Curated Optogenetic Publication Database

Abstract: Flocculation is a key technological trait in lager brewing, governing fermentation performance, yeast recovery, and beer quality. In the allo-aneuploid hybrid yeast Saccharomyces pastorianus, the genetic basis of flocculation remains poorly resolved due to its complex dual sub-genome architecture. Here, we systematically re-annotated and functionally characterized the complete FLO gene repertoire of the Group II strain CBS 1483. Thirteen FLO genes were identified, including allelic variants and a previously uncharacterized adhesin, Flo12, containing a Hyphal_reg_CWP domain instead of the canonical PA14 lectin-binding domain. Structural modeling revealed strong conservation of Ca²+-binding residues in PA14 domains, alongside repeat-region diversification likely contributing to functional variability. Using optogenetic expression in a FLO-null background, we demonstrated that SpcI-FLO9-1 and SpcI-FLO9-2_1 are the strongest drivers of flocculation, exhibiting NewFlo-like sugar sensitivity. Transcriptomic analysis during 17°P wort fermentation showed dynamic induction of these genes coinciding with flocculation onset. Surprisingly, deletion of both loci in CBS 1483 did not abolish but only delayed sedimentation in wort, accompanied by improved maltose utilization and attenuation. These findings reveal functional redundancy and compensatory mechanisms within the FLO network of lager yeast, highlighting the genetic complexity underlying flocculation, and providing a molecular framework to inform yeast selection, strain development, and optimization of the lager fermentation processes.IMPORTANCEFlocculation, the process by which yeast cells aggregate and settle, is essential for producing clear, high-quality lager beer, and for efficient yeast recovery during brewing. However, the genetic basis of this trait in lager yeast has remained poorly understood because these strains possess unusually complex hybrid genomes. In this study, we systematically identified and characterized the complete set of flocculation genes in the industrial lager yeast Saccharomyces pastorianus CBS 1483. We demonstrated that lager yeast flocculation is not controlled by a single dominant gene, but instead emerges from the combined action of several moderately active adhesion proteins that are expressed at low levels during fermentation. Surprisingly, deleting the two strongest candidate genes only delayed, rather than eliminated, sedimentation, revealing a robust compensatory network that preserves brewing performance. These findings refine the current understanding of yeast flocculation and provide a molecular framework for developing brewing strains with improved fermentation efficiency, product consistency, and flavor quality.

Abstract: Embryonic myeloid (EM) cells are the first immune-cell population to emerge during development and must disperse throughout the embryo to act as the first line of defence against infection. Although EM cells migrate directionally toward wounds, how they navigate through unwounded tissues during early colonization remains unclear. Here we show that EM cell dispersion in Xenopus embryos is driven, at least in part, by cell-on-cell migration, an important yet underappreciated phenomenon, guided by dynamic tissue flows. We have established a new ex vivo EM cell migration system that allows for live imaging, computational analyses and optogenetic manipulation. We find that local ectodermal tissue flows repolarize EM cell protrusions and bias their directional migration. Disrupting these flows, both ex vivo and in vivo, either genetically or mechanically, impairs EM cell dispersion. Our findings reveal that mechanical cues generated by surrounding tissue flows coordinate immune-cell migration during development, highlighting an overlooked mechanism by which collective tissue dynamics guide individual cell behaviour.

Abstract: Liquid-liquid phase separation (LLPS) plays a central role in intracellular compartmentalization and gene regulation. Although optogenetic optoDroplet systems are widely used to study LLPS in living cells, the mechanisms underlying the transition from dynamic liquid-like condensates to operationally defined gel-like assemblies remain poorly understood because intermediate assembly states are difficult to monitor in situ. Here, we developed a confocal laser microspectroscopy-based approach to characterize the assembly dynamics of the fused sarcomas (FUS)n-mCherry-CRY2 optoDroplet system in living NIH3T3 cells. Quantitative fluorescence analysis enabled the identification and discrimination of three distinct assembly states: monomeric, dimer/oligomer intermediate, and gel-like. Passive micro-rheology, fluorescence recovery after photobleaching (FRAP), and automated morphological analyses further demonstrated progressive reductions in molecular mobility and increasing structural rigidity during condensate maturation. Three-dimensional spatial analysis revealed that mature condensates exhibit a distinct core-shell organization consisting of an operationally defined gel-like core, a dimer/oligomer-rich intermediate layer, and an outer monomer-rich boundary layer with liquid-like properties. Upon cessation of blue-light stimulation, the outer layer rapidly dissolved, whereas the central gel-like core remained intact, indicating the acquisition of physical irreversibility. These findings establish fluorescence microspectroscopy as a quantitative approach for resolving molecular assembly states during condensate maturation and demonstrate that gelation proceeds from the condensate center toward the periphery. This framework provides new insights into pathological liquid-to-gel phase transitions associated with neurodegenerative diseases.

