1) specifically infecting the nitrogen-fixing cyanobacterium Anabaena sp. PCC 7120. Super-resolution microscopy revealed heterogeneous infection outcomes even among adjacent cells within the same filament, highlighting host-level defense variability. Comparative genomics placed these phages within a cluster that includes the previously isolated phages A-1(L) and N1, and revealed a conserved [~]30 kb invertible genomic region flanked by inverted repeats (IR). This region exists as two stable isomers simultaneously within phage populations, a previously unreported genomic plasticity trait in cyanophages. Consistent with such plasticity, infection kinetics were modulated by light and nitrogen availability, indicating a resource-responsive infection strategy. The novel phages encode an alkaline phosphatase of non-cyanobacterial origin, which was highly upregulated during infection indicating a role in phosphate acquisition in phosphate-limited waters. In contrast, they lack a tnpB gene present in A-1(L), which is identical in sequence to five genes in the host genome. We detected protein-coding potential for both strands of tnpB and upregulated transcription during infection, consistent with a role in this process. The 347 residues protein encoded on the tnpB reverse strand exhibited only limited similarity to other proteins or folding potential, underlining its novelty. Our work illustrates how genomic isomerism, accessory genes, and environmental sensing collectively drive functional diversification in cyanophages, providing insights into phage-host coevolution and the impact of phages on cyanobacterial blooms in dynamic freshwater ecosystems.

microbiology↗

bioRxiv2026

MNS induces antiviral protection and suppresses inflammation

BackgroundIdentifying safe and broad-spectrum antiviral and anti-inflammatory agents remains an urgent need in infectious and inflammatory diseases. Here, we demonstrated that MNS (NSC170724), a small-molecule nitrovinyl benzodioxole, enhanced antiviral defense while limiting excessive inflammation. MethodsThe antiviral activity of MNS was evaluated in multiple cell lines and mouse infection models across DNA and RNA viruses. Virus-induced and LPS-induced inflammatory responses were assessed using RT-qPCR, ELISA and western blotting. Bulk RNA-seq and ATAC-seq were performed to define transcriptional and epigenetic mechanisms. ResultsMNS significantly suppressed viral infection in vitro and improved survival in four lethal viral infection models, accompanied by reduced viral loads and attenuated tissue injury. MNS also diminished virus-triggered and LPS-triggered inflammatory cytokine production in macrophages and multiple mouse organs, and protected mice from LPS-induced endotoxic lethality. Multi-omics profiling showed that MNS broadly repressed LPS-induced inflammatory transcriptional programs and reversed chromatin accessibility gains across promoters and transcription start sites. Joint analysis of RNA-seq and ATAC-seq data demonstrated consistent downregulation of pivotal inflammatory pathways, such as NF-{kappa}B, Toll-like receptor, and TNF signaling. ConclusionsWith potent activity against viral replication and inflammation in cellular and animal models, MNS emerges as a promising candidate for the treatment of viral infections and hyperinflammatory conditions.

systems biology↗

bioRxiv2026

Functional activation reveals a repressor-gated fourth type VI secretion system in clinical Pseudomonas aeruginosa

Pseudomonas aeruginosa is a major opportunistic pathogen in which the type VI secretion system (T6SS) contributes to interbacterial competition and virulence. While most strains encode three T6SSs, we here functionally characterize a fourth system (H4-T6SS) in a clinical P. aeruginosa isolate. The H4-T6SS gene cluster is transcriptionally silent under laboratory conditions. By promoter rewiring, we demonstrate that the H4-T6SS assembles a contractile machine and mediates effector secretion and robust inter- and intraspecific antagonism. We identify a dominant pore-forming effector and its secretion pathway and a silent yet conserved nuclease effector. Genetic and biochemical analyses show that H4-T6SS expression is activated by an H4-encoded transcriptional regulator and repressed by H-NS family proteins. Analysis of over 1,200 clinical isolates reveals that the H4-T6SS is distributed across lineages and geographic regions. Finally, we observe functional crosstalk between H4- and H2-T6SSs. Together, these findings establish H4-T6SS as a functional and repressor-gated secretion system that expands the competitive arsenal of clinical P. aeruginosa.

