Curated Optogenetic Publication Database

Abstract: Inducible CRISPR/Cas systems enable spatiotemporal control of genome editing in response to chemical, optical, biological, or physical stimuli. By restricting genome-editing activity to defined conditions, these systems may reduce off-target exposure and immune burden while improving tumor-selective control, making them attractive tools for precision oncology.

Abstract: Chimeric antigen receptor (CAR) T-cell therapy has revolutionized the treatment of hematologic cancers but encounters challenges, including severe treatment-related toxicities, a highly suppressive tumor microenvironment (TME), limited long-term persistence, and poor trafficking/infiltration into solid tumors. This review outlines recent genetic engineering strategies to address these issues and enhance the safety, durability, and efficacy of CAR-T cell therapy. To reduce cytokine release syndrome and neurotoxicity, methods such as affinity-tuned and humanized scFvs, hinge/TM optimization, and ITAM calibration have been developed, along with programmable "switch-off" and "switch-on" systems that include suicide genes, antibody-bridging switches, and optogenetic or hypoxia-gated circuits. TME remodeling strategies utilize nanomaterials for targeted cytokine delivery, cell-surface "backpack" systems, and engineered oncolytic viruses that release cytokines or checkpoint-blocking agents. For durability and resistance to exhaustion, precise genome engineering techniques, including CRISPR-based editing and multiplexed shRNA platforms, were employed to target inhibitory receptors and exhaustion-driving transcriptional programs. Additionally, chemokine-receptor engineering and local biomaterial-based delivery systems are discussed as ways to enhance CAR-T trafficking and intratumoral persistence. These innovations collectively point toward integrated, patient-specific CAR-T platforms that incorporate safety controls, metabolic and transcriptional flexibility, and enhanced trafficking through the TME to broaden clinical use.

Abstract: Cells in many naturally occurring organisms routinely cooperate to control their extracellular pH in a dynamic and reversible manner, but this capability has been underexplored in synthetic biology. Here, we sought to engineer a microbial system that switches between two states -high and low extracellular pH- with minimal human intervention. We accomplished this by combining: (1) a genetic circuit that produces recombinant urease under the control of a light-inducible promoter; (2) a degradation tag on urease to accelerate the high-to-low pH transition; and (3) optimization of several environmental factors, including media composition, replenishment rate, and light exposure patterns. The system raises the pH when urease is produced and hydrolyzes urea in the media to produce ammonia; it lowers the pH as a byproduct of the cell's native metabolism when urease production ceases. We demonstrate that the optimized system cycles continuously for up to 14 days with minimal performance loss. Overall, our system demonstrates synthetic pH control in an engineered living system and highlights challenges and potential solutions for using such systems outside of the context of typical laboratory manipulation.

Abstract: Optogenetic tools-light-responsive proteins that enable to regulate specific cellular activities, study biological processes, and develop new therapies-are attractive approaches for achieving endogenous gene regulation under minimally invasive conditions. Our first step in constructing an optogenetic system to regulate endogenous Drosophila gene expression was to identify inhibitory anti-CRISPR (Acr) proteins that block CRISPRa-mediated activation. Next, we inserted optogenetic protein LOV2 into these Acrs, tested for their ability to optogenetically modulate endogenous gene upregulation through the CRISPRa-based flySAM system in Drosophila, and found that the photoswitchability of these prototypes was weak. We therefore engineered an optimized Acr-LOV2 fusion module by refining length of intrinsically disordered and ordered regions (IDR and IOR) of Acrs. This optimization yielded a variant with significantly greater sensitivity to blue-light-induced endogenous gene upregulation than the prototypes, leading to new in vivo discoveries. In addition, this work provides insights for in vivo functional characterization of the IDR and the IOR of these small-sized proteins. Together, these findings establish a robust optogenetic toolbox for precise, light-controlled endogenous gene regulation in Drosophila.