Abstract: Transcription involves initiation, pausing, elongation, and termination. Suppressor of Ty5 (SPT5) regulates promoter-proximal pausing and elongation, but how it orchestrates both steps during dynamic developmental changes in gene expression remains unclear. Here, using rapid optogenetic depletion in Drosophila embryos, we uncover different consequences of SPT5 removal at different developmental stages. In early embryos, SPT5 depletion causes a shift of RNA polymerase II (Pol II) from the canonical pausing site to the +1 nucleosome, which is strongly positioned. In late embryos, SPT5 depletion similarly reduces pausing at the canonical site, but the transcriptional machinery can overcome the +1 nucleosome-which appears more labile at this time point-moving into the gene body. This results in lethality and both up- and downregulation of expression, depending on the balance between Pol II entering the gene body and defective elongation. This is intensified for genes naturally increasing or decreasing their expression, indicating that SPT5 contributes to fine-tuning dynamic expression changes.

Abstract: Temporal regulation of cyclic adenosine monophosphate (cAMP) using photoactivated adenylyl cyclase (bPAC) is critical in research; however, commercial optogenetic systems are costly. Here, we present a protocol for optogenetic control of cAMP in human telencephalic organoids using an easy-to-build Arduino-driven light-emitting diode (LED) stimulation system. We describe steps for assembling the system, writing code for LED regulation, and preparing induced pluripotent stem (iPS) cells for optogenetic experiments. We then detail procedures for optogenetic activation of cAMP to control cell-fate decisions. For complete information on the generation and use of this protocol, please refer to Shimada et al.1.

Abstract: Optogenetics, a biotechnology that combines optical and genetic strategies to regulate cellular and tissue functions with high spatial and temporal precision, serves as a powerful tool-level regulatory technology widely employed to investigate cellular processes and elucidate disease mechanisms. Nanotechnology-driven optogenetics, which incorporates nanomaterials/nanostructures to improve the efficacy and broaden the applications of optogenetic systems, synergistically integrates the precision, tunability, and multifunctionality of nanotechnology with the spatiotemporal control inherent to optogenetics. The development not only enhances the flexibility and efficiency of optogenetic technology but also advances the field toward therapeutic-grade interventions. In this review, we summarize recent advances in nanotechnology-driven optogenetics, with a particular emphasis on three key areas: (1) nanostructured light sources for the precise activation of optogenetic systems, which include both externally light-stimulated systems and systems that operate independently of external light sources; (2) nanotechnology-enabled targeted delivery of light-sensitive proteins and genetic constructs to ensure efficient modulation of optogenetic pathways, which specifically involves the nanotechnology-assisted gene, protein, and recombinase enzyme delivery approaches; and (3) nanotechnology-driven therapeutic applications of optogenetics, including CAR T cell immunotherapy, cancer treatment, neurological interventions, and cardiac therapies. We further discuss the current challenges facing this emerging field and outline future research directions. This review aims not only to highlight recent breakthroughs but also to position nanotechnology-driven optogenetics as a promising tool for next-generation precision medicine.

