Curated Optogenetic Publication Database

Abstract: Here, we present a gene regulation strategy enabling programmable control over eukaryotic translational initiation. By excising the natural poly-adenylation (poly-A) signal of target genes and replacing it with a synthetic control region harboring RNA-binding protein (RBP)-specific aptamers, cap-dependent translation is rendered exclusively dependent on synthetic translation initiation factors (STIFs) containing different RBPs engineered to conditionally associate with different eIF4F-binding proteins (eIFBPs). This modular design framework facilitates the engineering of various gene switches and intracellular sensors responding to many user-defined trigger signals of interest, demonstrating tightly controlled, rapid and reversible regulation of transgene expression in mammalian cells as well as compatibility with various clinically applicable delivery routes of in vivo gene therapy. Therapeutic efficacy was demonstrated in two animal models. To exemplify disease treatments that require on-demand drug secretion, we show that a custom-designed gene switch triggered by the FDA-approved drug grazoprevir can effectively control insulin expression and restore glucose homeostasis in diabetic mice. For diseases that require instantaneous sense-and-response treatment programs, we create highly specific sensors for various subcellularly (mis)localized protein markers (such as cancer-related fusion proteins) and show that translation-based protein sensors can be used either alone or in combination with other cell-state classification strategies to create therapeutic biocomputers driving self-sufficient elimination of tumor cells in mice. This design strategy demonstrates unprecedented flexibility for translational regulation and could form the basis for a novel class of programmable gene therapies in vivo.

Abstract: Diabetes treatment and rehabilitation are usually a lifetime process. Optogenetic engineered designer cell-therapy holds great promise in regulating blood glucose homeostasis. However, portable, sustainable, and long-term energy supplementation has previously presented a challenge for the use of optogenetic stimulation in vivo. Herein, we purpose a self-powered optogenetic system (SOS) for implantable blood glucose control. The SOS consists of a biocompatible far-red light (FRL) source, FRL-triggered transgene-expressing cells, a power management unit, and a flexible implantable piezoelectric nanogenerator (i-PENG) to supply long-term energy by converting biomechanical energy into electricity. Our results show that this system can harvest energy from body movement and power the FRL source, which then significantly enhanced production of a short variant of human glucagon-like peptide 1 (shGLP-1) in vitro and in vivo. Indeed, diabetic mice equipped with the SOS showed rapid restoration of blood glucose homeostasis, improved glucose, and insulin tolerance. Our results suggest that the SOS is sufficiently effective in self-powering the modulation of therapeutic outputs to control glucose homeostasis and, furthermore, present a new strategy for providing energy in optogenetic-based cell therapy.

Abstract: Pulsing cellular dynamics in genetic circuits have been shown to provide critical capabilities to cells in stress response, signaling and development. Despite the fascinating discoveries made in the past few years, the mechanisms and functional capabilities of most pulsing systems remain unclear, and one of the critical challenges is the lack of a technology that allows pulsatile regulation of transgene expression both in vitro and in vivo. Here, we describe the development of a synthetic BRET-based transgene expression (LuminON) system based on a luminescent transcription factor, termed luminGAVPO, by fusing NanoLuc luciferase to the light-switchable transcription factor GAVPO. luminGAVPO allows pulsatile and quantitative activation of transgene expression via both chemogenetic and optogenetic approaches in mammalian cells and mice. Both the pulse amplitude and duration of transgene expression are highly tunable via adjustment of the amount of furimazine. We further demonstrated LuminON-mediated blood-glucose homeostasis in type 1 diabetic mice. We believe that the BRET-based LuminON system with the pulsatile dynamics of transgene expression provides a highly sensitive tool for precise manipulation in biological systems that has strong potential for application in diverse basic biological studies and gene- and cell-based precision therapies in the future.

Abstract: It is widely understood that CRISPR-Cas9 technology is revolutionary, with well-recognized issues including the potential for off-target edits and the attendant need for spatiotemporal control of editing. Here, we describe a far-red light (FRL)–activated split-Cas9 (FAST) system that can robustly induce gene editing in both mammalian cells and mice. Through light-emitting diode–based FRL illumination, the FAST system can efficiently edit genes, including nonhomologous end joining and homology-directed repair, for multiple loci in human cells. Further, we show that FAST readily achieves FRL-induced editing of internal organs in tdTomato reporter mice. Finally, FAST was demonstrated to achieve FRL-triggered editing of the PLK1 oncogene in a mouse xenograft tumor model. Beyond extending the spectrum of light energies in optogenetic toolbox for CRISPR-Cas9 technologies, this study demonstrates how FAST system can be deployed for programmable deep tissue gene editing in both biological and biomedical contexts toward high precision and spatial specificity.

Abstract: The ability to control the activity of CRISPR-dCas9 with precise spatiotemporal resolution will enable tight genome regulation of user-defined endogenous genes for studying the dynamics of transcriptional regulation. Optogenetic devices with minimal phototoxicity and the capacity for deep tissue penetration are extremely useful for precise spatiotemporal control of cellular behavior and for future clinic translational research. Therefore, capitalizing on synthetic biology and optogenetic design principles, we engineered a far-red light (FRL)-activated CRISPR-dCas9 effector (FACE) device that induces transcription of exogenous or endogenous genes in the presence of FRL stimulation. This versatile system provides a robust and convenient method for precise spatiotemporal control of endogenous gene expression and also has been demonstrated to mediate targeted epigenetic modulation, which can be utilized to efficiently promote differentiation of induced pluripotent stem cells into functional neurons by up-regulating a single neural transcription factor, NEUROG2 This FACE system might facilitate genetic/epigenetic reprogramming in basic biological research and regenerative medicine for future biomedical applications.

Abstract: With the increasingly dominant role of smartphones in our lives, mobile health care systems integrating advanced point-of-care technologies to manage chronic diseases are gaining attention. Using a multidisciplinary design principle coupling electrical engineering, software development, and synthetic biology, we have engineered a technological infrastructure enabling the smartphone-assisted semiautomatic treatment of diabetes in mice. A custom-designed home server SmartController was programmed to process wireless signals, enabling a smartphone to regulate hormone production by optically engineered cells implanted in diabetic mice via a far-red light (FRL)-responsive optogenetic interface. To develop this wireless controller network, we designed and implanted hydrogel capsules carrying both engineered cells and wirelessly powered FRL LEDs (light-emitting diodes). In vivo production of a short variant of human glucagon-like peptide 1 (shGLP-1) or mouse insulin by the engineered cells in the hydrogel could be remotely controlled by smartphone programs or a custom-engineered Bluetooth-active glucometer in a semiautomatic, glucose-dependent manner. By combining electronic device-generated digital signals with optogenetically engineered cells, this study provides a step toward translating cell-based therapies into the clinic.