microbiology↗

bioRxiv2026

NINJ1 mediates hepatic ischemia-reperfusion injury

Hepatic ischemia-reperfusion injury (IRI) results from interrupted perfusion to the liver and contributes to acute liver dysfunction such as following liver transplantation. Lytic cell death pathways are major drivers of IRI and the subsequent inflammatory response. The transmembrane protein ninjurin-1 (NINJ1) was identified as the key executor of terminal plasma membrane rupture across multiple lytic cell death pathways implicated in hepatic IRI. We hypothesized that NINJ1-mediated lytic cell death drives IRI and that its therapeutic inhibition would mitigate liver IRI. Using human liver specimens, we found that NINJ1 is highly expressed in human liver tissue and that its activation correlates with early allograft dysfunction in patients undergoing liver transplantation. Utilizing a segmental hepatic IRI model in mice and rats, Ninj1 genetic deletion or pharmacologic inhibition diminished acute liver injury. Mice with hepatocyte- or macrophage-specific Ninj1 knockout both had reduced hepatocellular injury following IRI, suggesting that NINJ1 within both populations contributes to the resulting liver injury. Mechanistically, we found that hepatocytes and Kupffer cells are highly susceptible to hypoxia-induced NINJ1-mediated plasma membrane rupture, which can be pharmacologically prevented. These data position NINJ1 as a potential new therapeutic target to limit hepatic IRI, with important implications for organ preservation during liver transplantation.

immunology↗

bioRxiv2026

A live-imaging system for Arabidopsis leaf primordia at early stages

Live imaging is one of the most powerful methods to reveal the morphogenesis of plant organs. However, the highly three-dimensional structure of plant organs always poses technical challenges. For example, the basal region of leaf primordia is rarely observed because of the shape of leaf primordia and the sudden shift in geometry at the point where the leaf primordium connects to the hypocotyl. In this work, we developed a new live-imaging system that is suitable for observing the developmental process of the basal region of Arabidopsis leaf primordia at early stages. Using this system, we achieved continuous observation of the basal region of early Arabidopsis leaf primordia for more than 50 hours.

plant biology↗

bioRxiv2026

Mid-infrared light produces anti-ageing effects through resonant absorption by living organisms

Aging is commonly viewed as a consequence of cumulative molecular damage and declining stress resilience. However, this perspective overlooks the possibility that aging may primarily arise from an efficiency loss of integrated cellular operation. Here, we show that aging can be systemically prevented by enhancing cellular efficiency using frequency-specific mid-infrared (MIR) light, as revealed by organismal-, cellular- and molecular-level analyses of Caenorhabditis elegans. Remarkably, super-weak 34-THz MIR light ([~]1 W mm-{superscript 2}) prolonged the median lifespan of worms by 60%, delaying aging onset and preventing abrupt "cliff-edge" mortality, without detectable thermal effects. At the cellular level, MIR exposure enhanced global gene transcription in parallel with the establishment of a mitochondrial state of high-efficiency energy metabolism, together preserving youthful cellular homoeostasis during aging. These cellular effects were associated with vibrational modes of phosphate groups in nucleic acids and mitochondrial phospholipids within the 33-35 THz range, providing a frequency-matched molecular context for MIR modulation. Together, our results support an efficiency-first, systemic anti-aging model in which frequency-specific MIR light enhances the integrated kinetic efficiency of cellular systems, spanning gene transcriptional dynamics and mitochondrial bioenergetic flux, rather than the activation of damage-repair or stress-response pathways. These findings advance our understanding of light-matter interactions in living systems and establish a non-biochemical, physical strategy for modulating aging.

biophysics↗

bioRxiv2026

Salmonella exploits metal-responsive UMPylation to enable intracellular biphasic growth and antibiotic tolerance

Salmonella develops profound antibiotic tolerance during in vivo infection, directly contributing to antimicrobial treatment failure. Although phenotypic heterogeneity in bacterial growth rates underlies this tolerance, the exact mechanisms remain elusive. Here, we reveal that Salmonella exhibits a biphasic growth pattern within macrophages, with initial antibiotic levels modulating lag phase duration. Crucially, this growth plasticity is orchestrated by metal ion homeostasis and a reversible PTM-mediated protein phase transitions. Mechanistically, under stress conditions, enhanced Mn2+ uptake activates the UMPylator YdiU, which modifies critical translational machinery components such as elongation factor TufA. These modifications induce protein aggregation, leading to translational arrest and growth stasis. Conversely, upon stress alleviation, Salmonella accumulates Mg2+ and effluxes Mn2+, then the de-UMPylase YdiV is activated, which resolves protein aggregates to enable rapid bacterial resuscitation and proliferation. This metal-responsive proteostatic switch represents an evolutionarily conserved strategy for bacterial stress adaptation, with broad implications for combating antimicrobial tolerance. SIGNIFICANCE STATEMENTAntibiotic tolerance, a key cause of treatment failure in chronic infections, is often linked to bacterial phenotypic heterogeneity, yet the underlying regulatory mechanisms remain largely understood. This study uncovers a fundamental survival strategy employed by intracellular pathogen Salmonella, where host-derived manganese acts as a critical signal to induce a reversible, dormant state. We demonstrate that stress-induced Mn2+ influx activates widespread protein UMPylation, which functions as a molecular glue to drive the aggregation of essential proteins, thereby halting bacterial growth and conferring antibiotic tolerance. Crucially, this state is reversible upon stress relief through Mn2+ efflux and subsequent de-UMPylation, enabling rapid bacterial resuscitation. As an evolutionarily conserved mechanism of adaptive tolerance, this metal-ion-responsive switch, mediated by reversible protein UMPylation, identifies a key target for developing therapeutics against persistent bacterial infections.

