Curated Optogenetic Publication Database

Abstract: Smart microscopy is transforming biological imaging by integrating real-time analysis with adaptive acquisition to enhance imaging efficiency. Whereas many emerging implementations are event-driven and focus on on-demand data acquisition to reduce phototoxicity, we here present 'outcome-driven' microscopy, a framework combining smart microscopy with optogenetics to control cell biological processes and achieve predefined outcomes. We validate this approach using light-based control of cell migration and nucleocytoplasmic transport, demonstrating robust spatiotemporal control of cellular behaviour in single cells and in cell populations.

Abstract: Gap junctions mediate rapid signal transduction between contiguous cells, which are indispensable for multicellular organisms to coordinate cellular activities across numerous physiological processes. However, precise control of gap junctions remains elusive. Herein, we present CarGAP, a single-component chemo-optogenetic tool that utilizes the C-terminal adenosylcobalamin (AdoB12) binding domain of a photoreceptor protein (i.e., CarHC) to achieve reversible control over both vertebrate and invertebrate gap junctions with spatiotemporal precision. The vertebrate CarGAP (i.e., Cx-CarGAP), created by genetically fusing connexins with CarHC in mammalian cells, can efficiently block the gap junction channels through AdoB12-induced protein oligomerization and subsequently reinstate them via green light-induced protein disassembly. We further introduced the CarGAP system (i.e., Inx-CarGAP) to the Drosophila ovary, enabling reversible control over the heterotypic gap junctions formed by innexin2 (Inx2) and innexin4 (Inx4, also known as zero population growth, Zpg), thereby uncovering the roles of gap junctions in stem cell-niche interactions. This study illustrates CarGAP as a generalizable chemo-optogenetic tool for interrogating the functions of gap junctions in various biological contexts.

Abstract: When mammalian cells are exposed to stress, they co-ordinate the condensation of stress granules (SGs) through the action of proteins G3BP1 and G3BP2 (G3BPs) and, simultaneously, undergo a massive reduction in translation. Although SGs and G3BPs have been linked to this translation response, their overall impact has been unclear. Here we investigate the question of how, and indeed whether, G3BPs and SGs shape the stress translation response. We find that SGs are enriched for mRNAs that are resistant to the stress-induced translation shutdown. Although the accurate recruitment of these stress-resistant mRNAs does require the context of stress, a combination of optogenetic tools and spike-normalized ribosome profiling demonstrates that G3BPs and SGs are necessary and sufficient to both help prioritize the translation of their enriched mRNAs and help suppress cytosolic translation. Together, these results support a model in which G3BPs and SGs reinforce the stress translation programme by prioritizing the translation of their resident mRNAs.

Abstract: Light-sensitive proteins allow organisms to perceive and respond to their environment, and have diversified over billions of years. Among these, Light-Oxygen-Voltage (LOV) domains are widespread photosensors that control diverse physiological processes and are increasingly used in optogenetics. Yet, the evolutionary constraints that shaped their protein dynamics and thereby their functional diversity remain poorly resolved. Here we systematically characterize the dynamics of 21 natural LOV core domains, significantly extending the spectroscopically resolved catalog through the addition of 18 previously unstudied variants. Using time-resolved spectroscopy, we uncover an exceptional kinetic diversity spanning from picoseconds to days and identify distinct functional clusters within the LOV family. These clusters reflect evolutionary branching, including a divergence of ≈1.0 billion years between investigatedLOV variants from plants and ≈0.4 billion years of separation within one of these functional clusters. Individual variants with extreme photocycles emerge as promising anchor points for optogenetic applications, ranging from highly efficient adduct formation to ultrafast recovery. Beyond natural diversity, we introduce a LOV domain generated by artificial intelligence-guided protein design. Despite being sequentially remote from its maternal template, this variant retains core photocycle function while exhibiting unique biophysical properties, thereby occupying a new region on the biophysical landscape. Our work emphasizes how billions of years of evolution defined LOV protein dynamics, and how protein design can expand this repertoire, engineering next-generation optogenetic tools.

