Curated Optogenetic Publication Database

Abstract: The actin cytoskeleton and its nanoscale organization are central to all eukaryotic cells-powering diverse cellular functions including morphology, motility, and cell division-and is dysregulated in multiple diseases. Historically studied largely with purified proteins or in isolated cells, tools to study cell type-specific roles of actin in multicellular contexts are greatly needed. DeActs are recently created, first-in-class genetic tools for perturbing actin nanostructures and dynamics in specific cell types across diverse eukaryotic model organisms. Here, ChiActs are introduced, the next generation of actin-perturbing genetic tools that can be rapidly activated in cells and optogenetically targeted to distinct subcellular locations using light. ChiActs are composed of split halves of DeAct-SpvB, whose potent actin disassembly-promoting activity is restored by chemical-induced dimerization or allosteric switching. It is shown that ChiActs function to rapidly induce actin disassembly in several model cell types and are able to perturb actin-dependent nano-assembly and cellular functions, including inhibiting lamellipodial protrusions and membrane ruffling, remodeling mitochondrial morphology, and reorganizing chromatin by locally constraining actin disassembly to specific subcellular compartments. ChiActs thus expand the toolbox of genetically-encoded tools for perturbing actin in living cells, unlocking studies of the many roles of actin nano-assembly and dynamics in complex multicellular systems.

Abstract: The cGAS-STING signaling pathway has emerged as a promising target for immunotherapy development. Here, we introduce a light-sensitive optogenetic device for control of the cGAS/STING signaling to conditionally modulate innate immunity, called 'light-inducible SMOC-like repeats' (LiSmore). We demonstrate that photo-activated LiSmore boosts dendritic cell (DC) maturation and antigen presentation with high spatiotemporal precision. This non-invasive approach photo-sensitizes cytotoxic T lymphocytes to engage tumor antigens, leading to a sustained antitumor immune response. When combined with an immune checkpoint blocker (ICB), LiSmore improves antitumor efficacy in an immunosuppressive lung cancer model that is otherwise unresponsive to conventional ICB treatment. Additionally, LiSmore exhibits an abscopal effect by effectively suppressing tumor growth in a distal site in a bilateral mouse model of melanoma. Collectively, our findings establish the potential of targeted optogenetic activation of the STING signaling pathway for remote immunomodulation in mice.

Abstract: Biomolecular condensates participate in the regulation of gene transcription, yet the relationship between nuclear condensation and transcriptional activation remains elusive. Here, we devised a biotinylated CRISPR-dCas9-based optogenetic method, light-activated macromolecular phase separation (LAMPS), to enable inducible formation, affinity purification, and multiomic dissection of nuclear condensates at the targeted genomic loci. LAMPS-induced condensation at enhancers and promoters activates endogenous gene transcription by chromatin reconfiguration, causing increased chromatin accessibility and de novo formation of long-range chromosomal loops. Proteomic profiling of light-induced condensates by dCas9-mediated affinity purification uncovers multivalent interaction-dependent remodeling of macromolecular composition, resulting in the selective enrichment of transcriptional coactivators and chromatin structure proteins. Our findings support a model whereby the formation of nuclear condensates at native genomic loci reconfigures chromatin architecture and multiprotein assemblies to modulate gene transcription. Hence, LAMPS facilitates mechanistic interrogation of the relationship between nuclear condensation, genome structure, and gene transcription in living cells.

Abstract: Membrane contact sites (MCSs) mediate crucial physiological processes in eukaryotic cells, including ion signaling, lipid metabolism, and autophagy. Dysregulation of MCSs is closely related to various diseases, such as type 2 diabetes mellitus (T2DM), neurodegenerative diseases, and cancers. Visualization, proteomic mapping and manipulation of MCSs may help the dissection of the physiology and pathology MCSs. Recent technical advances have enabled better understanding of the dynamics and functions of MCSs. Here we present a summary of currently known functions of MCSs, with a focus on optical approaches to visualize and manipulate MCSs, as well as proteomic mapping within MCSs.

Abstract: Optogenetics combines optics and genetics to enable non-invasive interrogation of cell physiology at an unprecedented high spatiotemporal resolution. Here, we introduce Opto-CRAC as a set of genetically-encoded calcium actuators (GECAs) engineered from the calcium release-activated calcium (CRAC) channel, which has been tailored for optical control of calcium entry and calcium-dependent physiological responses in non-excitable cells and tissues. We describe a detailed protocol for applying Opto-CRAC as an optogenetic tool to achieve photo-tunable control over intracellular calcium signals and calcium-dependent gene expression in mammalian cells.

Abstract: Genetically encoded photoswitches have enabled spatial and temporal control of cellular events to achieve tailored functions in living cells, but their applications to probe the structure-function relations of signaling proteins are still underexplored. We illustrate herein the incorporation of various blue light-responsive photoreceptors into modular domains of the stromal interaction molecule 1 (STIM1) to manipulate protein activity and faithfully recapitulate STIM1-mediated signaling events. Capitalizing on these optogenetic tools, we identify the molecular determinants required to mediate protein oligomerization, intramolecular conformational switch, and protein-target interactions. In parallel, we have applied these synthetic devices to enable light-inducible gating of calcium channels, conformational switch, dynamic protein-microtubule interactions and assembly of membrane contact sites in a reversible manner. Our optogenetic engineering approach can be broadly applied to aid the mechanistic dissection of cell signaling, as well as non-invasive interrogation of physiological processes with high precision.