The ever-increasing complexity of synthetic gene networks and applications of synthetic biology requires precise and orthogonal gene expression systems. Of particular interest are systems responsive to light as they enable the control of gene expression dynamics with unprecedented resolution in space and time. While broadly used in mammalian backgrounds, however, optogenetic approaches in plant cells are still limited due to interference of the activating light with endogenous photoreceptors. Here, we describe the development of the first synthetic light-responsive system for the targeted control of gene expression in mammalian and plant cells that responds to the green range of the light spectrum in which plant photoreceptors have minimal activity. We first engineered a system based on the light-sensitive bacterial transcription factor CarH and its cognate DNA operator sequence CarO from Thermus thermophilus to control gene expression in mammalian cells. The system was functional in various mammalian cell lines, showing high induction (up to 350-fold) along with low leakiness, as well as high reversibility. We quantitatively described the systems characteristics by the development and experimental validation of a mathematical model. Finally, we transferred the system into A. thaliana protoplasts and demonstrated gene repression in response to green light. We expect that this system will provide new opportunities in applications based on synthetic gene networks and will open up perspectives for optogenetic studies in mammalian and plant cells.

中文翻译：

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用于控制哺乳动物和植物细胞中转基因表达的绿光响应系统

合成基因网络的日益复杂和合成生物学的应用要求精确和正交的基因表达系统。特别令人感兴趣的是对光敏感的系统，因为它们能够以空前的时空分辨率控制基因表达动态。然而，尽管在哺乳动物背景中广泛使用，但是由于活化光与内源性光感受器的干扰，植物细胞中的光遗传学方法仍然受到限制。在这里，我们描述了第一个合成的光响应系统的开发，该系统可对哺乳动物和植物细胞中的基因表达进行有针对性的控制，该系统对植物的感光细胞具有最小活性的光谱的绿色范围做出响应。嗜热栖热菌可控制哺乳动物细胞中的基因表达。该系统可在多种哺乳动物细胞系中发挥作用，显示出高诱导率（最多350倍），渗漏率低，可逆性高。我们通过数学模型的开发和实验验证来定量描述系统特性。最后，我们将系统转移到拟南芥原生质体中，并证明了对绿光的基因抑制。我们希望该系统将为基于合成基因网络的应用提供新的机会，并为哺乳动物和植物细胞的光遗传学研究开辟前景。