Photoactivated RNA Binding in a Blue-Light Receptor Enables Optoribogenetics

感光感受器是自然界中各种依赖光的生物适应的基础。在光遗传学中，光感受器具有可逆性、时空分辨率和最小的侵入性，同时也是可遗传编码的致动器，用于控制细胞和有机体的生理。自然产生的光感受器和过多的工程代表现在能够对多种细胞过程进行多功能的光遗传干预。尽管如此，直接作用于RNA的光遗传致动器一直很少。最近，我们从多部分放线杆菌中发现了光-氧-电压(LOV)受体PAL，从而缩小了这一差距。PAL序列-特异性地与短RNA适配子结合，在蓝光下亲和力增强约100倍，从而使迄今无法获得的光学遗传应用成为可能。目前的合作提案旨在获得PAL和最近发现的两个同源物中光调节和RNA结合的分子理解。相关的见解为PAL衍生物的工程提供了信息，使其具有改变和增强的特征，特别是RNA序列特异性和对光的响应。我们将利用光调控的PAL：适配子相互作用在细胞培养和体内两个方面对微小RNA的剪接和内源基因的表达调控进行时空控制。这项计划中的研究为新的光调节致动器铺平了道路，该致动器适用于对RNA在自然界中履行的各种角色进行精确的光学遗传询问。

Sensory photoreceptors underlie various light-dependent organismal adaptations in nature. In optogenetics, photoreceptors double as genetically encodable actuators for the control of cellular and organismal physiology with reversibility, spatiotemporal resolution and minimal invasiveness. Naturally occurring photoreceptors and a plethora of engineered representatives now enable ver-satile optogenetic intervention into manifold cellular processes. That notwithstanding, there has been a dearth of optogenetic actuators that would directly act on RNA. We have recently closed this gap with the discovery of the light-oxygen-voltage (LOV) receptor PAL from the actinobacte-rium Nakamurella multipartita. PAL sequence-specifically binds short RNA aptamers with affinity that is enhanced by around 100-fold under blue light and thereby enables hitherto inaccessible optoribogenetic applications. The current, collaborative proposal aims at obtaining a molecular understanding of light regulation and RNA binding in PAL and two recently discovered homologs. Pertinent insight informs the engineering of PAL derivatives with altered and enhanced traits, par-ticularly RNA sequence specificity and response to light. We will harness the light-regulated PAL:aptamer interaction for the spatiotemporal control of micro RNA splicing and expression reg-ulation of endogenous genes, both in cell culture and in vivo. The planned research paves the way towards novel light-regulated actuators suitable for the precise optoribogenetic interrogation of the myriad roles RNA fulfills in nature.