Abstract: Tumor recurrence, metastasis, and therapeutic resistance remain major challenges in oncology, driving the need for advanced therapeutic strategies with improved precision and controllability. Optogenetics, which enables light-mediated regulation of cellular functions, has emerged as a promising modality for cancer therapy by offering unparalleled spatiotemporal precision. This capability allows dynamic control of intracellular signaling and transgene expression, enabling selective targeting of malignant cells while minimizing damage to surrounding tissues. However, clinical translation is hindered by key challenges, including inefficient in vivo delivery of optogenetic components, limited tissue penetration of activating light, and suboptimal performance of existing tools. Addressing these barriers requires a convergence of molecular engineering and materials science, wherein advanced biomaterials play a critical role in enabling gene delivery and overcoming tissue-penetration limitations in complex tumor environments. In this review, we provide a comprehensive oriented overview of optogenetics in oncology. We first analyze the molecular mechanisms and engineering principles of representative optogenetic tools, with a focus on LOV- and CRY2-based systems. We then highlight recent advances in biomaterial-assisted optogene delivery and light delivery strategies, emphasizing their material-dependent mechanisms that enable precise spatiotemporal control in vivo. Furthermore, we summarize emerging preclinical applications in cancer immunotherapy, gene regulation, and intracellular signaling control. Finally, we discuss key challenges in biosafety, kinetic optimization, and clinical scalability, and outline future directions that integrate optogenetics with functional materials and intelligent design to realize clinically viable platforms. This review aims to provide a framework for the development of clinically viable optogenetic platforms for next-generation cancer therapy.

Abstract: The cytoskeleton is a dynamic intracellular network that governs cell shape, migration, division, and mechanotransduction. Precise spatiotemporal control of cytoskeletal regulation is essential for understanding how these processes are coordinated in physiology and disease, yet conventional pharmacological and genetic approaches often lack sufficient resolution or reversibility. Optogenetic technologies provide a powerful alternative by enabling light-controlled, noninvasive manipulation of cytoskeletal regulators with high temporal precision and subcellular specificity. This review summarizes recent advances in genetically encoded optogenetic tools for interrogating cytoskeletal dynamics. We discuss core design strategies, including allosteric regulation, light-induced oligomerization, heterodimerization, and dissociation, and highlight representative applications targeting actin filaments, microtubules, and upstream signaling pathways such as Rho family GTPases. We conclude by outlining current limitations and emerging directions, including improved tissue penetration, reduced phototoxicity, and multiplexed optical control, which are expected to further expand the utility of optogenetics in cytoskeleton research.

Abstract: Photosensory protein domains, derived from nature, are foundational for optogenetic protein engineering. Tailoring their properties enables their full exploitation for optogenetic regulation in basic research and applied bioengineering applications. Here, we present a simple, yet powerful strategy based on random mutagenesis coupled to high-throughput screening that allowed altering the most fundamental properties of the widely used nMag/pMag photodimerization system: its light sensitivity and activation. Variants were characterized in vivo in bacteria by flow cytometry and during the entire growth curve by spectrofluorometry. We identify mutations that either increase or decrease the light sensitivity at sub-saturating light intensities, while also improving the light activation and dark-to-light fold change. Notably, light sensitivity and activation levels could be changed independently. In addition, we demonstrated that the shapes of the dose-response curves can be finely tuned. This broadens the applicability of the Magnets photosensors for optogenetic regulation strategies.

Abstract: An optogenetic split-GAL4 system, ShineGAL4, allows genes to be manipulated with unprecedented spatiotemporal precision. Here, we convert a panel of 14 GAL4 drivers widely used in Drosophila research into their ShineGAL4 counterparts. Homology assisted CRISPR knock-in (HACK) is used to replace GAL4 with the GAL4 DNA binding domain fused to a Magnet photoswitch. We show that the resulting ShineGAL4 drivers enable gene expression to be rapidly induced by light specifically in fat body, muscles, enterocytes, oenocytes, Malpighian tubules, neurons, neuroblast lineages, glial subtypes or in all glia. We also develop an optogenetic cassette for photoactivation of GAL4 in 'silent' FLP-out clones. This panel of optogenetic tools will enable precise spatiotemporal control of gene expression in a wide range of different Drosophila tissues and cell-types.

Abstract: Microbial optogenetic tools can regulate gene expression with spatial and temporal precision, offering excellent potential for single-cell resolution studies. However, bacterial optogenetic systems have primarily been deployed for population-level experiments. It is not always clear how these tools perform in single cells, where stochastic effects can be substantial. In this study, we focus on optogenetic Cre recombinase and compare the performance of three variants (OptoCre-REDMAP, OptoCre-Vvd, and PA-Cre) for their population-level and single-cell activity. We quantify recombination efficiency, expression variability, and activation dynamics using reporters which produce changes in fluorescence or antibiotic resistance following light-induced Cre activity. We find that optogenetic recombinase performance can be reporter-dependent. Further, single-cell analysis reveals highly heterogeneous activity, with substantial variation in the efficiency and timing of recombinase activity from cell to cell. These findings suggest important criteria for selecting optogenetic recombinases and indicate areas for optimization to improve single-cell capabilities of bacterial optogenetic tools.