Abstract: Microtubules are crucial regulators of plant development and are organized by a suite of microtubule-associated proteins (MAPs) that can rapidly remodel the array in response to various cues. This complexity has inspired countless studies into microtubule function from the subcellular to tissue scale, revealing an ever-increasing number of microtubule-dependent processes. Developing a comprehensive understanding of how local microtubule configuration, dynamicity, and remodeling drive developmental progression requires new approaches to capture and alter microtubule behavior. In this review, we will introduce the technological advancements we believe are poised to transform the study of microtubules in plant cells. In particular, we focus on (1) advanced imaging and analysis methods to quantify microtubule organization and behavior, and (2) novel tools to target specific microtubule populations in vivo. By showcasing innovative methodologies developed in non-plant systems, we hope to motivate their increased adoption and raise awareness of possible means of adapting them for studying microtubules in plants.

Abstract: LitR is a blue-green light-sensing transcriptional regulator that uses coenzyme B12 as a chromophore. In this study, we developed a genome-integrative light-inducible expression (iLiEX) system in Streptomyces griseus NBRC 13350, a Gram-positive bacterium that produces streptomycin. The system incorporates LitR, transcriptional amplification module T7 RNA polymerase, and a serine integrase. Using iLiEX, we achieved light-dependent overproduction of catechol-2,3-dioxygenase and β-glucuronidase (GUS) at levels comparable to those from a high-copy plasmid. Notably, GUS activity was 39-fold higher than with the constitutively strong ermE* promoter. The iLiEX system was also functional in S. coelicolor, S. lividans, S. albus J1074, and S. avermitilis. We improved iLiEX in two key ways: by optimizing the ribosome-binding site of T7 RNA polymerase to increase expression, and by introducing the T7 lysozyme gene to reduce leaky transcription. The system's versatility was improved by shortening the T7 promoter from 89 to 44 bp. For simple visualization on agar plates, light-dependent overexpression of fluorescent proteins, a chromogenic protein, and a brown pigment synthesis enzyme was demonstrated. High-level production of secreted enzymes, including laccase and transglutaminase, was also confirmed. Overall, we developed a single-copy light-inducible overexpression system with broad functionality across multiple Streptomyces species.

Abstract: Presynaptic accumulation of misfolded α-synuclein (α-syn) and altered synaptic transmission are considered early events in the pathogenesis of Parkinson's disease (PD), suggesting a potential causal link between these two events. However, the mechanisms by which α-syn aggregation induces synaptic dysfunction and the subsequent progressive neurodegeneration remain elusive. In the present study we leveraged the high temporal resolution of the Light-Inducible Protein Aggregation (LIPA) system in vivo and in human dopaminergic neurons to explore the early sequence of α-syn-induced pathological events leading to synaptopathy. We observed that nigrostriatal axonal transport and presynaptic accumulation of α-syn aggregates altered the activity of different neuronal populations in the mouse striatum. The results of histological and metabolite analyses show that presynaptic accumulation of α-syn induced a shift in the activation pattern of D1- and D2-expressing striatal medium spiny neurons, caused an increase in the size and density of dopaminergic synapses, and disrupted striatal dopamine signaling. Altogether, our findings reveal that the accumulation of α-syn in dopaminergic terminals triggered early presynaptic impairments, which subsequently altered striatal neuronal activity. Our study provides new insights into the molecular mechanisms underlying early synaptopathy in PD.

Abstract: The CRISPR-Cas9 system has been proven to be a powerful tool for gene editing in living cells and shows great potential in genetic disease treatment. Anti-CRISPR (Acr)-based optogenetic tools could spatiotemporally regulate the activity of CRISPR-Cas9, thereby improving the precision and safety of gene editing. However, these tools could only regulate a certain Cas9 protein because of the high specificity of Acr used, limiting their further application. In this study, we developed a new optogenetic tool named CASANOVA-A5 (CRISPR-Cas9 activity switching via a novel optogenetic variant of AcrIIA5) by inserting the blue light sensor AsLOV2 into AcrIIA5 with a broad inhibition spectrum. We proved that the CASANOVA-A5 could regulate the gene editing activity of SpCas9, SaCas9, NmeCas9, and St1Cas9 in a blue light-dependent manner. Additionally, we engineered AcrIIA5-LOV9 by integrating the blue light-dependent degron module LOV9, showing obvious optical regulation for SpCas9. Together, our work demonstrates two feasible methods to engineer the Acrs to potent optogenetic tools and suggests systematic strategies for further optimization.

