Curated Optogenetic Publication Database

Abstract: Engineered bacteria-based cancer therapy has increasingly been considered to be a promising therapeutic strategy due to the development of synthetic biology. Wherein, engineering bacteria-mediated photodynamic therapy (PDT)-immunotherapy shows greater advantages and potential in treatment efficiency than monotherapy. However, the unsustainable regeneration of photosensitizers (PSs) and weak immune responses limit the therapeutic efficiency. Herein, we developed an engineered bacteria-based delivery system for sequential delivery of PSs and checkpoint inhibitors in cancer PDT-immunotherapy. The biosynthetic pathway of 5-aminolevulinic acid (5-ALA) was introduced into Escherichia coli, yielding a supernatant concentration of 172.19 mg/L after 10 h of growth. And another strain was endowed with the light-controllable releasement of anti-programmed cell death-ligand 1 nanobodies (anti-PD-L1). This system exhibited a collaborative effect, where PDT initiated tumor cell death and the released tumor cell fragments stimulated immunity, followed by the elimination of residual tumor cells. The tumor inhibition rate reached 74.97%, and the portion of activated T cells and inflammatory cytokines were reinforced. The results demonstrated that the engineered bacteria-based collaborative system could sequentially deliver therapeutic substance and checkpoint inhibitors, and achieve good therapeutic therapy. This paper will provide a new perspective for the cancer PDT-immunotherapy.

Abstract: Vital organismal processes, including development, differentiation and adaptation, involve altered gene expression. Although expression is frequently controlled at the transcriptional stage, various regulation mechanisms operate at downstream levels. Here, we leverage the photoreceptor NmPAL to optogenetically induce RNA refolding and the translation of bacterial mRNAs. Blue-light-triggered NmPAL binding disrupts a cis-repressed mRNA state, thereby relieves obstruction of translation initiation, and upregulates gene expression. Iterative probing and optimization of the circuit, dubbed riboptoregulator, enhanced induction to 30-fold. Given action at the mRNA level, the riboptoregulator can differentially regulate individual structural genes within polycistronic operons. Moreover, it is orthogonal to and can be wed with other gene-regulatory circuits for nuanced and more stringent gene-expression control. We thus advance the pAurora2 circuit that combines transcriptional and translational mechanisms to optogenetically increase bacterial gene expression by >1000-fold. The riboptoregulator strategy stands to upgrade numerous regulatory circuits and widely applies to expression control in microbial biotechnology, synthetic biology and materials science.

Abstract: Optogenetics' advancement has made light induction attractive for controlling biological processes due to its advantages of fine-tunability, reversibility, and low toxicity. The lactose operon induction system, commonly used in Escherichia coli, relies on the binding of lactose or isopropyl β-d-1-thiogalactopyranoside (IPTG) to the lactose repressor protein LacI, playing a pivotal role in controlling the lactose operon. Here, we harnessed the light-responsive light-oxygen-voltage 2 (LOV2) domain from Avena sativa phototropin 1 as a tool for light control and engineered LacI into two light-responsive variants, OptoLacIL and OptoLacID. These variants exhibit direct responsiveness to light and darkness, respectively, eliminating the need for IPTG. Building upon OptoLacI, we constructed two light-controlled E. coli gene expression systems, OptoE.coliLight system and OptoE.coliDark system. These systems enable bifunctional gene expression regulation in E. coli through light manipulation and show superior controllability compared to IPTG-induced systems. We applied the OptoE.coliDark system to protein production and metabolic flux control. Protein production levels are comparable to those induced by IPTG. Notably, the titers of dark-induced production of 1,3-propanediol (1,3-PDO) and ergothioneine exceeded 110% and 60% of those induced by IPTG, respectively. The development of OptoLacI will contribute to the advancement of the field of optogenetic protein engineering, holding substantial potential applications across various fields.

Abstract: Bacteria must constantly probe their environment for rapid adaptation, a crucial need most frequently served by two-component systems (TCS). As one component, sensor histidine kinases (SHK) control the phosphorylation of the second component, the response regulator (RR). Downstream responses hinge on RR phosphorylation and can be highly stringent, acute, and sensitive because SHKs commonly exert both kinase and phosphatase activity. With a bacteriophytochrome TCS as a paradigm, we here interrogate how this catalytic duality underlies signal responses. Derivative systems exhibit tenfold higher red-light sensitivity, owing to an altered kinase-phosphatase balance. Modifications of the linker intervening the SHK sensor and catalytic entities likewise tilt this balance and provide TCSs with inverted output that increases under red light. These TCSs expand synthetic biology and showcase how deliberate perturbations of the kinase-phosphatase duality unlock altered signal-response regimes. Arguably, these aspects equally pertain to the engineering and the natural evolution of TCSs.