Abstract: Synthetic microbial consortia (SMCs) represent a paradigm shift from monocultures to multi-strain systems that leverage ecological interactions for enhanced environmental adaptation and bioproduction. This review systematically sorts out engineering strategies for constructing stable SMCs, focusing on three core principles regarding host selection based on obligate mutualism (e.g., auxotrophs), pathway modularization to resolve metabolic conflicts, and dynamic regulation using tools like quorum sensing and optogenetics. We demonstrate the efficacy of SMCs in diverse applications including high-value compound synthesis and lignocellulosic biomass conversion through consolidated bioprocessing and inhibitor mitigation. SMCs enabling advanced functions in engineered living materials, environmental remediation, and biomedical innovation via division of labor are also described. Despite such progress, challenges in scalability and real-time control of SMCs under industrial conditions remain. We conclude that SMCs serve to bridge evolutionary ecology and biotechnology, offering robust solutions for sustainable biomanufacturing and beyond.

Abstract: Various blue-light photoreceptor proteins have photo-responsive domains known as light, oxygen, voltage (LOV) domains, which are extensively distributed in plants, algae, fungi, and bacteria. When exposed to blue light, the flavin chromophore and a highly conserved cysteine residue form a covalent adduct on a microsecond time scale. LOV domains are common photosensory modules that can be applied to optogenetics, regulated synthesis of reactive oxygen species, and fluorescence microscopy. This review explores the photocycle kinetics and applications of various LOV domains, which have been explored for confocal microscopy, two-photon microscopy, and super-resolution microscopy. Many LOV domains have been derived and modulated for use in different types of microscopic applications. Molecular understanding, diversity of LOV domains, and versatile photo-physical characteristics of these proteins have immense potential for the development of useful probes for various microscopy tools. There is a great demand for perspective research on LOV domain proteins for harnessing their possible optobiotechnological applications.

Abstract: Single-cell resolution studies have transformed our understanding of microbial systems, revealing substantial cell-to-cell heterogeneity and complex dynamic behaviors. This review describes recent advances in using optogenetics, where light-sensitive proteins control cellular processes, to investigate microbial behavior at the individual cell level. We discuss studies where optogenetic approaches have enabled high-resolution analysis of properties such as relative cell positioning, subcellular localization, morphology, and gene expression dynamics. In addition, we highlight emerging feedback and event-driven control methods that dynamically modulate cellular states using light signals. By leveraging light's unique capabilities for spatial and temporal manipulation, researchers can now probe cellular characteristics with unprecedented precision. We anticipate significant advances as researchers introduce more sophisticated dynamically patterned light signals for single-cell microbial research.

Abstract: Molecular biology has benefited enormously from repurposed tools-many enzymes and antibodies evolved for other functions but are now essential for interrogating biological function by manipulating proteins or nucleic acids. In contrast, lipids have remained technically difficult to visualize or manipulate in cells. This review introduces tools that bring lipid biology into reach for molecular cell biologists, using familiar experimental approaches. We first describe adaptations of immunofluorescence and live-cell imaging of fluorescent molecules to track lipids. Then, we discuss tools for manipulating lipid levels, including pharmacologic inhibitors, synthetic biology platforms for inducible lipid generation or degradation, and optogenetic systems for precise temporal control. While some methods remain technically demanding, most tools are now broadly accessible. Our goal is to offer a practical framework for integrating lipid biology into mainstream cell biology experiments.

Abstract: Precise regulation of protein abundance is essential for understanding dynamic cellular processes and for advancing therapeutic development. However, existing approaches lack the spatiotemporal resolution required to these cellular processes. Recent advances in optogenetics have enabled the design of optogenetic targeted protein degradation systems (Opto-TPD) allowing reversible and non-invasive control of protein stability with high spatiotemporal precision. In this review, we systematically summarize the design principles of Opto-TPD tools, including those based on light-oxygen-voltage (LOV)-domain conformational systems, light-inducible dimerization systems, and light-controlled degradation tool expression systems. We further highlight their applications in probing protein function, modulating signaling pathways, and therapeutic translations. By comparing the mechanistic features, performance, and limitations of each platform, we aim to provide a comprehensive resource for guiding future tool optimization. Altogether, these Opto-TPD tools represent a powerful and versatile complement to existing protein manipulation technologies, expanding the toolbox for precise control of protein homeostasis in living systems.