Abstract: Embryonic cells must interpret morphogen signals that vary in both time and space, but the rules by which they decode these dynamics remain unclear. Here we combine optogenetics with human 2D gastruloids to define minimal WNT signaling rules for germ-layer patterning. We block endogenous WNT secretion to create a “blank canvas” and reconstitute signaling using light-gated LRP6. Systematic temporal scans reveal a narrow competence window when the onset and duration of WNT signaling specify mesoderm; this window is shifted by cell density and amplified by BMP priming, whereas identical WNT inputs outside it invert germ-layer order or generate alternative mesodermal subtypes. Using micromirror-based illumination, we restricted WNT activation to a mid-ring during this temporal window; combined with BMP4, this fully restored germ layer domains with boundaries sharper than those generated by ligand stimulation. Thus, precise spatiotemporal control of a single pathway is sufficient to optically rebuild germ-layer architecture and reveals WNT as a temporal morphogen.

Abstract: Cellular signaling cascades rely on transfer of information from one protein to another or within a single protein. To facilitate signal integration, specific structural motifs evolved that allow signal processing and also enable modular downstream response integration, facilitating sophisticated regulatory mechanisms. On a structural level, especially coiled-coil helices are frequently observed as signaling motifs. In diguanylate cyclases (DGCs) featuring GGDEF domains, N-terminal coiled-coils frequently activate systems by rearrangements of the interdimer active site. The variety of sensory domains that modulate this structural equilibrium in response to different stimuli highlights the importance of DGCs in bacterial adaptation. One interesting example of sensor DGCs is blue light-activated light-oxygen-voltage (LOV)-GGDEF couples. Here, we describe molecular details of a two-stage mechanism that allows tight dark-state inhibition while enabling high enzymatic activities upon illumination, achieving fold changes exceeding 10,000-fold. Using an in vivo activity assay, we screened amino acid substitutions at the inhibitory interface and the sensor-effector linker region to identify variants that promote enzymatic activity in the dark. In combination with chimeras of LOV and GGDEF domains preventing inhibitory interface formation, we successfully stabilized elongated active-state conformations and confirmed the role of the inhibitory interface between sensor and effector in the tight dark-state inhibition. Interestingly, the initially generated chimeras are still light regulatable as long as the linker sequence is not stabilized in either inhibiting or stimulating coiled-coil register. Our results offer valuable insights for potential optogenetic applications but also demonstrate inherent challenges associated with Methylotenera sp. LOV-activated DGCs.

Abstract: Tumor initiation remains one of the least understood events in cancer biology, largely due to the challenge of dissecting the intricacy of the tumorigenic process in laboratory settings. The insufficient biological complexity of conventional in vitro systems makes animal models the primary experimental approach to study tumorigenesis. Despite providing valuable insights, these in vivo models function as experimental black boxes with limited spatiotemporal resolution of cellular dynamics during oncogenesis. In addition, their use raises ethical concerns, further underscoring the need for alternative ex vivo systems. Here we provide a detailed protocol to integrate state-of-the-art microfabrication, tissue engineering and optogenetic approaches to generate topobiologically complex miniature colons ('mini-colons') capable of undergoing tumorigenesis in vitro. We describe the key methodology for the generation of blue light-inducible oncogenic cells, the establishment of hydrogel-based mini-colon scaffolds within microfluidic devices, the development of mini-colons and the induction of spatiotemporally controlled tumorigenesis. This protocol enables the formation and long-term culture of complex cancerous tissues that capture in vivo-like tumoral biology while offering real-time and single-cell resolution analyses. It can be implemented in 4-6 weeks by researchers with prior experience in 3D cell culture techniques. We anticipate that these methodological guidelines will have a broad impact on the cancer research community by opening new avenues for tumorigenesis studies.