Abstract: Engineered yeast-living drugs offer significant potential for oral protein therapy but remain limited by poor intestinal retention and lack of precise spatiotemporal control over therapeutic delivery. Here, we developed an ingestible hydrogel-encapsulated optogenetic yeast system controlled by a wearable electronic device for programmable interleukin-10 (IL-10) delivery in inflammatory bowel disease (IBD). We engineered Pichia pastoris with an EL222-based blue-light-inducible circuit achieving rapid, stringent IL-10 secretion (50% saturated concentration within just 1 h upon induction). The yeast was encapsulated in a photo-crosslinkable HAMA hydrogel that was gelated within 60 s upon blue-light exposure, exhibiting robust mechanical stability and tissue adhesion. A lightweight (2.27 g) wearable Bluetooth-controlled electronic device delivered targeted blue light transdermally, enabling simultaneous in situ hydrogel crosslinking and optogenetic activation in yeast cells. This system significantly extended the time yeast stays in the mouse gut, with reporter gene expression lasting over 5 h, while the unencapsulated control group lasted less than 2 h. In DSS-induced colitis mice, our strategy reduced proinflammatory cytokines (IL-1β, IL-6, and TNF-α) by >60%, restored intestinal barrier integrity, and normalized gut microbiota diversity to near-healthy levels, which performed significantly better than in other groups. This integrated "extracorporeal instruction-intracorporeal response" paradigm establishes a digitally programmable platform for precision live biotherapeutics. However, limited blue-light tissue penetration remains a key constraint; future work should focus on near-infrared photosensitive systems, programmable-degradation hydrogels, and wireless closed‑loop feedback to advance clinical translation.

Abstract: Endoplasmic reticulum-plasma membrane (ER-PM) contact sites are dynamic membrane interfaces that regulate essential cellular processes, including calcium signaling and lipid homeostasis. Emerging evidence suggests that contact-site geometry and organization critically influence these functions, yet existing tools cannot systematically manipulate individual geometric parameters while providing high-fidelity visualization in living cells. Here, we develop complementary chemical and optical platforms for inducible ER-PM contact-site reconstitution. A nontoxic, reversible abscisic acid-inducible system based on the plant-derived ABIcs/PYLcs pair and a rapidly reversible optogenetic iLID/SspB system enable robust visualization and dose-dependent control of contact-site assembly and disassembly. Increasing inducer dose selectively increases contact-site density and total contact area per cell without substantially changing the average size of individual contact sites. In contrast, systematic variation of tether length or tether abundance selectively increases average contact-site size and total contact area without altering contact-site density, providing orthogonal strategies to modulate contact-site density and size. Importantly, engineered contact sites recruit ER-PM contact-site-associated proteins, including MAPPER, Kv2.1, and Stromal Interaction Molecule 1 (STIM1), demonstrating key organizational features of native ER-PM contacts. MAPPER recruitment is maintained across tether lengths tested, whereas Kv2.1/STIM1 recruitment depends strongly on tether length, with longer tethers promoting recruitment by increasing intermembrane spacing. In contrast, varying tether abundance has little effect on protein recruitment, revealing distinct roles for intermembrane spacing and tether density in organizing ER-PM contact sites. Together, this work establishes a quantitative platform for engineering ER-PM contact sites with defined geometric properties and provides a framework for investigating how membrane contact-site architecture regulates cellular function.

Abstract: Spatiotemporal control over cell fate and behavior within bioprinted constructs remains a key challenge in tissue engineering. Optogenetics offers versatile potential for non-invasive regulation of biological processes. Yet, its integration within large-scale, cell-laden bioprinted materials is still limited, especially considering the spatial constraints of existing light delivery methods. In this study, we introduce a novel approach that repurposes tomographic volumetric bioprinting to enable post-printing stimulation of photosensitive protein-switches and optogenetic circuits in cells deep within hydrogel constructs. By converging different bioprinting approaches, computer vision, context-aware model generation, and synthetic biology and cell engineering, we demonstrated selective activation of a fluorescent, light-responsive protein probe within multi-material centimeter-scale constructs. Moreover, leveraging a multi-wavelength volumetric bioprinter, we further demonstrate this concept by selectively stimulating cells expressing a near-infrared optogenetic system that triggers gene expression and the induction of pancreas-specific transcription factors. The described methods provide platforms for remote, repeatable, and localized control of biological events in volumetric constructs, opening new possibilities for advanced tissue models, and dynamic tuning of cell-mediated protein production in engineered living systems.