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: 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: 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 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: 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: 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: Recent advances in protein structure prediction by artificial intelligence have enabled the rational design of engineered enzymes with enhanced activity and precise regulatory features. Here, we report the AlphaFold3-guided enhancement of MagMboI, a photoactivatable restriction enzyme designed for light-controlled top-down genome engineering. MagMboI is derived from the type II restriction enzyme MboI and functions through a split-protein strategy in which its N- and C-terminal fragments are fused to light-inducible dimerization modules. Upon exposure to blue light, these domains heterodimerize, restoring nuclease activity in a controlled manner. Using AlphaFold3, we modeled the structure of the MagMboI-DNA complex and gained structural insights into the interaction between MagMboI and its target DNA recognition sequence (5'-GATC-3') required for Mg2+-dependent DNA cleavage. Comparing neighboring split-site variants, we identified an alternative split that increases the MagMboI-DNA interface area and enhances complex stability relative to the original construct. This redesigned variant (designated MagMboI-plus) preserves α-helical integrity while strengthening protein-DNA contacts. Although MagMboI-plus, when introduced in Saccharomyces cerevisiae cells, exhibited slightly increased DNA-cleavage activity in vivo upon blue light activation, it was found to induce more pronounced genomic rearrangements compared to the original MagMboI construct. These findings demonstrate that AlphaFold3-based prediction can accelerate functional improvements in engineered enzymes, providing a strategy for developing light-controlled genome engineering tools.

Abstract: Optogenetically regulated enzymes offer unprecedented spatiotemporal control over protein activity, intermolecular interactions, and intracellular signaling. Many design strategies have been developed for their fabrication based on the principles of intrinsic allostery, oligomerization or 'split' status, intracellular compartmentalization, and steric hindrance. In addition to employing photosensory domains as part of the traditional optogenetic toolset, the specificity of effector domains has also been leveraged for endogenous applications. Here, we discuss the dynamics of light activation while providing a bird's eye view of the crafting approaches, targets, and impact of optogenetic enzymes in orchestrating cellular functions, as well as the bottlenecks and an outlook into the future.

Abstract: Targeted protein degradation is a powerful tool for biological research, cell therapy, and synthetic biology. However, conventional methods often depend on pre-fused degrons or chemical degraders, limiting their wider applications. Here we develop a guided protein labeling and degradation system (GPlad) in Escherichia coli, using de novo designed guide proteins and arginine kinase (McsB) for precise degradation of various proteins, including fluorescent proteins, metabolic enzymes, and human proteins. We expand GPlad into versatile tools such as antiGPlad, OptoGPlad, and GPTAC, enabling reversible inhibition, optogenetic regulation, and biological chimerization. The combination of GPlad and antiGPlad allows for programmable circuit construction, including ON/OFF switches, signal amplifiers, and oscillators. OptoGPlad-mediated degradation of MutH accelerates E. coli evolution under protocatechuic acid stress, reducing the required generations from 220 to 100. GPTAC-mediated degradation of AroE enhanced the titer of 3-dehydroshikimic acid to 92.6 g/L, a 23.8% improvement over the conventional CRISPR interference method. We provide a tunable, plug-and-play strategy for straightforward protein degradation without the need for pre-fusion, with substantial implications for synthetic biology and metabolic engineering.

Abstract: Clustered regularly interspaced short palindromic repeats (CRISPR) technology represents a landmark advance in the field of gene editing. However, conventional CRISPR/Cas systems are limited by inadequate temporal and spatial control. In recent years, the development of optically controlled CRISPR (Opto-CRISPR) technology has offered a novel solution to this issue. As a combination of optogenetics and the CRISPR technology, the Opto-CRISPR technology enables dynamic space-time-specific gene editing and regulation in cells and organisms. In this review, we concisely introduce the basic principles of Opto-CRISPR, summarize its operational mechanisms, and discuss its applications and recent advances across various research fields. In addition, this review analyzes the limitations of Opto-CRISPR, aiming to provide a reference for the development of this emerging field.