Abstract: Optogenetic tools have been used in a wide range of microbial engineering applications that benefit from the tunable, spatiotemporal control that light affords. However, the majority of current optogenetic constructs for bacteria respond to blue light, limiting the potential for multichromatic control. In addition, other wavelengths offer potential benefits over blue light, including improved penetration of dense cultures and reduced potential for toxicity. In this study, we introduce OptoCre-REDMAP, a red light inducible Cre recombinase system in Escherichia coli. This system harnesses the plant photoreceptors PhyA and FHY1 and a split version of Cre recombinase to achieve precise control over gene expression and DNA excision in bacteria. We optimized the design by modifying the start codon of Cre and characterized the impact of different levels of induction to find conditions that produced minimal basal expression in the dark and full activation within four hours of red light exposure. We characterized the system’s sensitivity to ambient light, red light intensity, and exposure time, finding OptoCre-REDMAP to be reliable and flexible across a range of conditions. The system exhibits robust light-sensitive behavior, responding to red light while remaining inactive under blue light, making it suitable for future applications in synthetic biology that require multichromatic control.

Abstract: The ideal engineered microbial smart-drug should be capable of functioning on demand at specific sites in vivo. However, precise regulation of engineered microorganisms poses challenges in the convoluted and elongated intestines. Despite the promising application potential of optogenetic regulation strategies based on light signals, the poor tissue penetration of light signals limits their application in large experimental animals. Given the rapid development of ingestible electronic capsules in recent years, taking advantage of them as regulatory devices to deliver light signals in situ to engineered bacteria within the intestines has become feasible. In this study, we established an electronic-microorganism signaling system, realized by two Bluetooth-controlled luminescent electronic capsules were designed. The “Manager” capsule is equipped with a photosensor to monitor the distribution of engineered bacteria and to activate the optogenetic function of the bacteria by emitting green light. The other capsule, “Locator”, can control the in situ photopolymerization of hydrogels in the intestines via ultraviolet light, aiding in the retention of engineered bacteria at specific sites. These two electronic capsules are expected to work synergistically to regulate the distribution and function of engineered bacteria in vivo, and their application in the treatment of colitis in pigs is currently being investigated, with relevant results to be updated subsequently.

Abstract: Optogenetics is a powerful tool for spatiotemporal control of gene expression. Several light-inducible gene regulators have been developed to function in bacteria, and these regulatory circuits have been ported to new host strains. Here, we developed and adapted a red-light-inducible transcription factor for Shewanella oneidensis. This regulatory circuit is based on the iLight optogenetic system, which controls gene expression using red light. A thermodynamic model and promoter engineering were used to adapt this system to achieve differential gene expression in light and dark conditions within a S. oneidensis host strain. We further improved the iLight optogenetic system by adding a repressor to invert the genetic circuit and activate gene expression under red light illumination. The inverted iLight genetic circuit was used to control extracellular electron transfer within S. oneidensis. The ability to use both red- and blue-light-induced optogenetic circuits simultaneously was also demonstrated. Our work expands the synthetic biology capabilities in S. oneidensis, which could facilitate future advances in applications with electrogenic bacteria.

Abstract: The discovery of antibiotics has noticeably promoted the development of human civilization; however, antibiotic resistance in bacteria caused by abusing and overusing greatly challenges human health and food safety. Considering the worsening situation, it is an urgent demand to develop emerging nontraditional technologies or methods to address this issue. With the expanding of synthetic biology, optogenetics exhibits a tempting prospect for precisely regulating gene expression in many fields. Consequently, it is attractive to employ optogenetics to reduce the risk of antibiotic resistance. Here, a blue light-controllable gene expression system was established in Escherichia coli based on a photosensitive DNA-binding protein (EL222). Further, this strategy was successfully applied to repress the expression of β-lactamase gene (bla) using blue light illumination, resulting a dramatic reduction of ampicillin resistance in engineered E. coli. Moreover, blue light was utilized to induce the expression of the mechanosensitive channel of large conductance (MscL), triumphantly leading to the increase of streptomycin susceptibility in engineered E. coli. Finally, the increased susceptibility of ampicillin and streptomycin was simultaneously induced by blue light in the same E. coli cell, revealing the excellent potential of this strategy in controlling multidrug-resistant (MDR) bacteria. As a proof of concept, our work demonstrates that light can be used as an alternative tool to prolong the use period of common antibiotics without developing new antibiotics. And this novel strategy based on optogenetics shows a promising foreground to combat antibiotic resistance in the future.