Abstract: Inducing tumor cell pyroptosis represents a promising anticancer strategy; however, uncontrolled pyroptosis not only restricts the production of viral delivery vectors but also poses a risk of systemic damage, thereby limiting the translational application of pyroptosis-based therapies. The development of genetic tools that enable precise spatiotemporal control over pyroptosis remains challenging. To address this, we developed PyroRACS, a bioorthogonal optogenetic inducer for precise pyroptosis induction. PyroRACS utilizes an engineered red-light-activatable Cre-ON genetic switch (RACS) to drive the expression of the gasdermin D N-terminal domain, enabling tunable initiation of pyroptosis without relying on endogenous signaling pathways. We validated robust pyroptosis induction by PyroRACS in multiple cell lines. PyroRACS exhibited high controllability by selectively ablating cancer cells in vitro with precise spatiotemporal resolution. Moreover, its superior controllability enabled the production of the adenovirus vector and allowed "all-in-one" delivery of PyroRACS. In a tumor-bearing mouse model, spatially restricted induction of pyroptosis by PyroRACS resulted in effective tumor suppression, with no detectable systemic toxicity observed under the tested conditions. Collectively, PyroRACS provides a novel optogenetic tool for precise manipulation of pyroptosis, facilitating fundamental research and advancing pyroptosis-based precision oncology therapeutics.

Abstract: Optogenetic approaches enable spatiotemporal control of signaling proteins, yet their integration with microfluidic assays to study confined cell migration remains challenging. Herein, we present a protocol for the optogenetic activation of PI3K/Akt signaling in confined cells. We detail procedures for applying stimulation to induce localized Akt activation at the cell's leading edge. This protocol enables real-time manipulation of subcellular signaling dynamics during confined migration. For additional information on the use of this protocol, please refer to Lee et al.1.

Abstract: The complexity of the eukaryotic cell cycle complicates experiment design and data interpretation, limiting our understanding of how cells coordinate cell cycle processes. Traditional perturbation methods, including knockouts, deletions, and arrest-inducing chemicals, are limited by compensatory feedback interactions or pleiotropic side effects. Inducible synthetic systems offer greater specificity but often rely on external inducers, making rapid reversibility difficult. Here, we developed OPTO-Cln2, an optogenetic tool for light-controlled and reversible regulation of G1 progression in budding yeast. Using time-lapse microscopy, we show that OPTO-Cln2-strains rapidly switch between normal and altered G1 progression. Combining OPTO-Cln2 with a readout of TORC1 and PKA activity, we find that oscillatory signaling dynamics is coordinated with G1 progression. Finally, we show that OPTO-Cln2 enables at least two cycles of synchronous arrest and release in liquid cultures. This system provides a powerful approach for studying cell cycle dynamics and the coordination of cell growth with division.

Abstract: Toll-like receptors (TLRs) are single-pass transmembrane proteins that initiate innate immune responses through recognition of pathogen-associated molecular patterns, including lipopolysaccharide, flagellin, and microbial nucleic acids. In mammals, TLRs are expressed in both immune and non-immune cells, where they activate cytokine expression through the myeloid differentiation primary response 88 (MyD88) signaling pathway and subsequently engage the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) or interferon regulatory factor 3 / interferon regulatory factor 7 (IRF3/IRF7) pathways. To enable optical control of TLR function, the extracellular domains of several homodimeric TLRs, including TLR3, TLR4, and TLR5, are replaced with the photosensory core module of the bacterial phytochrome DrBphP. The resulting chimeric receptors activate the NF-κB and IRF3/IRF7 pathways in mammalian cells in a far-red-light-dependent manner. The MyD88 pathway is further reprogrammed to induce caspase activation instead of cytokine production, thereby creating a synthetic system that links TLR stimulation to caspase signaling. This strategy establishes a versatile optogenetic platform for far-red-light control of innate immune and cell death pathways.