Abstract: Saccharomyces cerevisiae is a widely used chassis in metabolic engineering. Due to the Crabtree effect, it preferentially produces ethanol under high-glucose conditions, limiting the synthesis of other valuable metabolites. Conventional metabolic engineering approaches typically rely on irreversible genetic modifications, making it insufficient for dynamic metabolic control. In contrast, optogenetics offers a reversible and tunable method for regulating cellular metabolism with high temporal precision. In this study, we engineered the pyruvate decarboxylase isozyme 1 (Pdc1) by inserting the photosensory modules (AsLOV2 and cpLOV2 domains) into rationally selected positions within the enzyme. Through a growth phenotype-based screening system, we identified two blue light-responsive variants, OptoPdc1D1 and OptoPdc1D2, which enable light-dependent control of enzymatic activity. Leveraging these OptoPdc1 variants, we developed opto-S. cerevisiae strains, MLy-9 and MLy-10, which demonstrated high efficiency in modulating both cell growth and ethanol production. These strains allow reliable regulation of ethanol biosynthesis in response to blue light, achieving a dynamic control range of approximately 20- to 120-fold. The opto-S. cerevisiae strains exhibited dose-dependent production in response to blue light intensity and pulse patterns, confirming their potential for precise metabolic control. This work establishes a novel protein-level strategy for regulating metabolic pathways in S. cerevisiae and introduces an effective method for controlling ethanol metabolism via optogenetic regulation.

Abstract: Light Oxygen Voltage (LOV) domains are important widespread receptors of blue light that also found applications in optogenetics and imaging. While LOV domains from mesophiles are relatively well characterized, their counterparts from thermophilic microorganisms remain understudied. Here, we express two constructs of a LOV domain belonging to a histidine kinase from Meiothermus ruber, MrLOV and MrLOVe, and show that they are photoactive, with recovery time values of 21 and 27 min, respectively, and thermostable. Crystal structures reveal that MrLOV, which lacks helices A'α and Jα, forms a parallel dimer, whereas MrLOVe is a tetramer organized as an antiparallel dimer of two parallel dimers interacting via helices Jα. One MrLOVe dimer is symmetric, and the other is asymmetric, with conformational differences mirroring activation-related changes in other LOV domains. Our data provide the structural basis for understanding and engineering of thermophilic LOVs and pave the way for development of thermostable and photostable LOV-derived optogenetic tools and flavin-based fluorescent proteins.

Abstract: Multimerization and phase separation represent two paradigms for organizing receptor tyrosine kinases (RTKs). However, their functional distinctions from the perspective of biomolecular organization remain unclear. Here, we present CORdensate, a light-controllable condensation system combining two synergistic photoactuators: oligomeric Cry2 and heterodimeric LOVpep/ePDZ. Engineering single-chain photoswitches, we achieve four biomolecular organization patterns ranging from monomerization to phase separation. CORdensate exhibits constant assembly and disassembly kinetics. Applying CORdensate to mimic pathogenic RTK granules establishes the role of phase separation in activating ALK and RET. Moreover, assembling ALK and RET through varying organization patterns, we highlight the superior organizational ability of phase separation over multimerization. Additionally, CORdensate-based RTK granules suggest that phase separation broadly and robustly activates RTKs. This study introduces a optogenetic tool for investigating biomolecular condensation.

Abstract: The robustness and broad applicability of an optogenetic tool depends heavily on the properties of the underlying photoreceptor protein and its cognate binding partner - the light responsive ‘core’. Current red light optogenetic systems for use in mammalian cells all rely on phytochrome based photoreceptors. These are large (70 kDa) proteins that act as dimers, thereby enforce dimerization on attached proteins. Naturally occurring or engineered binding partners can function effectively in certain cases, but large size, complex mode of interaction, background binding, relatively weak affinity and/or low fold changes between on and off states are significant limitations. Using structure-based design and directed evolution we developed a small (17 kDa) monomeric bilverdin binding photoreceptor FenixS, and a highly selective, high-affinity binder, Ash1 (6 kDa). Negligible off-state binding and a >1200-fold increase in binding affinity upon 700 nm illumination result in a high performance, ultra-low background, light responsive core for a diverse range of applications. An optogenetic tool for red light activation of transcription in mammalian cells based on the FenixS-Ash1 core exhibits robust performance without the need for biliverdin supplementation.