Abstract: The transition of bacteria from an individualistic to a biofilm lifestyle profoundly alters their biology. During biofilm development, the bacterial cell-cell adhesions are a major determinant of initial microcolonies, which serve as kernels for the subsequent microscopic and mesoscopic structure of the biofilm, and determine the resulting functionality. In this study, the significance of bacterial cell-cell adhesion dynamics on bacterial aggregation and biofilm maturation is elucidated. Using photoswitchable adhesins between bacteria, modifying the dynamics of bacterial cell-cell adhesions with periodic dark-light cycles is systematic. Dynamic cell-cell adhesions with liquid-like behavior improve bacterial aggregation and produce more compact microcolonies than static adhesions with solid-like behavior in both experiments and individual-based simulations. Consequently, dynamic cell-cell adhesions give rise to earlier quorum sensing activation, better intermixing of different bacterial populations, improved biofilm maturation, changes in the growth of cocultures, and higher yields in fermentation. The here presented approach of tuning bacterial cell-cell adhesion dynamics opens the door for regulating the structure and function of biofilms and cocultures with potential biotechnological applications.

Abstract: Biomolecular condensates are dynamic subcellular compartments that lack surrounding membranes and can spatiotemporally organize the cellular biochemistry of eukaryotic cells. However, such dynamic organization has not been realized in prokaryotes that naturally lack organelles, and strategies are urgently needed for dynamic biomolecular compartmentalization. Here we develop a light-switchable condensate system for on-demand dynamic organization of functional cargoes in the model prokaryotic Escherichia coli cells. The condensate system consists of two modularly designed and genetically encoded fusions that contain a condensation-enabling scaffold and a functional cargo fused to the blue light-responsive heterodimerization pair, iLID and SspB, respectively. By appropriately controlling the biogenesis of the protein fusions, the condensate system allows rapid recruitment and release of cargo proteins within seconds in response to light, and this process is also reversible and repeatable. Finally, the system is demonstrated to dynamically control the subcellular localization of a cell division inhibitor, SulA, which enables the reversible regulation of cell morphologies. Therefore, this study provides a new strategy to dynamically control cellular processes by harnessing light-controlled condensates in prokaryotic cells.

Abstract: Epstein-Barr virus (EBV) is detected in 40% of patients with Hodgkin lymphoma (HL). During latency, EBV induces epigenetic alterations to the host genome and decreases the expression of pro-apoptotic proteins. The present study aimed to evaluate the expression levels of mRNA molecules and the end product of proteins for the JAK/STAT and NF-κB pathways, and their association with clinicopathological and prognostic parameters in patients with EBV-positive and -negative classical Hodgkin lymphoma (CHL).

Abstract: The regulation of gene expression by light enables the versatile, spatiotemporal manipulation of biological function in bacterial and mammalian cells. Optoribogenetics extends this principle by molecular RNA devices acting on the RNA level whose functions are controlled by the photoinduced interaction of a light-oxygen-voltage photoreceptor with cognate RNA aptamers. Here light-responsive ribozymes, denoted optozymes, which undergo light-dependent self-cleavage and thereby control gene expression are described. This approach transcends existing aptamer-ribozyme chimera strategies that predominantly rely on aptamers binding to small molecules. The optozyme method thus stands to enable the graded, non-invasive, and spatiotemporally resolved control of gene expression. Optozymes are found efficient in bacteria and mammalian cells and usher in hitherto inaccessible optoribogenetic modalities with broad applicability in synthetic and systems biology.

