Optogenetics & Synthetic Biology Interdisciplinary Research Center, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Mei Long Road, Shanghai 200237, China.
Shanghai Frontiers Science Center of Optogenetic Techniques for Cell Metabolism, School of Pharmacy, East China University of Science and Technology, 130 Mei Long Road, Shanghai 200237, China.
ACS Synth Biol. 2023 Mar 17;12(3):664-671. doi: 10.1021/acssynbio.2c00410. Epub 2023 Mar 9.
The light-regulated Gal4-UAS system has offered new ways to control cellular activities with precise spatial and temporal resolution in zebrafish and Drosophila. However, the existing optogenetic Gal4-UAS systems suffer from having multiple protein components and a dependence on extraneous light-sensitive cofactors, which increase the technical complexity and limit the portability of these systems. To overcome these limitations, we herein describe the development of a novel optogenetic Gal4-UAS system (ltLightOn) for both zebrafish and Drosophila based on a single light-switchable transactivator, termed GAVPOLT, which dimerizes and binds to gene promoters to activate transgene expression upon blue light illumination. The ltLightOn system is independent of exogenous cofactors and exhibits a more than 2400-fold ON/OFF gene expression ratio, allowing quantitative, spatial, and temporal control of gene expression. We further demonstrate the usefulness of the ltLightOn system in regulating zebrafish embryonic development by controlling the expression of lefty1 by light. We believe that this single-component optogenetic system will be immensely useful in understanding the gene function and behavioral circuits in zebrafish and Drosophila.
光调控 Gal4-UAS 系统为斑马鱼和果蝇提供了新的方法，可以精确地控制细胞活动，具有精确的时空分辨率。然而，现有的光遗传学 Gal4-UAS 系统存在多个蛋白组成部分，并且依赖于外来的光敏感辅助因子，这增加了技术复杂性并限制了这些系统的可移植性。为了克服这些限制，我们在此描述了一种基于单个光可开关转录激活子 GAVPOLT 的新型光遗传学 Gal4-UAS 系统（ltLightOn）的开发，该系统可用于斑马鱼和果蝇，GAVPOLT 二聚化并结合到基因启动子上，在蓝光照射下激活转基因表达。ltLightOn 系统独立于外源辅助因子，表现出超过 2400 倍的 ON/OFF 基因表达比，允许对基因表达进行定量、空间和时间控制。我们进一步通过用光控制 lefty1 的表达来证明 ltLightOn 系统在调节斑马鱼胚胎发育中的有用性。我们相信，这种单组分光遗传学系统将在理解斑马鱼和果蝇中的基因功能和行为回路方面非常有用。