microbiology↗

bioRxiv2026

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↗

bioRxiv2026

How different AI models understand cells differently

AI single-cell foundation models (scFMs) are believed to be able to learn essential relations in cell transcriptomics with the attention modules in Transformer, but there is no method to reveal what they actually learned. We observed that different models may grasp different aspects of relations. To unravel the mystery, we propose scGeneLens, a framework for dissecting how scFMs perceive cells. We employed a sparse block attention to replace the original attention mechanism to concentrate attentions into a few dominant gene-gene relations, used attention propagation to trace how the relations propagate across Transformer layers, and used integrated gradients to disentangle the relative contributions of gene identity and expression in cell representations. We applied it to scFoundation and scGPT and show that they exhibit pluralistic perceptions of cells: scFoundation emphasizes relations among cell-type marker genes, resulting in stronger cell-type separability, whereas scGPT focuses more on genes involved in shared cellular pathways and core biological activities, leading to representations that generalize across conditions. The framework provides a unified lens for probing what scFMs learn about cells and offers actionable insights for the design of future cellular foundation models. Our code can be seen in https://anonymous.4open.science/r/scGeneLens-B771/.

bioinformatics↗

bioRxiv2026

Prime-InDel version 1: A web tool providing InDel markers for targeted regions narrowing

While high-density SNP genotyping enables genome-wide assays in rice (Oryza sativa L.) and other crops, PCR-based markers--particularly those derived from insertion-deletion (InDel) variations--remain crucial for fine-mapping. Currently, readily available primer information for high-density InDel rice markers is still limited. We present the first version of PrimeInDel (https://rfgb.rmbreeding.cn/search/variation/primeIndel), an online tool integrating three sets of InDel markers: an 8K-set and a 316K-set from the 3K Rice Genomes (3K-RG) project, and a 22K-set compiled from published literatures. In user cases, primers from PrimeInDel proved highly effective, narrowing a cold-tolerance QTL (qSR2) from 2.7 Mb to 200 kb and a heading-date QTL (qDeh1.1) from 239.5 kb to 83.6 kb. Our work provides the community with a robust, PCR-based InDel primer, accessible via an online data set, which will facilitate germplasm genotyping and gene identification in rice breeding.

plant biology↗

bioRxiv2026

Pan-genome and multi-omics analyses reveal the genomic basis of symbiotic evolution in the mycoheterotrophic medicinal orchid Gastrodia elata

Mycoheterotrophy represents an extreme evolutionary strategy in which plants abandon photosynthesis and become obligately dependent on fungal partners for carbon and nutrients. Gastrodia elata, a medicinal orchid forming long-term symbioses with Armillaria and Mycena, provides a compelling system to investigate the genomic basis of obligate plant-fungus mutualism. Here, we generate a chromosome-level genome assembly (1.09 Gb) of a dark-red G. elata accession and construct a pan-genome from 12 Gastrodia accessions to dissect the evolutionary consequences of mycoheterotrophy. Phylogenomic analyses reveal pervasive hybridization across cultivated germplasm and demonstrate that tuber morphology more accurately reflects intraspecific relationships than stem color, challenging the traditional color-based taxonomy. Comparative genomic analyses reveal a pronounced degeneration of photosynthetic capacity. We detect no intact nuclear-encoded rbcS loci, which would preclude assembly of the Rubisco holoenzyme and thus canonical Calvin-Benson carbon fixation. Notably, a subset of photosynthesis-related genes is retained and transcriptionally active, suggesting their co-option into non-photosynthetic regulatory roles. Despite millions of years of intimate association with fungal partners, we detect no evidence of fungal-derived horizontal gene transfer based on a multi-step phylogenomic validation pipeline, indicating that G. elata-fungus mutualism is sustained through metabolic exchange rather than genetic integration. We further identify the Gastrodia antifungal protein (GAFP) gene family as a key molecular innovation underlying symbiotic homeostasis. The recently derived Class 1 GAFP lineage exhibits distinct domain architecture, tandem expansion, and elevated expression, consistent with enhanced regulation of fungal overgrowth. Finally, we delineate a specialized nutrient-acquisition framework adapted to obligate heterotrophy, in which fungal trehalose hydrolysis supplies hexose carbon via an expanded sugar transporter protein (STP) repertoire, while organic nitrogen is assimilated through amino acid and oligopeptide transporters coupled with urease-mediated conversion, compensating for the loss of nitrate uptake and reduction pathways. Together, these results establish a pan-genomic blueprint for symbiotic adaptation and reductive genome evolution in G. elata.