Abstract: Molecular tools for optogenetic control allow for spatial and temporal regulation of cell behavior. In particular, light controlled protein degradation is a valuable mechanism of regulation because it can be highly modular, used in tandem with other control mechanisms, and maintain functionality throughout growth phases. Here, we engineered LOVdeg, a tag that can be appended to a protein of interest for inducible degradation in Escherichia coli using blue light. We demonstrate the modularity of LOVdeg by using it to tag a range of proteins, including the LacI repressor, CRISPRa activator, and the AcrB efflux pump. Additionally, we demonstrate the utility of pairing the LOVdeg tag with existing optogenetic tools to enhance performance by developing a combined EL222 and LOVdeg system. Finally, we use the LOVdeg tag in a metabolic engineering application to demonstrate post-translational control of metabolism. Together, our results highlight the modularity and functionality of the LOVdeg tag system, and introduce a powerful new tool for bacterial optogenetics.

Abstract: By applying sensory photoreceptors, optogenetics realizes the light-dependent control of cellular events and state. Given reversibility, noninvasiveness, and exquisite spatiotemporal precision, optogenetic approaches enable innovative use cases in cell biology, synthetic biology, and biotechnology. In this chapter, we detail the implementation of the pREDusk, pREDawn, pCrepusculo, and pAurora optogenetic circuits for controlling bacterial gene expression by red and blue light, respectively. The protocols provided here guide the practical use and multiplexing of these circuits, thereby enabling graded protein production in bacteria at analytical and semi-preparative scales.

Abstract: Optogenetics is an attractive synthetic biology tool for controlling the metabolic flux distribution. Here, we demonstrated optogenetic flux ratio control of glycolytic pathways consisting of the Embden-Meyerhof-Parnas (EMP), pentose phosphate (PP), and Entner-Doudoroff (ED) pathways by illuminating multicolor lights using blue light-responsive EL222 and green/red light-responsive CcaSR in Escherichia coli. EL222 forms a dimer and binds to a particular DNA sequence under blue light; therefore, target gene expression can be reduced or induced by inserting a recognition sequence into its promoter regions. First, a flux ratio between the PP and ED pathways was controlled by blue light using EL222. After blocking the EMP pathway, the EL222-recognition sequence was inserted between the -35 and -10 regions of gnd to repress the PP flux and was also inserted upstream of the -35 region of edd to induce ED flux. After adjusting light intensity, the PP:ED flux ratios were 60:39% and 29:70% under dark and blue light conditions, respectively. Finally, a CcaSR-based pgi expression system was implemented to control the flux ratio between the EMP and PP + ED pathways by illuminating green/red light. The EMP:PP:ED flux ratios were 80:9:11%, 14:35:51%, and 33:5:62% under green, red, and red and blue light, respectively.

Abstract: Microbial biosynthesis, as an alternative method for producing quantum dots (QDs), has gained attention because it can be conducted under mild and environmentally friendly conditions, distinguishing it from conventional chemical and physical synthesis approaches. However, there is currently no method to selectively control this biosynthesis process in a subset of microbes within a population using external stimuli. In this study, we have attained precise and selective control over the microbial biosynthesis of QDs through the utilization of an optogenetically engineered Escherichia coli (E. coli). The recombinant E. coli is designed to express smCSE enzyme, under the regulation of eLightOn system, which can be activated by blue light. The smCSE enzymes use L-cysteine and Cd2+ as substrates to form CdS QDs. This system enables light-inducible bacterial biosynthesis of QDs in precise patterns within a hydrogel for information encryption. As the biosynthesis progresses, the optical characteristics of the QDs change, allowing living materials containing the recombinant E. coli to display time-dependent patterns that self-destruct after reading. Compared to static encryption using fluorescent QD inks, dynamic information encryption based on living materials offers enhanced security.

Abstract: Anti-CRISPR (Acr) proteins are encoded by mobile genetic elements to overcome the CRISPR immunity of prokaryotes, displaying promises as controllable tools for modulating CRISPR-based applications. However, characterizing novel anti-CRISPR proteins and exploiting Acr-related technologies is a rather long and tedious process. Here, we established a versatile plasmid interference with CRISPR interference (PICI) system in Escherichia coli for rapidly characterizing Acrs and developing Acr-based technologies. Utilizing the PICI system, we discovered two novel type II-A Acrs (AcrIIA33 and AcrIIA34), which can inhibit the activity of SpyCas9 by affecting DNA recognition of Cas9. We further constructed a circularly permuted AcrIIA4 (cpA4) protein and developed optogenetically engineered, robust AcrIIA4 (OPERA4) variants by combining cpA4 with the light-oxygen-voltage 2 (LOV2) blue light sensory domain. OPERA4 variants are robust light-dependent tools for controlling the activity of SpyCas9 by approximately 1000-fold change under switching dark-light conditions in prokaryotes. OPERA4 variants can achieve potent light-controllable genome editing in human cells as well. Together, our work provides a versatile screening system for characterizing Acrs and developing the Acr-based controllable tools.

