Department of Chemistry, School of Science, The University of Tokyo, Tokyo, Japan.
Photochem Photobiol. 2018 Sep;94(5):1071-1076. doi: 10.1111/php.12955. Epub 2018 Jul 12.
Transcriptional regulation is a useful strategy for gene therapy and for biomedical research. Unlike chemically regulated transcriptional approaches, spatiotemporal control of transcription using optogenetic tools is a powerful technology for the analysis of single cells. For light to penetrate into tissues, it is desired to use photoreceptors absorbing red/far-red light with a low-molecular mass applicable for the use of virus vectors, and a photoswitch using the photoreceptor needs to be constructed as a single expression vector. Herein, we describe an optogenetic tool based on Arabidopsis thaliana phytochrome (Phy) B and its binding partner, phytochrome-interacting factor (PIF) 6. We generated a truncated PhyB, which allowed for reversible association with PIF6 by red/far-red light illumination. The red light illumination only for 5 min induced PhyB translocation from the cytoplasm into the nucleus by the association with PIF6, resulting in transcriptional activation based on Gal4 DNA-binding domain and the upstream activating sequence of Gal system. The nucleocytoplasmic shuttling vector using PhyB and PIF6 might be applicable for transcriptional regulation in tissue experiments.
转录调控是基因治疗和生物医学研究的一种有效策略。与化学调控转录方法不同，使用光遗传学工具对转录进行时空控制是分析单细胞的强大技术。为了使光穿透组织，需要使用吸收红光/远红光的低分子量光受体，适用于病毒载体的使用，并且需要构建使用该光受体的光开关作为单个表达载体。本文中，我们描述了一种基于拟南芥光敏色素（Phy）B及其结合伴侣光敏色素相互作用因子（PIF）6 的光遗传学工具。我们生成了一个截断的 PhyB，它允许通过红光/远红光照射与 PIF6 可逆结合。红光仅照射 5 分钟，通过与 PIF6 的结合将 PhyB 从细胞质转运到细胞核中，从而基于 Gal4 DNA 结合域和 Gal 系统的上游激活序列激活转录。使用 PhyB 和 PIF6 的核质穿梭载体可能适用于组织实验中的转录调控。