Abstract: Microbial coculture can integrate advantages and overcome the metabolic imbalance of individual species. Programming strain interactions represents a common routine for synthetic microbial communities with distinct species, which causes difficulties and redundant workloads in interaction construction before being available as chassis hosts. This study explores yeast-bacterium consortium without engineered interactions for the co-inducible relayed synthesis of natural products. The Komagataella phaffii-Escherichia coli consortium is explored for co-growth under selected conditions. Low-level glucose- and blue light-responsive transcriptional systems are rebuilt separately for each host, allowing single-signal co-induced activation of compound synthesis in coculture. Pathway redirection, genome mining, and rewiring of key targets for acyl donor degradation result in efficient production of the reporter molecule simvastatin (26.2 mg l-1) through living consortium cultured on simple carbon source. Inducible biosynthesis of another reporter compound (2S)-naringenin (165.6 mg l-1) further validates the extendibility of this community. The described platform represents a breakthrough in engineering microbial consortium for biosynthesis.

Abstract: Primordial germ cells (PGCs) are the first cells specified in the Drosophila embryo and are precursors to the germline. Their formation requires suppression of somatic fates, achieved by degrading the receptor tyrosine kinase Torso at the posterior pole through the ubiquitin ligase adaptor germ cell-less (GCL). Although Torso is known to antagonize PGC formation, the underlying mechanisms remained unclear. Here, we combine optogenetic Ras activation and Ras effector loop mutants to show that Ras suppresses PGC formation independently of the canonical Raf/MEK/ERK pathway. We identify an unexpected early role for Torso in activating phosphoinositide 3-kinase (PI3K), generating membrane domains enriched in phosphatidylinositol (3,4,5)-trisphosphate (PIP3). Elevated PI3K activity disrupts PGC formation, while reduced PI3K activity creates ectopic PGCs. We demonstrate that GCL remodels the posterior pole membrane by suppressing Torso-dependent PI3K activation. Clearing PIP3 enables myosin II enrichment, allowing for PGC formation. Together, our findings reveal how antagonistic Torso and GCL activities establish the soma-germline boundary by organizing cortical lipids.

Abstract: Many molecular signals are known to be organized in subcellular regions termed microdomains. One example entails Rho GTPases, which control a range of cell behaviors through their functional patterning in spatiotemporal units. While fluorescent biosensors have allowed precise measurement of Rho GTPase activity in living cells, computational tools to delineate and track signaling microdomains in time-lapse image sequences of such biosensors do not exist. Here, we introduce a method that centers on the notion of activation time series coordination to identify signaling microdomains in space and time. After validating the algorithm with simulated microdomains, we show that our method identifies domains that support current hypotheses on the signaling architecture governing Rho GTPase organization and also respond to perturbation with optogenetics. A record of this paper's transparent peer review process is included in the supplemental information.

Abstract: Microtubules form dynamic cytoskeletal scaffolds essential for intracellular transport, organelle positioning, and spatial organization of signaling. Their architecture and function are continuously remodeled through the concerted actions of microtubule-associated proteins (MAPs), post-translational modifications (PTMs), and molecular motors. To precisely interrogate these processes in living systems, we developed a genetically encoded, single-component optogenetic platform for spatiotemporal control of microtubule organization and dynamics. By harnessing light-induced oligomerization to regulate microtubule association, this system supports reversible microtubule labeling and plus-end tracking, localized control of tubulin PTMs, optically regulated kinesin-driven cargo transport, and inducible microtubule severing within a unified design strategy. Using these tools, we reveal how local microtubule integrity governs lysosomal trafficking and endoplasmic reticulum (ER)-associated signaling dynamics. Collectively, this modular optogenetic toolkit bridges molecular design with cytoskeletal function, offering a versatile platform to dissect how dynamic cytoskeletal architectures coordinate intracellular organization, transport, and signaling.