Abstract: The efficacy of antitumor immunotherapy is closely associated with the expansion of tumor-infiltrating CD8+ T cells. However, within the tumor microenvironment, CD8+ T cells often exhibit reduced proliferation due to persistent exposure to tumor antigens. The cytokine IL-2 is a potent growth factor that can drive the expansion of tumor-infiltrating lymphocytes. While its clinical application has been severely limited by systemic toxicity and in vivo instability. To address these challenges, we have developed a dual-responsive system (EcNIL-2@UCNP/Gel-CTX) leveraging the hypoxic tropisms of E. coli Nissle 1917(EcN). This system is capable of producing IL-2 in situ upon near-infrared (NIR) irradiation and releasing low-dose cyclophosphamide (CTX) in response to matrix metalloproteinase-2 (MMP-2) in the tumor microenvironment. The EcNIL-2@UCNP/Gel-CTX system not only drives the expansion of CD8+ T cells and boost the activity of NK cells but also reduces Treg cell populations, thereby remodeling the immune microenvironment and eliciting robust tumor-specific immune responses in H22 subcutaneous tumors in mice and confers long-term protection against tumor rechallenge by promoting the generation of durable memory T cells. Our findings provide an both light and tumor microenvironment responsive platform for enhanced cancer immunotherapy.

Abstract: The modification of key enzymes for chemical production plays a crucial role in enhancing the yield of targeted products. However, manipulating key nodes in specific signalling pathways remains constrained by traditional gene overexpression or knockout strategies. Discovering and designing optogenetic tools enable us to regulate enzymatic activity or gene expression at key nodes in a spatiotemporal manner, rather than relying solely on chemical induction throughout production processes. In this review, we discuss the recent applications of optogenetic tools in the regulation of microbial metabolites, plant sciences and disease therapies. We categorize optogenetic tools into five classes based on their distinct applications. First, light-induced gene expression schedules can balance the trade-off between chemical production and cell growth phases. Second, light-triggered liquid-liquid phase separation (LLPS) modules provide opportunities to co-localize and condense key enzymes for enhancing catalytic efficiency. Third, light-induced subcellular localized photoreceptors enable the relocation of protein of interest across various subcellular compartments, allowing for the investigation of their dynamic regulatory processes. Fourth, light-regulated enzymes can dynamically regulate production of cyclic nucleotides or investigate endogenous components similar with conditional depletion or recovery function of protein of interest. Fifth, light-gated ion channels and pumps can be utilized to investigate dynamic ion signalling cascades in both animals and plants, or to boost ATP accumulation for enhancing biomass or bioproduct yields in microorganisms. Overall, this review aims to provide a comprehensive overview of optogenetic strategies that have the potential to advance both basic research and bioindustry within the field of synthetic biology.

Abstract: The soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein syntaxin mediates neuronal exocytosis and self-assembles into large clusters in the plasma membrane. The formation and function of these clusters, and whether they promote or inhibit synaptic-vesicle fusion, remain unclear. Here using optogenetic control of syntaxin clustering in vitro and in vivo, as a light-inducible gain-of-function assay, we show that light-enhanced clustering reduces both spontaneous and triggered vesicle fusion, and this impairs mouse hunting behavior. Cluster formation is induced by liquid-liquid phase separation (LLPS) of the SNARE domain of syntaxin. For the regulatory mechanism, Munc18, which is known to alter syntaxin conformation, acts to reduce LLPS for cluster formation, thereby promoting active syntaxin. These results suggest that exocytosis regulation involves LLPS-induced syntaxin clusters that serve as a syntaxin reservoir from which Munc18 captures syntaxin monomers to form a syntaxin-Munc18 complex, setting the stage for efficient fusion.

Abstract: Coenzyme Q10 biosynthesis in Escherichia coli is constrained by kinetic mismatches between precursor synthesis and methylation, alongside bioenergetic uncoupling. We implemented an optogenetic phase-control strategy integrating dynamic light induction, ribosome binding site (RBS) engineering, and real-time membrane potential (ΔΨ) feedback. Temporal coordination of 1-deoxy-D-xylulose-5-phosphate synthase (DXS) and UbiG methyltransferase (UbiG) via a 6-h phase delay reduced methylglyoxal shunt flux by 41 ± 3% (p < 0.01) through enhanced precursor channeling. Membrane hyperpolarization to - 90 ± 2 mV (relative to - 70 mV in controls) triggered voltage-gated UbiG membrane localization (62 ± 3%) and ATP-driven S-adenosylmethionine regeneration, increasing methylation efficiency 2.3-fold. Multivariate modeling identified ΔΨ and acetate as critical control parameters, enabling optimized fermentation (dissolved oxygen (DO) 15-20%, pH 6.7-6.9). The engineered strain achieved 0.63 ± 0.07 g/L CoQ10 in 5-L bioreactors-a 4.3-fold improvement over the static control strain (0.15 ± 0.02 g/L)-with 82.5% carbon efficiency and 25.8% glycerol-to-product yield. This work establishes bioenergetically coupled temporal control as a scalable paradigm for membrane-bound isoprenoid biomanufacturing. KEY POINTS: • Phase-driven enzyme synchronization via optogenetics resolves kinetic mismatch. • Membrane hyperpolarization gates enzyme localization and ATP regeneration. • Model-integrated bioenergetic-process control enhances CoQ10 production efficiency.