genomics↗

bioRxiv2026

Regulation of CtISWI activity by AutoN and HSS domain

Chromatin remodeler imitation switch (ISWI) plays an important role in regulating chromatin structure through sliding and spacing nucleosomes. Despite the enormous progress in regulatory elements and mechanisms of the activity of ISWI in recent years, there are still some unclear structures and mechanisms in different species. Here, we studied the ATPase activity and nucleosome binding affinity of Chaetomium thermophilum ISWI (hereafter referred to as CtISWIWT) and several mutants, further proving the importance of these mutated residues in the inhibition of AutoN. We also analyzed the effects of dsDNA and ssDNA on ATPase activity of CtISWI, suggesting the potential interaction between HSS and ATPase domain. Notably, we provided a predicted structural model based on the sequence of CtISWIWT, proposing a two-step activating mechanism of conformation change and activity regulation. Taken together, our findings elucidate a different model of ISWI self-maintenance and action, providing a new mechanism of regulation supporting chromatin remodeling. HighlightsO_LIStructural modeling of ISWI: a chromatin remodeler that couples to ATP hydrolysis to slide and space composition of nucleosome. C_LIO_LIThe ATPase activity of ISWI can be stimulated by exogenous DNA, with opposite promoting effects by dsDNA and ssDNA. C_LIO_LIThe mechanism by which ISWI is activated upon binding with nucleosome has been the subject of debate, and a more comprehensive mechanism for regulation of ISWI activity. C_LI SignificanceIn the past decades, a variety of regulatory mechanisms of the activity of chromatin remodeling factor ISWI have been proposed. Based on the hypothesis of nucleosome complex structure analysis, these studies attempted to explore the mechanism of chromatin remodeling, a gene expression regulation activity. However, previous studies have basically focused on the binding and regulation mechanism of ISWI ATPase domain and nucleosomes, without mentioning the activity mode of full-length ISWI. Therefore, our study mainly focuses on the nearly full-length ISWI containing HSS domain, exploring the mechanism of the active state transition of ISWI in remodeling activities from this perspective. It enriches and supplements the research on chromatin remodeling, an important physiological activity.

biochemistry↗

bioRxiv2026

A computational framework for designing micron-scale crisscross DNA megastructures

Crisscross polymerization enables the assembly of hundreds of unique DNA origami slats into micron-sized structures with nanoscale precision. To design these megastructures, thousands of handle sequences from a fixed library must be assigned to individual slats to encode the desired binding architecture. This complexity presents two major challenges: handles must be selected to minimize parasitic interactions that compete with desired assembly, and the fabrication of hundreds of unique slats creates a substantial logistical burden. Here, we develop a unified framework that standardizes the design and fabrication of crisscross megastructures. We use an evolutionary algorithm to optimize handle assignment and minimize parasitic binding between slats, paired with a graph-based algorithm that expands the handle library. Together, these algorithms enable the assembly of large, multi-layered megastructures that otherwise would be produced at negligible yields. We have released this framework as #-CAD, an open-source graphical application that integrates these algorithms, streamlines laboratory workflows, and makes crisscross DNA origami more broadly accessible.

synthetic biology↗

bioRxiv2026

Bright calcium-modulated bioluminescent indicators for activity imaging and red photon-assisted synaptic transmission

Bioluminescent calcium sensors have unique uses in neuroscience and neuroengineering, enabling noninvasive imaging of neuronal activity and contactless activation of opsin-expressing neurons. However, the speed, range, and robustness of non-invasive imaging and rewiring or neuronal activity are limited by the radiance and dynamic range of existing bioluminescent calcium sensors. Here, we report the stepwise engineering of improved cyan-excitable red fluorescent proteins (mCyRFP4 and dCyRFP4), an improved red bioluminescent protein based on NanoLuc and CyRFP4 (Antares3), and an improved bioluminescent calcium sensor based on Antares3 (CaMBI3). Antares3 is 3-fold brighter than its predecessor, while CaMBI3 responds with an overall dynamic range of 24-fold, enabling high-sensitivity detection of calcium dynamics in muscle and neuronal tissues in vivo. Finally, a CaMBI3 variant and a red-shifted opsin ChRmine enabled red photon-assisted synaptic transmission in C. elegans. CaMBI3 thus facilitates genetically targeted non-invasive imaging and rewiring of neural activity in living animals.

neuroscience↗