Abstract: Optogenetics is a powerful tool for spatiotemporal control of gene expression. Several light-inducible gene regulators have been developed to function in bacteria, and these regulatory circuits have been ported into new host strains. Here, we developed and adapted a red light-inducible transcription factor for Shewanella oneidensis. This regulatory circuit is based on the iLight optogenetic system, which controls gene expression using red light. Promoter engineering and a thermodynamic model were used to adapt this system to achieve differential gene expression in light and dark conditions within a S. oneidensis host strain. We further improved the iLight optogenetic system by adding a repressor to invert the genetic circuit and activate gene expression under red light illumination. The inverted iLight genetic circuit was used to control extracellular electron transfer (EET) within S. oneidensis. The ability to use both red and blue light-induced optogenetic circuits simultaneously was demonstrated. Our work expands the synthetic biology toolbox of Shewanella, which could facilitate future advances in applications with electrogenic bacteria.

Abstract: Base editors, which enable targeted locus nucleotide conversion in genomic DNA without double-stranded breaks, have been engineered as powerful tools for biotechnological and clinical applications. However, the application of base editors is limited by their off-target effects. Continuously expressed deaminases used for gene editing may lead to unwanted base alterations at unpredictable genomic locations. In the present study, blue-light-activated base editors (BLBEs) are engineered based on the distinct photoswitches magnets that can switch from a monomer to dimerization state in response to blue light. By fusing the N- and C-termini of split DNA deaminases with photoswitches Magnets, efficient A-to-G and C-to-T base editing is achieved in response to blue light in prokaryotic and eukaryotic cells. Furthermore, the results showed that BLBEs can realize precise blue light-induced gene editing across broad genomic loci with low off-target activity at the DNA- and RNA-level. Collectively, these findings suggest that the optogenetic utilization of base editing and optical base editors may provide powerful tools to promote the development of optogenetic genome engineering.

Abstract: The ability to perform sophisticated, high-throughput optogenetic experiments has been greatly enhanced by recent open-source illumination devices that allow independent programming of light patterns in single wells of microwell plates. However, there is currently a lack of instrumentation to monitor such experiments in real time, necessitating repeated transfers of the samples to stand-alone analytical instruments, thus limiting the types of experiments that could be performed. Here we address this gap with the development of the optoPlateReader (oPR), an open-source, solid-state, compact device that allows automated optogenetic stimulation and spectroscopy in each well of a 96-well plate. The oPR integrates an optoPlate illumination module with a module called the optoReader, an array of 96 photodiodes and LEDs that allows 96 parallel light measurements. The oPR was optimized for stimulation with blue light and for measurements of optical density and fluorescence. After calibration of all device components, we used the oPR to measure growth and to induce and measure fluorescent protein expression in E. coli. We further demonstrated how the optical read/write capabilities of the oPR permit computer-in-the-loop feedback control, where the current state of the sample can be used to adjust the optical stimulation parameters of the sample according to pre-defined feedback algorithms. The oPR will thus help realize an untapped potential for optogenetic experiments by enabling automated reading, writing, and feedback in microwell plates through open-source hardware that is accessible, customizable, and inexpensive.

Abstract: Biotechnology offers many opportunities for the sustainable manufacturing of valuable products. The toolbox to optimize bioprocesses includes extracellular process elements such as the bioreactor design and mode of operation, medium formulation, culture conditions, feeding rates, and so on. However, these elements are frequently insufficient for achieving optimal process performance or precise product composition. One can use metabolic and genetic engineering methods for optimization at the intracellular level. Nevertheless, those are often of static nature, failing when applied to dynamic processes or if disturbances occur. Furthermore, many bioprocesses are optimized empirically and implemented with little-to-no feedback control to counteract disturbances. The concept of cybergenetics has opened new possibilities to optimize bioprocesses by enabling online modulation of the gene expression of metabolism-relevant proteins via external inputs (e.g., light intensity in optogenetics). Here, we fuse cybergenetics with model-based optimization and predictive control for optimizing dynamic bioprocesses. To do so, we propose to use dynamic constraint-based models that integrate the dynamics of metabolic reactions, resource allocation, and inducible gene expression. We formulate a model-based optimal control problem to find the optimal process inputs. Furthermore, we propose using model predictive control to address uncertainties via online feedback. We focus on fed-batch processes, where the substrate feeding rate is an additional optimization variable. As a simulation example, we show the optogenetic control of the ATPase enzyme complex for dynamic modulation of enforced ATP wasting to adjust product yield and productivity.