Abstract: Protein homeostasis, or proteostasis, is essential for cellular proteins to function properly. The buildup of abnormal proteins (such as damaged, misfolded, or aggregated proteins) is associated with many diseases, including cancer. Therefore, maintaining proteostasis is critical for cellular health. Currently, genetic methods for modulating proteostasis, such as RNA interference and CRISPR knockout, lack spatial and temporal precision. They are also not suitable for depleting already-synthesized proteins. Similarly, molecular tools like PROTACs and molecular glue face challenges in drug design and discovery. To directly control targeted protein degradation within cells, we introduce an intrabody-based optogenetic toolbox named Flash-Away. Flash-Away integrates the light-responsive ubiquitination activity of the RING domain of TRIM21 for protein degradation, coupled with specific intrabodies for precise targeting. Upon exposure to blue light, Flash-Away enables rapid and targeted degradation of selected proteins. This versatility is demonstrated through successful application to diverse protein targets, including actin, MLKL, and ALFA-tag fused proteins. This innovative light-inducible protein degradation system offers a powerful approach to investigate the functions of specific proteins within physiological contexts. Moreover, Flash-Away presents potential opportunities for clinical translational research and precise medical interventions, advancing the prospects of precision medicine.

Abstract: Precise control of gene expression is one of the fundamental goals of synthetic biology. Whether the objective is to modify endogenous cellular function or induce the expression of molecules for diagnostic and therapeutic purposes, gene regulation remains a key aspect of biological systems. Over time, advances in protein engineering and molecular biology have led to the creation of gene circuits capable of inducing the expression of specific proteins in response to external stimulus such as light. These optogenetic, or light-activated circuits hold significant potential for gene therapy as a tool for regulating the expression of therapeutic genes within cells. However, the applications of optogenetic systems can be limited by the lack of efficient ways for light delivery inside cells or tissue. Our approach to address this challenge is to harness the power of bioluminescence to produce light directly inside cells using a luminescent enzyme. Combined with a photosensitive transcription factor, we report the development of a fully genetically encoded optogenetic circuit for control of gene expression. Furthermore, we utilized a magneto sensitive protein to engineer a split protein version of this luminescent enzyme, where its reconstitution is driven by a 50mT magnetic stimulus. Thus, resulting in a first-of-its-kind gene circuit activated by a combination of light and magnetic stimulus. We expect this work to advance the implementation of light-controlled systems without the need of external light sources, as well as serve as a basis for the development of future magneto-sensitive tools.

Abstract: Regulation of cancer cells by their environment contributes to tumorigenesis and drug response, though the extent to which the oncogenic state can alter a cell's perception of its environment is not clear. Prior studies found that EML4-ALK, a receptor tyrosine kinase (RTK) fusion oncoprotein, suppresses transmembrane receptor signaling through EGFR. Moreover, suppression was reversed with targeted ALK inhibition, thereby promoting survival and drug tolerance. Here we tested whether such modulation of EGFR was common among other RTK fusions, which collectively are found in ∼5% of all cancers. Using live- and fixed-cell microscopy in isogenic and patient-derived cell lines, we found that a wide variety of RTK fusions suppress transmembrane EGFR and sequester essential adaptor proteins in the cytoplasm, as evidenced by the localization of endogenous Grb2. Targeted therapies rapidly released Grb2 from sequestration and potentiated EGFR. Synthetic optogenetic analogs of RTK fusions confirmed that cytoplasmic sequestration of Grb2 was sufficient to suppress perception of extracellular EGF and could do so without driving signaling from the synthetic fusion itself, demonstrating that fusion signaling and suppression of EGFR could be functionally decoupled. Our study uncovers that a large number of RTK fusions simultaneously act as both activators and suppressors of signaling, the mechanisms of which could be exploited for new biomimetic therapies that enhance cell killing and suppress drug tolerance.