Abstract: Cell therapy holds great promise for cancer immunotherapy, but its clinical efficacy is severely hindered by poor post-transplant cell survival, low homing efficiency, and host immune clearance. To address these challenges, this study develops a novel light-controlled immunotherapy strategy that integrates a red/far-red light genetic switch with single-cell encapsulation engineering. The red/far-red light (660/730 nm) reversible regulatory system enables precise spatiotemporal control over the expression of therapeutic proteins in engineered cells (e.g., CAR-T or engineered HEK 293T cells), allowing on-demand activation of anti-tumor immune responses. On this basis, a mild enzyme-mediated single-cell encapsulation technique is further employed to rapidly form a protective hydrogel coating in situ on the cell surface, thereby enhancing the survival of transplanted cells under hostile in vivo microenvironments. This strategy combines precise gene expression regulation with physical protection, improving therapeutic outcomes without the need for genomic modification of the cells. It provides a new paradigm for developing safe, controllable, and efficient cancer immunotherapy. Key features • Using a 660/730 nm red/far-red light reversible switch, deep tissue penetration enables spatiotemporal precise control of tumor-targeted therapeutic proteins. • Achieving rapid and gentle in situ gelation encapsulation of single-cell surfaces through HRP-pHLIP membrane anchoring and HA-dopamine enzymatic crosslinking. • Targeted strategies to overcome post-transplant hypoxia, inflammatory stress, and pulmonary first-pass entrapment, physically enhancing early cell survival prior to reaching the target tissue. • This experimental protocol requires at least three days.

Abstract: Regulation of gene transcription based on clustered regularly interspaced short palindromic repeats (CRISPR) is a powerful tool for constructing synthetic gene circuits in Saccharomyces cerevisiae. The current CRISPR-based regulatory approaches primarily focus on inhibiting the binding of dCas9 protein to single guide RNA (sgRNA) or blocking target site recognition. However, these regulation strategies are often at a single level, and their sensitivity still needs to be improved. In this study, the gene regulatory approaches at the translational and post-translational levels were integrated with optogenetic control patterns to attain very sensitive multi-level precision regulation of the dCas9 protein, thereby facilitating flexible regulation of transcription levels of target genes. This strategy was used to regulate the transcription levels of fluorescent proteins, resulting in up to 2.58-fold increase in the fluorescence intensity of mCherry compared to that without regulation. This CRISPR-based multi-level optogenetic system should be extremely helpful in understanding gene regulatory networks and in designing robust genetic circuits for synthetic biology.

Abstract: In cell biology, optical techniques can measure cells' internal states (biosensors) and stimulate cellular responses (optogenetics). Yet the design of all-optical experiments is often manual: a predetermined stimulus pattern is applied to cells, biosensors are measured over time, and data are processed offline. Here, we develop PyCLM, a Python-based suite enabling closed-loop measurement, image segmentation, and optogenetic control of thousands of cells per experiment. We showcase PyCLM on diverse applications, including performing feedback control on single cells and delivering developmental signaling patterns to Drosophila embryos. We compare single-cell versus tissue-scale optogenetic control of epithelial migration, revealing that fast and slow waves of receptor tyrosine kinase activity determine the direction of tissue movement, matching prior in vivo observations in zebrafish and mouse. PyCLM enables simple setup of dynamic experiments to probe cell and tissue properties and provides a first step toward real-time control of single-cell states at the tissue scale.

Abstract: Deficits in levels of phosphatidylinositol-4,5-bisphosphate [PI(4,5)P2] mediate Alzheimer's disease (AD) pathogenesis and etiology. The depletion of PI(4,5)P2 has been reported in AD in both human brain and animal models. Repletion of the synaptic pool of PI(4,5)P2, through haploinsufficiency of the degrading enzyme, Synaptojanin 1, ameliorated behavioral deficits in a mouse model of AD, in spite of accumulating amyloid. In order to refine the contribution of PI(4,5)P2 to AD, we used optogenetic translocation of the PI(4,5)P2-synthesizing enzyme, phosphoinositide phosphate 4 kinase2A (PIP4K2A) to the plasma membrane using light inducible dimerizable cryptochrome 2 (CRY-2) and the transcription factor CRY2-binding domain (CIBN) fused to the plasma membrane-targeting motif (CAAX). Spatiotemporally controlled production of phosphatidylinositol-4,5-bisphosphate [PI(4,5)P2] at the plasma membrane induced amelioration of behavioral deficits in a mouse model of AD. Imaging mass spectrometry confirmed alteration of specific PI(4,5)P2 acyl species, di-oleate, indicating that precise PI(4,5)P2 species may ultimately be leveraged for therapeutic intervention.