Abstract: Synthetic genetic oscillators can serve as internal clocks within engineered cells to program periodic expression. However, cell-to-cell variability introduces a dispersion in the characteristics of these clocks that drives the population to complete desynchronization. Here we introduce the optorepressilator, an optically controllable genetic clock that combines the repressilator, a three-node synthetic network in E. coli, with an optogenetic module enabling to reset, delay, or advance its phase using optical inputs. We demonstrate that a population of optorepressilators can be synchronized by transient green light exposure or entrained to oscillate indefinitely by a train of short pulses, through a mechanism reminiscent of natural circadian clocks. Furthermore, we investigate the system’s response to detuned external stimuli observing multiple regimes of global synchronization. Integrating experiments and mathematical modeling, we show that the entrainment mechanism is robust and can be understood quantitatively from single cell to population level.

Abstract: Splitting proteins with light- or chemically inducible dimers provides a mechanism for post-translational control of protein function. However, current methods for engineering stimulus-responsive split proteins often require significant protein engineering expertise and the laborious screening of individual constructs. To address this challenge, we use a pooled library approach that enables rapid generation and screening of nearly all possible split protein constructs in parallel, where results can be read out by using sequencing. We perform our method on Cre recombinase with optogenetic dimers as a proof of concept, resulting in comprehensive data on the split sites throughout the protein. To improve the accuracy in predicting split protein behavior, we develop a Bayesian computational approach to contextualize errors inherent to experimental procedures. Overall, our method provides a streamlined approach for achieving inducible post-translational control of a protein of interest.

Abstract: Optogenetic actuators have revolutionized the resolution at which biological processes can be controlled. In plants, deployment of optogenetics is challenging due to the need for these light-responsive systems to function in the context of horticultural light environments. Furthermore, many available optogenetic actuators are based on plant photoreceptors that might crosstalk with endogenous signaling processes, while others depend on exogenously supplied cofactors. To overcome such challenges, we have developed Highlighter, a synthetic, light-gated gene expression system tailored for in planta function. Highlighter is based on the photoswitchable CcaS-CcaR system from cyanobacteria and is repurposed for plants as a fully genetically encoded system. Analysis of a re-engineered CcaS in Escherichia coli demonstrated green/red photoswitching with phytochromobilin, a chromophore endogenous to plants, but also revealed a blue light response likely derived from a flavin-binding LOV-like domain. We deployed Highlighter in transiently transformed Nicotiana benthamiana for optogenetic control of fluorescent protein expression. Using light to guide differential fluorescent protein expression in nuclei of neighboring cells, we demonstrate unprecedented spatiotemporal control of target gene expression. We implemented the system to demonstrate optogenetic control over plant immunity and pigment production through modulation of the spectral composition of broadband visible (white) light. Highlighter is a step forward for optogenetics in plants and a technology for high-resolution gene induction that will advance fundamental plant biology and provide new opportunities for crop improvement.

Abstract: Recent progress in protein engineering has established optogenetics as one of the leading external non-invasive stimulation strategies, with many optogenetic tools being designed for in vivo operation. Characterization and optimization of these tools require a high-throughput and versatile light delivery system targeting micro-titer culture volumes. Here, we present a universal light illumination platform – Diya, compatible with a wide range of cell culture plates and dishes. Diya hosts specially-designed features ensuring active thermal management, homogeneous illumination, and minimal light bleedthrough. It offers light induction programming via a user-friendly custom-designed GUI. Through extensive characterization experiments with multiple optogenetic tools in diverse model organisms (bacteria, yeast and human cell lines), we show that Diya maintains viable conditions for cell cultures undergoing light induction. Finally, we demonstrate an optogenetic strategy for in vivo biomolecular controller operation. With a custom-designed antithetic integral feedback circuit, we exhibit robust perfect adaptation and light-controlled set-point variation using Diya.