Abstract: Intracellular optogenetics represents a rapidly advancing biotechnology that enables precise, reversible control of protein activity, signaling dynamics, and cellular behaviours using genetically encoded, light-responsive systems. Originally pioneered in neuroscience through channelrhodopsins to manipulate neuronal excitability, the field has since expanded into diverse intracellular applications with broad implications for medicine, agriculture, and biomanufacturing. Key to these advances are photoreceptors such as cryptochrome 2 (CRY2), light-oxygen-voltage (LOV) domains, and phytochromes, which undergo conformational changes upon illumination to trigger conditional protein-protein interactions, localization shifts, or phase transitions. Recent engineering breakthroughs-including the creation of red-light responsive systems such as MagRed that exploit endogenous biliverdin-have enhanced tissue penetration, minimized phototoxicity, and expanded applicability to complex biological systems. This review provides an overarching synthesis of the molecular principles underlying intracellular optogenetic actuators, including the photophysical basis of light-induced conformational changes, oligomerization, and signaling control. We highlight strategies that employ domain fusions, rational mutagenesis, and synthetic circuits to extend their utility across biological and industrial contexts. We also critically assess current limitations, such as chromophore dependence, light delivery challenges, and safety considerations, so as to frame realistic paths towards translation. Looking ahead, future opportunities include multi-colour and multiplexed systems, integration with high-throughput omics and artificial intelligence, and development of non-invasive modalities suited for in vivo and industrial applications. Intracellular optogenetics is thus emerging as a versatile platform technology, with the potential to reshape how we interrogate biology and engineer cells for therapeutic, agricultural, and environmental solutions.

Abstract: Epigenome editing has become a leading-edge technology of programmable, heritable and reversible control of gene expression in plants without changing the DNA sequence. CRISPR/dCas9 systems along with transcription activator-like effectors (TALEs) and zinc finger systems have made it possible to manipulate DNA methylation, histone modifications, and RNA epigenetic marks in a precise and locus-specific fashion. These tools have been used on major regulatory genes of flowering time, stress adjustment, and yield maximization in model and crop plants. This review synthesizes the current status of plant epigenome editing advances and highlights mechanistic innovations including SunTag, CRISPRoff/on and RNA m6A editing. It also emphasizes new paradigm shifts in chromatin reprogramming, including transcription-resistive chromatin states, locus-specific H3K27me3 demethylation, and nanobody-mediated chromatin targeting. Furthermore, it considers the consequences of these shifts in the context of trait stability and epigenetic inheritance. Moreover, the relative evaluation of dCas9-, TALE-, and ZFP-based platforms indicated that there are still enduring problems in the performance of delivery, off-target effects, and transgenerational stability. The review concludes with a conceptual framework connecting epigenome editing to climate-smart crop improvement and outlines future research priorities focused on combinatorial multi-omics integration and the development of environmentally responsive editing platforms.

Abstract: Cancer can result from abnormal regulation of cells by their environment, potentially because cancer cells may misperceive environmental cues. However, the magnitude to which the oncogenic state alters cellular information processing has not been quantified. Here, we apply pseudorandom pulsatile optogenetic stimulation, live-cell imaging, and information theory to compare the information capacity of receptor tyrosine kinase (RTK) signaling pathways in EML4-ALK-driven lung cancer (STE-1) and in non-transformed (BEAS-2B) cells. The average information rate through RTK/ERK signaling in STE-1 cells was less than 0.5 bit/hour, compared to 7 bit/hour in BEAS-2B cells, but increased to 3 bit/hour after oncogene inhibition. Information was transmitted by 50-70% of cells, whose channel capacity (maximum information rate) was estimated through in silico protocol optimization. In BEAS-2B cells, channel capacity of the parallel RTK/calcineurin pathway surpassed that of the RTK/ERK pathway. This study highlights information capacity as a sensitive metric for identifying disease-associated dysfunction and evaluating the effects of targeted interventions.