Abstract: Protein-protein interactions underpin virtually all biological processes in plants, from signal transduction and immune responses to development and stress adaptation. Despite their fundamental importance, the plant interactome remains far from complete, and existing maps are systematically biased by the technical limitations inherent to conventional detection platforms.

Abstract: Neurons acquire polarity by specifying one neurite as the axon, whereas the others become dendrites. But how this fundamental asymmetry is established remains unclear1. Neuronal polarization has been thought to rely primarily on growth cones that sense external cues2. Here we show that growth cones alone do not direct this process and that the soma acts as a central organizer of neuronal polarization. Using live imaging and genetic loss-of-function approaches in vivo, combined with optogenetic control and local cytoskeletal perturbations in cultured neurons, we uncover a soma-initiated oscillatory program that primes axon selection. Periodic actin branching that depends on the actin-related protein 2/3 (ARP2/3) complex at the soma remodels a global actomyosin network, thereby generating an actin wave that retracts neurites before propagating into a single neurite tip. Exposure to this wave relaxes local actomyosin contractility, which drives a transient microtubule-based protrusion and biases this neurite towards axon fate. As the cell exits this oscillatory stage, this neurite can overcome global inhibition and extend independently of ARP2/3, whereas actomyosin activity suppresses axon formation in the remaining neurites so that they subsequently become dendrites. This soma-driven mechanism ensures the emergence of a single axon independent of environmental cues and underpins the unidirectional information flow in neuronal circuits.

Abstract: Memory persistence, the ability to retain information over time, is a fundamental feature of long-term memory. Although astrocytes contribute to synaptic plasticity, the molecular mechanisms by which they support memory persistence remain unclear. Here we show that astrocytic ankyrin-2 (Ank2) is required for memory persistence in adult mice. Astrocyte-specific deletion of Ank2 impaired remote memory without affecting recent memory and disrupted the maintenance of long-term potentiation. Loss of Ank2 reduced astrocyte contacts with engram neurons and impaired astrocyte morphogenesis driven by brain-derived neurotrophic factor (BDNF) signaling through the truncated tropomyosin receptor kinase B receptor (TrkB.T1) and inositol 1,4,5-trisphosphate receptor type 2 (IP3R2). Consistent with this mechanism, astrocytic Ank2 was required for the enhancement of memory persistence by hippocampal BDNF infusion. Furthermore, selective optogenetic activation of astrocytic TrkB.T1 signaling enhanced remote memory, demonstrating that astrocytic BDNF signaling is sufficient to promote memory persistence. These findings identify astrocytic Ank2 as a key regulator of long-term memory persistence.

Abstract: Here we introduce OpenEvo, a fully open-source, low-cost turbidostat platform for automated continuous culture and directed evolution experiments. Existing tools are expensive, complex, or lack open-source hardware; OpenEvo addresses this gap with a complete, fully automated evolution platform with detailed, illustrated construction instructions for beginners, open-source software and firmware, priced around $300. An optional PC-based interface offers enhanced functionality, including remote access, programmable evolution cycles, programmable LED stimulation, and a data visualization tool. OpenEvo can cycle through three types of media for positive, negative, and neutral selection conditions, supporting a wide range of experimental designs. We validate the use of OpenEvo by evolving Haloferax volcanii to grow from 15% to 12% salt over ~150 cycles, ~1,000 hours. Evolved cells grew 55% faster than wild-type at 12% salt. Whole-genome sequencing of adapted cells found SNPs and large deletions. We also demonstrate positive and negative selection using the OpenEvo LEDs to drive optogenetics via a Phytochrome B-based optogenetic tool, with light as the selection stimulus during over 4000 hours of growth. OpenEvo lowers the technical and cost barriers for continuous evolution experiments, serves as a teaching tool, and is designed to grow an open community of users who share modifications.