Curated Optogenetic Publication Database

Abstract: Optogenetic tools have revolutionized the control of gene expression with high spatial and temporal resolution. Here we present a Single-chain Light-Activatable Transcriptional Reporter (SLATR), a system capable of fluorescently tagging target cells with minutes of white light stimulation. In its inactive, or dark state, a transcriptional factor is cytosolically bound, preventing nuclear translocation. White light irradiation triggers its release through the protease cleavage of a site that is sterically caged by the circularly permuted Avena sativa LOV2 (cpAsLOV2) domain. We discovered that cpAsLOV2 cages the cleavage site more efficiently than AsLOV2, achieving low background in the SLATR design. We demonstrate that SLATR exhibits a signal-to-background ratio between 3.4 and 36 and achieves reporter activation within 60 min of light stimulation. Furthermore, SLATR outperforms the only other single-chain light-activatable transcriptional reporter, LAUNCHER, with faster kinetics, greater light sensitivity, and markedly lower background under identical stimulation conditions. Our single-chain light-activatable transcriptional system expands the optogenetic toolkit though providing a simpler system for regulating gene expression with precise spatiotemporal control.

Abstract: To address the need for methods for tagging and manipulating neuronal ensembles underlying specific behaviors, we present an improved version of FLARE, termed cytoFLARE (cytosol-expressed FLARE). cytoFLARE incorporates cytosolic tethering of a transcription factor and expression of a more sensitive pair of calcium-sensing domains. We show that cytoFLARE captures more calcium- and light-dependent signals in HEK293T cells and higher signal-to-background ratios in neuronal cultures. We further establish cytoFLARE transgenic Drosophila models and apply cytoFLARE to label activated neurons upon sensory or optogenetic stimulation within a defined time window. Notably, through the cytoFLARE-driven expression of optogenetic actuators, we successfully reactivated and inhibited neurons involved in the larval nociceptive system. Our findings demonstrate the characterization and application of time-gated calcium integrators for both recording and manipulating neuronal activity in Drosophila larvae.