Molecular Light Switch for Controlled Gene Expression

描述(由申请人提供)：基因表达的实验操作是一种广泛使用的技术，对许多现代生物医学研究至关重要。控制重组基因的时间和表达水平的能力是生产用于基础研究和生物药物合成的重组蛋白的核心。无论是在实验室培养中还是在整个动物中，控制哺乳动物细胞基因表达的商业系统都依赖于小分子药物的应用来诱导或抑制转录。这些系统受到几个限制，包括毒性、靶外效应、缺乏与时间和洗脱有关的精确度，以及诱导的可逆性。此外，效应性药物和必要的无药物介质的成本可能会很高。需要新的技术来克服这些限制。目前的项目旨在开发一种使用长波(700-750 nm)光来诱导基因表达控制的新系统。除了避免与药物系统相关的副作用和成本外，光诱导控制还提供了对单个组织和细胞进行精确的空间和时间靶向的优势。光还避免了小分子诱导剂固有的摄取和运输要求。我们提出的策略将利用一种对长波长光自然反应的光感受器类，并将其转化为 变成控制目标基因转录的“分子光开关”。所选择的光感受器也是自然光可逆的，使工程转录开关能够施加前所未有的转录控制水平。该系统的成功开发不仅有可能彻底改变哺乳动物细胞培养中可诱导的基因表达，而且有可能推动整个动物的转基因表达研究和新的遗传疗法的开发。

DESCRIPTION (provided by applicant): Experimental manipulation of gene expression is a widely used technique that is crucial to much of modern biomedical research. The ability to control the timing and expression level of recombinant genes is central to the production of recombinant protein for basic research as well as for the synthesis of biological pharmaceuticals. Commercially available systems for controlling gene expression in mammalian cells, whether in laboratory cultures or in whole animals, are dependent on the application of small molecule drugs to induce or repress transcription. These systems suffer from several limitations, including toxicity, off-target effects, lack of precision in relation to timing and loation, and reversibility of induction. Moreover, the cost of the effector drug and necessary drug-free media can be burdensome. New technologies are needed to overcome these limitations. The current project aims to develop a new system for inducible control of gene expression using long-wavelength (700-750 nm) light. In addition to avoiding the side effects and costs associated with drug-based systems, light-inducible control offers the advantages of precise spatial and temporal targeting of individual tissues and cells. Light also avoids the uptake and transport requirements inherent in small molecule-based inducers. Our proposed strategy will exploit a photoreceptor class that is naturally responsive to long-wavelength light, and convert it into a "molecular light switch" that controls transcription of target genes. The selected photoreceptor is also naturally photoreversible enabling the engineered transcriptional switch to exert an unprecedented level of transcriptional control. Successful development of this system has the potential not only to revolutionize inducible gene expression in mammalian cell culture, but is also likely to advance the study of transgenic expression in whole animals and the development of novel genetic therapies.